IN HEALTH AND DISEASE - CNPNProteome Dynamics in Health and Diseases 3 Contents Workshops 5 Short...

IN HEALTH AND DISEASE

Sixth AnnualCanadian National Proteomics Network Symposium

Hilton Bonaventure Hotel, Montreal

April 14-16, 2014

2 Sixth Annual Symposium of the Canadian National Proteomics Network

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Proteome Dynamics in Health and Diseases 3

Contents

Workshops 5

Short Program 6

Travel award recipients 9

CNPN distinguish award presentation & invited lecture 13

Session: 1. BIOMARKER DISCOVERY 19

Session: 2. INTERACTOME 23

Session: 3. POST TRANSLATIONAL MODIFICATIONS AND CELL SIGNALING 27

Session: 4. CHEMICAL AND STRUCTURAL PROTEOMICS 31

Session: 5. TECHNOLOGY DEVELOPMENT 35

Session: 6. BIOINFORMATICS 39

Poster 43

Floor layout Hilton Hotel 68

List of participants 69

5Proteome Dynamics in Health and Diseases

Workshops

Sunday Apri l 13

Workshops - Rooms Fontaine G, H

8:30 Registration

9:00 - 12:00 Multiple Reaction MonitoringState-of-the-art in MRM assaysLectures (Room Lasalle) Christoph Borchers, University of Victoria - GBC Proteomics Centre, BC Romain Simon, University of Victoria - GBC Proteomics Centre, BC Christine Miller, Agilent Technologies, Santa Clara, CA

9:00 - 12:00 Clinical Proteomics 1Combining affinity enrichment, targeted and data independent acquisition approaches for biomarker discoveryLectures (Rooms Verdun/Lachine)Joelle Pérusse, Institut de Recherche Clinique de Montréal, QC Jennifer Frahm, Thermo Fisher Scientific, San Jose, CA Brigitte Simmons, AB/Sciex, ON

12:30 Lunch - Room Outremont

13:00 - 17:00 Clinical Proteomics 2MS-based tissue-imaging: improving diagnostic and health careLectures (Rooms Verdun/Lachine)Pierre Chaurand, Université de Montréal Richard Drake, Medical University of South Carolina, SC Josephine Bunch, The National Physical Laboratory, UK Heath Patterson, Université de Montréal, QC Timothy Garrett, University of Florida, FL

15:00 Symposium Registration

18:00 - 20:00 CNPN / Desorption Symposium 2014Welcome Reception


Monday Apri l 14

8:00 Registration

8:00 - 18:00 Poster Pin Up

8:20 Benoit Coulombe WELCOME ADDRESS

Session 1: BIOMARKER DISCOVERY (Chair: Benoit Coulombe) Rooms Verdun/Lachine

8:30 Invited Speaker

Proteome Centric Molecular Profiling of Cancer Sam M Hanash, Director, MD Anderson Cancer Center, University of Texas

9:10

Invited SpeakerIntegrated omic analysis of lung cancer reveals proteome signatures with prognostic impact Mike Moran, University of Toronto, Hospital for Sick Children, OCI

9:50 Development and application of a highly sensitive MSIA-SRM assay for quantification of the hypercholesterolemia-associated protein PCSK9 in plasma Joelle Pérusse, Institut de recherches cliniques de Montréal

10:15 MORNING COFFEE BREAK


Targeted proteomics for validating plasma-based biomarkers for ovarian cancer Ruth Hüttenhain, UCSF, University of California

11:25 Systematic validation of protein biomarker signatures for the stratification of aggressive prostate cancers Yunee Kim, University of Toronto

11:45 Improved Mass Spectrometry Sensitivity of Multiplexed Plasma Protein Quantification through Limited High pH LC Fractionation Romain Simon, University of Victoria, Genome BC Proteomics Centre

12:05 Short talks (5 min each)

LC-MS/MS based method for the absolute quantitation of NAPQI-modified serum albumin from in vivo rat plasma samples: monitoring acetaminophen toxicity Andre LeBlanc, Université du Québec à Montréal

The HDL proteome in acute coronary syndromes shifts to an inflammatory profile Zuhair Awan, McGill University

Urinary Exosomal Proteomic Analysis Reveals Candidate Biomarkers of Rats with Unilateral Ureteral Obstruction Dennis Orton, Dalhousie University

Using a combination of proteomics, lipidomics, metabolomics and chemical genomics, for discovery of natural anti-aging and anti-cancer compounds and defining mechanisms of their action Vincent Richard, Concordia University

12:30 LUNCH / TECH TALK

12:30-13:30 Thermo Techtalk (Outremont): Dawn of a New Era in Proteome-wide Quantification Using 10-plex Isobaric TagsPatrick Murphy, Harvard Medical School

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Monday Apri l 14

Session 2: INTERACTOME (Chair; Andrew Emili) - Rooms Verdun/Lachine


Deciphering human disease mutations through the atomic-resolution protein interactome network Haiyuan Yu , Cornell University

14:25 Profiling the RNA Maturation Landscape in Yeast Marlene Oeffinger, IRCM - (20 min)

14:45 Short talk (5 - 10 min each)

Dynamics of protein interactions of the MCM complex in response to DNA damage. Francois-Michel Boisvert, Université de Sherbrooke

Mapping the architecture of a human cell by proximity biotinylation coupled to mass spectrometry Christopher Go, Lunenfeld-Tanenbaum Research Institute

Characterizing the Human Ubiquitin Conjugating Enzyme (E2) Interactome Deborah Ng, University of Toronto

15:05 Mitochondrial Yeast Protein-Protein Interactions Reveal Diverse Complexes and Disease-Relevant Functional Relationships Mohan Babu, University of Regina

15:25 COFFEE AT POSTERS (Even Numbers),- Room Fontaine E F G H


Protein-protein interaction data: All that glitters is not gold Shoshana Wodak, Hospital for Sick Children, Toronto

16:55 Proximity labelling and mass spectrometry reveals novel roles for receptor protein tyrosine phosphatases Nicole St-Denis, Lunenfeld-Tanenbaum Research Institut

17:15 CNPN Annual General Assembly - Room Fontaine E F G H

18:00 - 20:00 POSTER SESSION (Room Fontaine E F G H) + MIXER (Cash Bar)

How does environment impact human health and natural resources at the genomic level? How can genomics help leverage Canada’s traditional resource sectors while preserving the environment?

Explore these questions and more with world-renowned genomics experts and visionaries from academia, industry, government and media. A poster session highlighting the work of graduate students and post-doctoral fellows will also be featured.

Register today at www.powerandpromise.cvent.com/2014

Stay connected with us:

November 24–26, 2014Ottawa Convention Centre, Ottawa, Ontario

#powerofgenomics @GenomeCanada

PromisePower

the

Genomics:the

Travel award recipients Award appliedAdam Rabalski, Western University student

Andrew Crowell, Dalhousie University student

Megan Lewicki, University of Toronto student

Syed Shaw, Ryerson University student

WuYang Jin, University of British Columbia student

Yunee Kim, University of Toronto student

Mario Navarrette, University of Manitoba young investigator

Razvan Albu, University of British Columbia young investigator

Xiaodong Wang, University of Victoria young investigator

Amber Couzens, Lunenfeld-Tanenbaum Research Institute young investigator

Sponsored by:

Travel award recipients


Tuesday Apri l 15

8:00 Registration

Session 3: POST TRANSLATIONAL MODIFICATIONS AND CELL SIGNALING (Chair: Pierre Thibault) - Rooms Verdun/Lachine


Phosphoproteomics in Cell Signaling and Cancer Scott Gerber, Geisel School of Medicine at University of Dartmouth


Phosphoproteomic Analysis Identifies the Tumor Suppressor PDCD4 as a RSK Substrate Negatively Regulated by 14-3-3 Philippe Roux, IRIC, Université de Montreal

9:30 Proteomic dissection of adaptor protein signaling complexes Nicolas Bisson, Universite Laval

9:50 System-level adaptation to osmotic stress in yeast unravelled using time-resolved phosphoproteomics Evgeny Kanshin, IRIC, Université de Montreal



Modulated ubiquitination of mitochondrial proteins following depolarization and pro-apoptotic stimuli Don Kirkpatrick, Genentech

11:25 Proteomics insight of heat-shock induced ubiquitination Thibault Mayor, University of British Columbia

11:45 Losing a BET in the nucleolus - unexpected consequences of bromodomain inhibition Jean-Philippe Lambert, Lunenfeld-Tanenbaum Research Institute

12:05 Short talks (5 min each)

Site-specific quantitation of lysine acetylation in isomeric peptides of histones H3 & H4 Nebiyu Abshiru, IRIC, Université de Montréal

Time-resolved phosphoproteomics following heat and cold shock provides novel insights into the regulation of cytokinesis and DNA replication Peter Kubiniok, IRIC, Université de Montreal

Proteomic analysis of cap-dependent translation identifies LARP1 as a key regulator of 5’TOP mRNA translation Marie Cargnelo, IRIC, Université de Montréal

Characterizing the SUMO Stress Response (SSR) in S. cerevisiae Megan Lewicki, University Health Network, University of Toronto


12:30-13:30 AB/Sciex Techtalk (Outremont)

Uncovering Meaningful Information in SWATH Data in Less Time

Steven Tate, AB/SCIEX, Concord, ON

PUSHING THE LIMITS IN MASS SPECTROMETRY

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Tuesday Apri l 15

Session 4: CHEMICAL AND STRUCTURAL PROTEOMICS (Chair: Christoph Borchers) - Rooms Verdun/Lachine


MS-based approaches for the elucidation of nucleic acid higher-order structure Daniele Fabris, University of Albany, The RNA Institute


Using structural mass spectrometry to probe the mechanism of regulation of lipid signaling complexes John Burke, University of Victoria, Genome BC Proteomics Centre


Towards solving protein structures by structural proteomics Evgeniy (Jenya) Petrotchenko, University of Victoria, Genome BC Proteomics Centre

15:05 Characterization of HNE modification of matrix metalloproteinase 13 in osteoarthritis patients using high-resolution tandem mass spectrometry and multiple reaction monitoring Makan Golizeh, Université du Québec à Montréal

15:25 COFFEE AT POSTERS (Odd Numbers) - Room Fontaine E F G H


Metabolic pathway discovery in ocean microbes through community proteomics David Walsh, Concordia University

16:55 Interaction network mapping reveals novel kinase-phosphatase associations within the Hippo pathway Amber Couzens, Lunenfeld-Tanenbaum Research Institute

17:10 Structural Proteomic Analysis of SOD1 Aggregation Nicholas Brodie, University of Victoria

17:25 Using Isotopically-Coded N-terminal Modification and Non-Specific Proteinase K Digestion for the Identification of Zero-Length Crosslinks Jason Serpa, University of Victoria, Genome BC Proteomics Centre

17:40 CNPN Distinguished Scientist Award Presentation and Talk

Protein TAILS tell Remarkables Tales Chris Overall, University of British Columbia

18:45 Drinks - Informal Fluidic Discussion

19:30 Conference Banquet and Student Awards - Room Outremont


CNPN distinguish award presentation & invited lecture –Dr. Christopher Overall, recipient of CNPN – Tony Pawson Proteomics Award for Outstanding Contribution and Leadership to the Canadian Proteomics Community

We are pleased to announce the 2014 recipient of the CNPN – Tony Pawson Proteomics Award, Dr. Christopher Overall of the University of British Columbia. This prize recognizes the remarkable achievement on the fundamental understanding and/or practice of proteomics in biological sciences. Dr. Overall is an international leader in proteomics recognized �� the systems level investigation of protein turnover by proteolysis, the term he coined, and in developing ‘polymers for proteomics’. His focus has been on understanding the role of proteinases in vivo, particularly in cancer, infection �� (many in top-tier journals) that have been cited > 12,600 times, he also has numerous patents at various stages. He holds the Tier 1 Canada Research Chair in Metalloproteinase Proteomics and Systems Biology, and has won several national and international awards, including CIHR Researcher of the Year award (2012) and lifetime achievement awards from the International Proteolysis Society and the Matrix Biology Society of Australia and New Zealand and trained 38 HQP, 10 of whom hold professorships.

�� Protein TAILS tell remarkable talesProteolytic processing generates new protein species with characteristic neo-N termini that are frequently accompanied by altered half-lives, function, interactions and location. The extent and rate of isoform !�� developmental stage. Monitoring in vivo �� but also obtains proof for the expression of ‘missing proteins’ in the human proteome. Proteolysis alters the protein sequence and results in neo-N termini and hence novel semi-tryptic N-terminal peptides ��"��#��$�� !�� %&��ionization and fragmentation properties over their fully tryptic counterparts, rendering these peptides �� '��*++��5��7��as Termini Orientated Proteomics-Human Amino Terminome (TOP-HAT) we utilize our N-terminomics ��7��9�� '��:�� #��;9�':#<�� mature protein N-termini and neo-N-termini of proteins. The success of TAILS is due in part to ‘polymers for proteomics’ that we have developed. In TOP-HAT we are analyzing less commonly studied or accessible cells in order to identify rare cell-restricted or developmental-stage restricted expression of proteins and their speciation. Recently we have determined the N terminome of human erythrocytes, � �� =� ��including 6 missing human proteins in erythrocytes and 10 in platelets. By analysis of pathological tissue we are mapping proteolytic pathways and their irreversible actions on signalling and cytokine pathways ��7�� outside-to-inside the cell.

CNPN distinguish award presentation & invited lecture


Wednesday Apri l 16

8:00 Registration

Session 5: TECHNOLOGY DEVELOPMENT - CNPN / Desorption shared session (Chairs: Alan Doucette, Pierre Chaurand) - Rooms Verdun/Lachine


Bringing Label-Free Quantitation to Top Down Proteomics

Neil Kelleher, Northwestern University

9:10 Data Dependent and Independent Strategies for Identifying Urinary Biomarkers (Talk sponsored by AB Sciex) Hanno Steen, Boston Children’s Hospital

9:30 Platelet-monocyte interactions in atherosclerosis: combining proteomic approaches helps discover a novel mechanism Jurgen Kast, University of British Columbia

9:50 Enhancement of Dynamic Range for Quantitative Proteomics Using Isotopic Labeling and High Field Asymmetric Waveform Ion Mobility Spectrometry (FAIMS) Eric Bonneil, Université de Montreal



Integrating Proteomics and Metabolomics: technical challenges Liang Li , University of Alberta

11:25 Identification of radical intermediates by immuno-spin trapping and mass spectrometry: site-specific detection of radicals on Î±-lactalbumin after a riboflavin-sensitized reactionMathilde Triquigneaux, Concordia University

11:45 Immobilized enzyme microreactor development and optimization as evaluated by capillary electrophoretic peptide mapping

Golfam Ghafourifar, Universite de Montreal

12:05 Short talks (5 min each) Peptide-mediated degradation of a death-inducing kinase as a novel therapy for stroke Wu Yang Jin, University of British Columbia

An Automated Immuno-MALDI Assay for the Clinical Determination of Plasma Renin Activity Robert Popp, University of Victory, Genome BC Proteomics Centre

Enhancement of lipid sensitivity for negative-ion tissue imaging by FTICR-MS using quercetin as a MALDI matrix Xiaodong Wang, University of Victory, Genome BC Proteomics Centre

Low-Evaporation Additives for Printing of Antibody Microarrays using Silicon Quill Pins Veronique Laforte, McGill University


12:30 - 13:30 Bruker Techtalk (Outremont)

1) ESI-FTICR analysis of nucleic acids: beyond mapping and sequencing applications Dan Fabris, University of Albany.

2) Comprehensive intact protein sequencing by combining multi-modal dissociation strategies

Gary Kruppa, Bruker Daltonics

Innovation with IntegrityProteomics

To Unravel the Proteome,Take an Integrated Approach

Solving the complexity of the proteome is far more challenging than fi rst imagined. That is why Bruker offers a portfolio of integratedcomplementary technologies that together comprise a multidimensional toolbox optimized to unlock the proteome’s complexity. Bottom-up and top-down analyses, intact protein analysis as well as in-depth protein characterization come together to illuminate a more detailed picture of the proteome, complete with biological context and confi dence in the quality of your MS data.

Visit us at the Canadian National Proteomics Network Annual Conference, Booth 1

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�� MALDI TOF-TOF

�� ESI-ITMS

�� ESI-MALDI-FTMS

�� Bioinformatics


Wednesday Apri l 16

Session 6: BIOINFORMATICS (Chair: Anne-Claude Gingras) Rooms Verdun/Lachine


MSstats: an R package for statistical analysis of quantitative mass spectrometry-based proteomic experimentsOlga Vitek, Purdue University

14:25 Treatment of PCP-SILAC data for monitoring interactome rearrangementsLeonard Foster, University of British Columbia

14:45 PSEA-Quant: Protein set enrichment analysis on label-free and label-based protein quantification data Mathieu Lavallée-Adam, Scripps Research Institute

15:05 Short talks (5 minutes each)

EGADS! Efficient GPU-Accelerated Database Search Andy Kong, University of Michigan

Automatic Peptide Selection for Targeted Proteomics Experiments Yassene Mohammed, University of Victory, Genome BC Proteomics Centre

An integrated proteomic analysis pipeline combining multiple open-source spectral matching algorithms Aleksandr Ignatchenko, The University Health Network

Investigation of aging-related protein insolubility by a MS-based proteomics and bioin-formatics approach Razvan Florin Albu, University of British Columbia

15:25 AFTERNOON COFFEE BREAK


Computational tools for untargeted proteomics using data independent acquisition mass spectrometry Alexey Nesvizhskii, University of Michigan

16:55 PEAKS Q: New Software for Protein Label Free QuantificationBin Ma, University of Waterloo

17:15 Subzero Temperature Chromatography and Top Down Mass Spectrometry for Spatially Resolved Protein Hydrogen Exchange MeasurementsChristoph Borchers, Genome BC Proteomics Center

17:35 Meeting Closes


NOTES


�� Sam M. Hanash University of Texas Houston, TX, USA

ABSTRACT: The MD Anderson Cancer Center has embarked on an ambitious initiative, designated the Moon-shot program, intended to reduce mortality associated with cancers for which there are promising opportunities that span from improved risk assessment and early detection to life saving innovative therapies. Proteomics is a major component of the moonshot program with a goal to unleash the power of proteomics to support driving ap-plications consisting of development of blood based biomarkers for early detection and for predictive, prognostic �� ?�'��?��'�� '�"�� human specimens, mouse models of cancer and cancer cell lines to identify candidate biomarkers and potential targets for therapeutics. The progress made and the opportunities and challenges involved in generating and min-ing petabytes of proteomics data will be presented

�� Mike Moran University of Toronto, Hospital For Sick Children, OCI Toronto, ON, Canada

Contributing Authors: L Li, SickKids; Y Wei, SickKids; C To, OCI; CQ Zhu, OCI; P Taylor, SickKids; V Ignatchenko, OCI; A Ignatchenko, OCI; J Tong, SickKids; NA Pham, OCI; W Zhang, SickKids; N Yanagawa, OCI; M Li, OCI; M Pintilie, OCI; G Liu, OCI; L Muthuswamy, OICR; FA Shepherd, OCI; MS Tsao, OCI; T Kislinger, OCI; MF Moran, SickKids, OCI, University of Toronto.

ABSTRACT: The cancer phenotype is a product of a set of processes, each subject to selective pressure and prone to dysregulation in cancer, acting at each stage of the sequence continuum DNA>RNA>Protein. Non-small cell lung carcinoma (NSCLC) has the highest cancer mortality worldwide, and with outcomes that have not im-proved in decades. We constructed a genetic map in the form of an integrated omic array in which ~400,000 mea-sures of DNA gene copy number, mRNA expression, and mass spec-based protein abundance were integrated from patient-matched NSCLC primary tumours, patient-derived xenograft (PDXs), and normal lung. Dysregulated proteins were encoded throughout the genome including, but not limited to, examples that map into cancer-asso-��[$�� %�� -ant to the cancer phenotype. The ability of a primary tumour to engraft is the best known prognostic indicator of recurrence in early stage NSCLC (John et al. Clin Cancer Res, 2011). Clustering revealed proteome signatures especially highly recapitulated between matched primary and PDX tumours. Genes encoding signature proteins �� overall survival. Therefore, the integrated omic analysis indicates that selective pressure for proteome remodeling is a key component of tumorigenesis and/or maintenance of the cancer phenotype, and that proteome signatures ��

Monday April 14

9:10 Invited

Speaker

8:30 Invited

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�� ! ��"�� #$��Joelle Pérusse IRCM - Institut de Recherche Clinique de Montréal Montréal, PQ, Canada

Contributing Authors: Joelle R Pérusse, Institut de Recherches Cliniques de Montréal, 110 Avenue des Pins Ouest, \��]� ��+^��*�_�`{|��}��?�=��9��#��=��7��{��'��\��#��}��|��\�� [��}��\��`��#�?��=��9��#��Biomarkers Research Initiatives in Mass Spectrometry Center, 790 Memorial Drive, Cambridge, MA, 02139, USA; Mary ��:��&��9��#��=��7��{��'��\��#��}��|��\�� [��Cambridge, MA, 02139, USA; Benoit Coulombe,Institut de Recherches Cliniques de Montréal, 110 Avenue des Pins Ouest, Montréal, PQ, H2W 1R7, Canada.

ABSTRACT: Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a key player in the regulation of circulating low density lipoprotein cholesterol (LDL-C). Both the distinct forms observed in plasma and posttranslational mod-��;+9\�<�� "�� 7 ��:[:"}� � ��:'#��7�� 7�� +9\��9��address this issue, and given the complexity and wide dynamic range of the plasma proteome, we have developed and applied a MSIA-SRM (Mass Spectrometry ImmunoAssay-Selected Reaction Monitoring) assay to quantify PCSK9. The SRM assay was optimized using heavy isotope-labelled peptides. The best antibody was determined by performing the enrichment step using MSIA tips coupled to commercial antibodies and analyzing the product by SRM. We detect peptides spanning all protein domains and a phosphorylation site by SRM analysis of MSIA-pro-cessed plasma digests spiked with heavy peptides. The calibration curve shows a LOD of 6 ng/ml, LOQ of 25 ng/ml and linearity of our assay (R^2=0.931-0.965) over PCSK9’s clinical range. The pipeline’s reproducibility was �� ;��<��`�� &�� 5��7�� 5�� +}#��'�� -��\#'�"#{\�� +}#��+9\��

%��&��&��'��Ruth Hüttenhain Cellular and Molecular Pharmacology, UCSF, University of California San Francisco, CA, USA

Contributing Authors: Meena Choi, Department of Statistics, Purdue University, West Lafayette, IN; Martin Soste, Insti-��=��9*��#��&� ��?�$�� # ��7�� }��9*��#��&� ��?�}��"��}��[��#��+��_��:��'$?�Lukas Reiter, Biognosys AG, Zurich, Switzerland; Oliver Rinner, Biognosys AG, Zurich, Switzerland; Atul Sethi, Institute of \� �� #��=�� 9*��#��&� ��?�[�� [�� =��_��*�� [�-na-Farber/Harvard Cancer Institute, Boston, MA; Olga Vitek, Department of Statistics, Purdue University, West Lafayette, '$?��"\� ��[�� :��:��#��?�{�� '��\� �� #��=�� 9*��#��&� �� #��#��&� ��

ABSTRACT: 9��7�� ;�}<��-prove management and outcome of the disease. Protein biomarker research is currently hampered by the limited �� -markers in complex samples such as blood plasma. Here, we describe a selected reaction monitoring (SRM)-�� "�� 7�� }�_�� -oped tailored publicly accessible libraries of SRM assays for biomarker relevant proteins, such as evidence-based cancer-associated proteins as well as N-glycoproteins. Anticipating the generation of large-scale SRM datasets in biomarker validation efforts, we developed two data analysis tools, mProphet and SRMstats, intended for auto-��5�� analysis of SRM datasets.We demonstrated that the biomarker validation strategy based on the SRM assays in �� 7��

Session: 1. Biomarker DiscoveryB

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for OC. The biomarker detection strategy was guided by a discovery-driven proteomic effort to detect potential $"� ��7�� }��'��7�� #{\�� }��-�� 7�� abundance difference comparing the patient groups were used to model novel putative protein signatures for the ��}��

��&��'��Yunee Kim University of Toronto Toronto, ON, Canada

Contributing Authors: Yunee Kim, Department of Medical Biophysics, University of Toronto; Salvador Mejia, Princess Margaret Cancer Center; Vladimir Ignatchenko, Princess Margaret Cancer Center; Cindy Yao, Department of Medical Bio-��9��?�� $�� [��\�� \� �� } �=�� \�� #�� ?�{��:��:��9��}�� }��{��}��\�� #�� ?��Gramolini, Department of Physiology, University of Toronto; Dean Troyer, Department of Microbiology and Molecular Cell =�� \�� #�� ?�+�� =��'��}��{��'��=��"��-��+ ��?��#��[��\�� \� �� } �=�� \�� #�� ?�{��[��7��[��} ��\� �� +�� !�� 9��\�� South Carolina; Thomas Kislinger, Princess Margaret Cancer Center.

ABSTRACT: Current prostate cancer (PCa) prognostic factors stratify patients into risk groups, but are inaccu-rate in predicting outcome, resulting in over-treatment of many men with indolent disease. Furthermore, multiple biopsies are required, subjecting patients to associated risks. Therefore, a pressing need in PCa management is improved prognostic factors that enable follow-up of men with low-risk disease without repeat needle biopsies. To �� !� ��"�� "��!�� 7��!��;�+#<�� +}��7�� "�+#��-gressive and indolent PCa revealed differentially expressed proteins based on spectral counts [Kim, MCP 2012]. {�� `�|�� ;��<�� SRM-MS. For each candidate, a corresponding crude heavy isotope-labelled peptide was synthesized and used ��#{\"\#�� $!�� -�� +#"��#'["#{\"\#��`��peptides, and a diagnostic signature consisting of 27 peptides, were generated using machine learning algorithms. Future experiments will be dedicated to developing a single standardized and multiplexed SRM-MS assay used �� +#"�� +#��7��^��9��5�� "�� +}�� -�� "��

�� (��)��*��+��+�*�-��Romain Simon University of Victoria, Genome BC Proteomics Centre Victoria, BC, Canada

Contributing Authors: Andrew J. Percy; Andrew G. Chambers; Christoph H. Borchers.

ABSTRACT: In the last few years, mass spectrometry has become a promising tool for protein quantitation by adding stable isotope-labeled standard (SIS) peptides to the digested matrix, separating the analytes by liquid chromatography (LC), and analysing the peptides by multiple reaction monitoring mode/mass spectrometry (MRM/\#<��'�� !�� :}"\{\%\#��

11:25

11:45

Monday April 14


�� 7��9��an approach, however, remains limited in blood plasma, especially at levels below 5 ng/mL, without antibody en-��"�� *��!��\{\��#'#��quantitative approach through an off-line, two dimensional reversed phase liquid chromatography (2D-RPLC) set-�� &�� *� ��"��9�� `�� "�� ;�� <� ��`��:}� ��9��concentrations covered an 8 order-of-magnitude range, from 15 mg/mL (vitamin D-binding protein) to 450 pg/mL (protein S100-B), which is more than an order-of-magnitude deeper than that attained without fractionation. The �� `� ��"��;��`��%:<�� `��"�� ̀ [�� 9�� evaluation of our multidimensional LC-MRM/MS method, along with its demonstrated improvement over an anti-body- and fractionation-free approach.

Short talks (Abstract in poster section)

*� �. ��&��&��"��/��)��&��0��(��Andre LeBlanc (poster #9) Université du Québec à Montréal (UQAM) Montréal, PQ, Canada

%��+�*��1��Zuhier Awan (poster #7) McGill University Montréal, PQ, Canada

2��3(��!��4��'��!�� 2��2��5&��Dennis Orton (poster #3) Dalhousie University Halifax, NS, Canada

2��&��6��6��&��6 ��Vincent Richard (poster #2) Concordia University Montréal, PQ, Canada

12:05

Session: 1. Biomarker DiscoveryB

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��'Haiyuan Yu Department of Biological Statistics & Computational Biology, Cornell University Ithaca, NY, USA

ABSTRACT: To better understand the molecular mechanisms and genetic basis of human disease, we combined the massive scale of network systems biology with the supreme resolution of traditional structural biology to ��"�� "��7�� "��_�� "��contradicting the previous assumption that only a few interface residues are mutation hot spots for disease. We �� |��"�� widely-accepted “guilt-by-association” principle does not apply to dominant mutations. Furthermore, recessive truncating mutations on the same interface are much more likely to cause the same disease, even if they are close ��$"�� protein products.

��!/�� *��7�� IRCM - Institut de Recherche Clinique de Montréal Montréal, PQ, Canada

Contributing Authors:�{��'��#��=�� #�� _�?�_��'�� Montréal, Montréal, PQ; Fridy P, Rockefeller University, New York, NY; Rogers R, Institute for Systems Biology, Seattle, WA; Kutlu B, Institute for Systems Biology, Seattle, WA; Saleem R, Institute for Systems Biology, Seattle, WA; Rout MP, Rocke-� ��$��7��$�?��'��#��=�� #�� _�?��\��'��cliniques de Montréal, Montréal, PQ.

ABSTRACT: '�� {$�� ?�they are not only the conveyer of genetic information in the form of mRNA, but can themselves be regulators of gene expression and other important cellular pathways. But regardless of which type, all RNAs are transcribed �� ;{$+<�� !�� _�� {$+��7��{$��+� �''�transcribed RNAs alone, and the, in comparison, relatively small number of known RNA maturation factors, several open questions remain; how much overlap between subsets of proteins exists for different Pol II transcripts? Are all mRNAs processed along the same pathway, or are there distinctions for different classes of transcripts? What is the intra-complex landscape, the direct interaction, within these RNPs?To obtain a comprehensive view on mRNP ��+��{$�� {$��7��mRNA maturation factors, from capping, splicing, 3’end processing and quality control to mRNA export. Our results indicate that while there is a certain degree of overlap, not all processing factors are associated with the same {$��\�� 7� ��interactions of an mRNP along its export pathway, using targeted crosslinking AP-MS. Taken together, our data provides a comprehensive overview of the vast RNA maturation landscape.

Monday April 14

14:25

13:45 Invited

Speaker



�� (��/��8Francois-Michel Boisvert (poster #21) Université de Sherbrooke Sherbrooke, PQ, Canada

��&��(��&�� Christopher Go (poster #19) Samuel Lunenfeld Research Institute, Mt. Sinai Hospital Toronto, ON, Canada

��9��+��2&�"��:��39��;3<=�� Deborah Ng (poster #23) University of Toronto Toronto, ON, Canada

��7��!��(�� !��-��!��Mohan Babu University of Regina Regina, SK, Canada

Contributing Authors: Ke Jin1,2; JamesVlasblom1; Matthew Jessulat1; Viktor Dieneko1, Sadhna Phanse1, Hiroyuki ��7�`��\��`�� \��=��`£�;+��<�� ¤�`[��=��{��Innovation Centre, University of Regina, SK, Canada; 2 Terrence Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, ON, Canada

ABSTRACT: Nuclear-encoded mitochondrial proteins (MPs) are widely conserved across eukaryotes, from unicel-lular organisms such as the budding yeast, to humans. While detailed mechanistic investigations of mitochondrial biosynthetic genes, complexes, and pathways have been performed extensively in isolation, an understanding of how these MPs function in coordination with other organelles remains unclear. Consequently, many questions regarding the role of mitochondrial dysfunction in the development of human disease remain unanswered. We �� !�� `��!�� \+��identifying hundreds of putative heteromeric MP complexes having extensive associations within mitochondria and ��!��"�� "��_�� !�� -ysis, morphological comparisons of deletion strains, and protein co-immunoprecipitation. Integration of these MP complexes with reported genetic interaction (GI) data revealed substantial crosstalk among MPs and non-MPs, not previously discernible from either physical or GIs alone, identifying novel factors critical for endoplasmic retic-ulum-mitochondrial organization, membrane morphology, and mitochondrial lipid transport. As more than one-third ��\+�� !�� 7�� new candidate disease-genes with putative roles in malignancies or neurodevelopment abnormalities. By taking a systematic, integrated look at inter- and extra-mitochondrial protein function, our analysis predicted and validated novel, disease-relevant mitochondrial gene functions.

Session: 2. INTERACTOMEIN

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��0��Shoshana J. Wodak((1)

Vlaams Instituut voor Biotechnologie, Department of Structural Biology Ghent, Belgium

Contributing Authors: James Vlasblom(1), Andrei Tourinsky(1), Shuye Pu(1) (1) Molecular Structure and Function program, Hospital for Sick Children, Toronto, Canada; (2) Department of Molecular Genetics, University of Toronto, Canada; (3) Depart-ment of Biochemistry, University of Toronto, Canada, (4) VIB Department of Structural Biology, VUB, Brussels Belgium.

ABSTRACT: Major progress has been achieved in the experimental techniques used for the detection of pro-tein interactions and in processing and analyzing the vast amount of data that they generate. However, we still �� methode. We will present an overview of major PPI datasets produced by high-throughput experimental meth-ods over the last 10 years. We will highlight the biases associated with different methods and their associated �� \��-cally, evidence will be presented that the poor overlap between the PPI datasets derived by different methods �� &�� "��-�� #�� ?�� 7- �� Going forward therefore, highest priority should be given to experimental and computational approaches capable of identifying the functional portions of the interactome as well as characterizing its non-functional complement.

Protein-protein interaction networks: the puzzling riches. Wodak SJ, Vlasblom J, Turinsky AL, Pu S. Curr Opin Struct Biol. 2013 Dec;23(6):941-53.

��(��&��Nicole St-Denis Lunenfeld-Tanenbaum Research Institute Toronto, ON, Canada

Contributing Authors: Zhen Yuan Lin, Lunenfeld-Tanenbaum Research Institute; Laurence Pelletier, Lunenfeld-Tanen-baum Research Institute; Anne-Claude Gingras, Lunenfeld-Tanenbaum Research Institute.

ABSTRACT: Regulated tyrosine phosphorylation plays a crucial role in controlling several important biological de-cisions in the cell, including proliferation, differentiation, adhesion, and cell death. At the plasma membrane—the initial interface with which the cell receives signals from its environment—tyrosine phosphorylation is regulated in large part by the balanced activities of two families of integral membrane enzymes—receptor tyrosine kinas-es (RTKs) and receptor protein tyrosine phosphatases (PTPRs). Abnormalities in tyrosine phosphorylation that disrupt proper cellular signalling are responsible for the development and progression of several human diseas-es, including cancer. Indeed, both RTKs and PTPRs are widely mutated in various forms of cancer. However, while several RTKs have been widely studied (and consequently exploited for therapeutic potential), the biological functions of the majority of PTPRs remain unknown.To gain insight into the cellular roles of various PTPRs, we �� ;�+"\#<��+9+{��However, in our standard AP-MS system, we were able to identify only a few interactors for each phosphatase. To further expand our network, we have since performed proximity biotinylation and mass spectroscopy (BioID-MS). The results demonstrate that BioID-MS can be quite useful for the characterization of proteins that are not ideal for �+"\#�� !�� =�-oID-MS results also highlight the diversity of environments in which PTPRs reside and function, and tremendously expand our understanding of the functional roles of this important family of enzymes.

16:15

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��Scott Gerber Geisel School of Medicine at Dartmouth Lebanon, NH, USA

Contributing Authors: Arminja Kettenbach, Sierra Cullati, Devin Schweppe, Jeff Milloy.

ABSTRACT: {�� "�� -ly all cellular signal transduction processes, including cell proliferation and differentiation, cell cycle progression, and apoptosis. In the context of human health, many oncogenes regulate the activity and expression of protein kinases, or are kinases themselves; thus, knowledge regarding dysregulated kinase pathways in human cancers has the potential to reveal mechanistic underpinnings of cellular transformation and/or novel entry points for ther-apeutic intervention for cancer care. Here, we present a summary of our studies that aim to uncover basic cell signaling mechanisms through the large-scale analysis of protein phosphorylation, their potential connections to the initiation and maintenance of cancer and the technologies that we have developed and employed to date to further these goals.

��%��>��!�#��&��/��!��&��?>�@�@Philippe Roux IRIC - Université de Montréal Montréal, PQ, Canada

Contributing Authors: Jacob A Galan, University of Montréal; Kathryn M Geraghty, Harvard Medical School; Genevieve :��\��]� ?��\��]� ?��9��7&��\��]� ?��}� ��\��]� ?��{��7�� #�� \��?�=�5��9��7�� School of Medicine; Bryan A. Ballif, University of Vermont; John Blenis, Harvard Medical School; Pierre Thibault, University of Montréal; Philippe P. Roux, IRIC - University of Montréal;

ABSTRACT: The Ras/mitogen-activated protein kinase (MAPK) signaling cascade regulates various biological functions, including cell growth and proliferation. As such, this pathway is frequently deregulated in several types of cancer, including most cases of melanoma. RSK (p90 ribosomal S6 kinase) is a MAPK-activated protein kinase required for melanoma growth, but relatively little is known about its exact function and the nature of its substrates. *�� 7�� {#�� 9�� {#��-�� `�"�"��_��-ized the phospho-dependent 14-3-3 interactome in melanoma cells, and found that a large proportion of 14-3-3 binding proteins are also potential RSK substrates. Our results show that RSK phosphorylates the tumor sup-pressor PDCD4 (programmed cell death protein 4) on two serine residues that regulate its subcellular localization and interaction with 14-3-3 proteins. We found that 14-3-3 binding promotes PDCD4 degradation, suggesting an important role for RSK in the inactivation of PDCD4 in melanoma. In addition to this tumor suppressor, our results suggest the involvement of RSK in a vast array of unexplored biological functions.

��(��Nicolas Bisson CHU de Québec, Cancer Research Centre, Université Laval Québec, PQ, Canada

Contributing Authors: Francois Chartier, Cancer Research Centre, Université Laval, Québec, PQ; Kevin Jacquet, Cancer Research Centre, Université Laval, Québec, PQ; Alice Beigbeder, Cancer Research Centre, Université Laval, Québec, PQ; Nicolas Bisson, Cancer Research Centre, Université Laval, Québec, PQ.

Tuesday April 15

9:10 Invited

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8:30 Invited

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9:30


ABSTRACT: Signaling pathways are commonly organized via inducible protein-protein interactions. For example, signals from tyrosine kinase receptors (RTK) are often relayed via adaptor proteins, which serve as hubs to recruit �� *�� 7�� _�� +"#{\�;��"� ��<�� &��!��7�� !��{9�� _�� the Grb2 adaptor nucleates a remarkably diverse set of protein complexes, involved in multiple aspects of cellular function. The data obtained delineates the highly detailed composition/organization of individual Grb2 complexes. Our approach also delivers key information about the parameters that govern the non-random formation of well-de-�� !�� {9�� 9�� "�� !�� cellular functions and phenotypes.

��Evgeny Kanshin IRIC - Université de Montréal Montréal, PQ, Canada

Contributing Authors: Louis-Philippe Bergeron Sandoval, University of Montréal; Pierre Thibault, Université de Montréal.

ABSTRACT: {�� "�� *�� 5�� 7�� "��!�� 9�� of dynamic changes in protein phosphorylation is a promising approach to dissect signaling events and correlate interactions between kinases, phosphatases and their substrates. Here we investigated dynamics of protein phos-�� 7��#��}�� the adaptation period with 2 min temporal resolution. Additionally we compared responses to two different salt ��_��&�� !�� ̀ �� suggesting that the proportion of regulated sites decreases while promiscuous phosphorylation increases during ��7�� 7��"��"��optimized to react to rapid emergency response, but also give rise to promiscuous phosphorylation at longer time �� !�� 7 �� suggest multiple regulatory functions of osmoregulation in cell cycle and morphogenesis.

��&�"��9�� Don Kirkpatrick Senior Scientist, GENENTECH San Francisco, CA, USA

ABSTRACT: The proteome of a cell is continuously sculpted in response to physiological and pathogenic stimuli. ��&�� 7�� of ubiquitin from lysine residues of protein substrates. Mass spectrometry has emerged as an ideal system for ��&�� -nated substrate, the position of ubiquitin is marked by a diglycine remnant (i.e. K-GG) on the resultant peptide. �� &� �� "�� +��7��#+��-vealing a cluster of substrates on the outer mitochondrial membrane (OMM). Separately, initiation of cell death by ��"��;��'�+��<��7��;��\��+'�"7��<��

Session: 3. POST TRANSLATIONAL MODIFICATIONS AND CELL SIGNALINGPO

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a distinct array of mitochondrial ubiquitination including the cell death regulators cytochrome C and SMAC/Diablo. Importantly, these proteins residue within the mitochondria and would not be accessible to the ubiquitin machinery under normal physiological conditions. The inducible ubiquitination of OMM proteins upon depolarizing stimuli and mitochondrial modulators of cell death upon pro-apoptotic stimuli are consistent known alterations of the mitochon-drial membrane integrity under these two conditions, respectively. This talk will focus on implementation of mass spectrometry proteomics methodologies to investigate the modulation of mitochondrial proteins by the ubiquitin system.

��'��&�"��8Thibault Mayor University of British Columbia Vancouver, BC, Canada

Contributing Authors: Fang Nancy, Chan Gerard, Zhu Mang, Comyn Sophie, Gsponer Joerg and Mayor Thibault.

ABSTRACT: Protein quality control pathways monitor the proteome to avoid the accumulation of misfolded pro-teins that are typically degraded by the ubiquitin proteasome system. Impairment of these pathways has been link to numerous neurodegenerative diseases like Parkinson’s and Huntington’s. Heat-shock induces an increase of both ubiquitination levels and proteasome-mediated degradation of misfolded proteins. Using quantitative mass spectrometry, we showed that heat-shock lead to the ubiquitination of mainly cytosolic proteins, including a large portion of intrinsically disordered polypeptides [1,2]. We recently found that the yeast Rsp5 ubiquitin ligase and its Nedd4 mammalian homologue were major ubiquitin ligases mediating this stress-response. We used again mass spectrometry to gain further insight into the range of Rsp5 substrates and the potential recognition mechanism of these heat-shock induced misfolded proteins. We found that a large portion of Rsp5 heat-shock induced sub-strates were cytosolic and contain PY Rsp5-interacting motifs that were more often located in structured regions. _� �� +�� {��"�-pendant ubiquitination upon heat-shock, and that a destabilizing point mutation near that PY motif led to further ubiquitination. We propose a model in which Rsp5 can directly target cytosolic misfolded proteins with PY motifs normally not exposed when folded.[1] Fang NN et al. (2011) Nat Cell Biol. [2] Ng AH et al. (2013) Mol. Cell. Prot.

*��43%��A��(��"��&��&��Jean-Philippe Lambert Lunenfeld-Tanenbaum Research Institute Toronto, ON, Canada

Contributing Authors:�\��7��9�� 7��:�� "9��{��'��?�#��+��$�� [��} �� \��#�� }��!��+�� #��7��$�� [��} �� \��#�� }��!��#��$�� [��} �� \��#�� }��!��+�� 7�� $�� [��} �� \��Structural Genomics Consortium, University of Oxford.Anne-Claude Gingras, Lunenfeld-Tanenbaum Research Institute.

ABSTRACT: :�� "�� highly enriched in the nucleus. Histone proteins in particular are heavily acetylated, which plays a crucial role in the regulation of gene expression by “loosening” chromatin, enabling DNA access to the transcription machin-ery. Histone acetylation is recognized by bromodomains, found in ~40 human proteins. Inhibitors targeting the ��%�� " ��=�9�� "��promise in cell-based models for numerous cancers as they decrease the expression of key oncogenes. However, the mechanism by which these inhibitors enact their effect remains unclear, preventing rational design of the next �� _��;�+<�� ;=��'[<�� ;\#<��=�9��;={[��={[��={[��={[9<��absence of bromodomain inhibitors. A consequence of inhibitor treatment was the almost complete collapse of ��7��=�9�� interaction platform on chromatin. However, BRD3 reacted unpredictably to one inhibitor, resulting in the formation

11:45

Tuesday April 15

11:25


�� 7 ��=��+"\#��=��'["\#��&��7 ��7��players in the regulation of ribosome biogenesis (mediated by the concerted action of all three RNA polymerases ;{$�+<<��+�� 7��by RNAPII, BRD3 may represent a novel regulatory link between RNAPI and RNAPII.


��"��+@�B�+>Nebiyu Abshiru (poster #31) IRIC- Université de Montréal Montréal, PQ, Canada

%��'��'��/��Peter Kubiniok (poster #25) IIRIC - Université de Montréal Montréal, PQ, Canada

��*�!�?��'��CD%5��!/��Marie Cargnello (poster #28) IRIC - Université de Montréal Montréal, PQ, Canada

��9��2 5��!��;��!=��8��Megan Lewicki (poster #32) University Health Network, University of Toronto Toronto, ON, Canada

Session: 3. POST TRANSLATIONAL MODIFICATIONS AND CELL SIGNALINGPO

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��&��Daniele Fabris Chemistry and Biological Sciences, University of Albany, The RNA Institute Albany, NY, USA

ABSTRACT: The discovery of ribozymes and riboswitches has keenly reasserted the critical role played by high-er-order structure in determining the function of nucleic acid sequences that do not code for actual proteins. Mass spectrometry-based approaches can provide valuable information about base-pairing and long-range interac-�� _�� strategies based on high-resolution and ion mobility spectrometry (IMS) mass spectrometry to investigate the structure-function relationships in non-coding viral RNA. We have demonstrated that the concerted application of footprinting and crosslinking methods can provide valid spatial constraints for modeling operations, leading to the solution of actual 3D structures. Top-down strategies can provide direct information about the position and strength of base-pairing interactions that stabilize higher-order structure. Putative structures are corroborated by IMS de-terminations that reveal the global topology of target RNA. The combination of antisense oligonucleotide probes and IMS analysis represents a very effective strategy for investigating the rearrangement of riboswitches into al-ternative conformations. These approaches constitute a valuable alternative for the investigation of systems that, �� &��!�� "�� structural biology. This is the case of the 5’ untranslated region (5’-UTR) of the genome of HIV-1 and its functional assemblies with the chaperone nucleocapsid (NC) protein. The results afforded by these approaches have been providing new insights into the processes of genome recognition, dimerization, and packaging, which will hopefully lead to the development of novel therapeutic strategies.

2��&�� (��John Burke University of Victoria, Genome BC Proteomics Centre Victoria, BC, Canada

Contributing Authors: Roger L. Williams, MRC Laboratory of Molecular Biology

ABSTRACT: Most cellular responses to extracellular stimuli have a common component of regulation arising from selective recruitment of a network of signalling complexes to membranes. However, studying the molec-� �� 7�� _��in understanding the phosphoinositide 3-kinase (PI3K) signalling pathway by applying a synthesis of hydrogen deuterium exchange mass spectrometry (HDX-MS), and X-ray crystallography. HDX-MS provides information on the conformation and solvent accessibility of protein amide hydrogens, and has allowed us to interrogate the dynamics of regulatory complexes of the class I PI3K isoforms, and how they interact with lipid membranes.Our application of this synthesis to PI3Ks elucidated novel aspects of PI3K regulation downstream of GPCRs and RTKs, as well as a unifying mechanism of how diverse oncogenic PI3K mutants upregulate activity. The impor-tance of regulating PI3K activity is highlighted by the fact that the PI3K p110Î± catalytic subunit (PIK3CA) is one of the most frequently mutated genes in cancer. Our HDX-MS results reveal novel details on the mechanism of activation of oncogenic mutations, and the molecular basis of how activating and inhibitory stimuli regulate PI3Ks. In addition I will also discuss our work using HDX-MS to optimise structural biology approaches, and future direc-tions examining the structural basis of the phosphoinositide 4 kinase family of enzymes in human disease.

14:25 Invited

Speaker

Tuesday April 15

13:45 Invited

Speaker


%��&��8Evgeniy Petrotchenko University of Victoria, Genome BC Proteomics Centre Victoria, BC, Canada

Contributing Authors:��+��7��"��=}�+��}��?�}��*��=��"��=}�Proteomics Centre.

ABSTRACT: Combination of the protein chemistry methods with modern mass spectrometry have crystallized �� interaction networks to the study of conformational changes of single proteins, can be addressed using multiple �� -tion which can be used as experimental constraints in protein-structure modeling. We believe that combination of numerous experimental structural proteomics constraints from multiple approaches will enable univocal deter-mination of single protein structures. Here, we present our recent developments in limited proteolysis, surface ��"��!�� 7�� ""� � � ��techniques -- for the solving unknown protein structures.

��9��+/3��(��?@��Makan Golizeh Université du Québec à Montréal (UQAM) Montréal, PQ, Canada

Contributing Authors:�¦\�7�� &��]��^�]��\��]� ��}��[��%+��Montréal, PQ, Canada][Jamilah Abusarah, Laboratoire de Recherche en Orthopédie, Hôpital du Sacré-Cœur de Montréal, PQ, Canada][Mohamed Benderdour, Laboratoire de Recherche en Orthopédie, Hôpital du Sacré-Cœur de Montréal, PQ, }��¨¦:7��# ��£��]��^�]��\��]� ��}��[��%+��\��]� ��+^��}��¨

ABSTRACT: Matrix metalloproteinase 13 (MMP-13) cleaves type II collagen to break down extracellular matrix ;�}\<�� !��&��7�� of joint degenerative diseases, such as osteoarthritis (OA). It is also observed that the lipid peroxidation product �"��!�"�";�<"�� ;*$�<��!�� mechanism of this interaction is not yet fully understood, it is suggested that this reactive electrophile potentially �� !��}\�� \\+�� !��{��\\+"`��©ª�*$��+�� digested by trypsin and analyzed by UHPLC-QqTOF-MS/MS. Data processing using ProteinPilot™ and Metabo- ��+� ��¬��7�� *$��\\+"`�� "�� -�� *$��!��-��!��*$�� :}"\{\�� !�� ;'+<��\�� &��*$�� \\+"`��*$�� 9�� \\+"`�%*$��

14:45 Invited

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15:05

Session: 4. CHEMICAL AND STRUCTURAL PROTEOMICSC

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��&��&��David Walsh Concordia University Montréal, PQ, Canada

Contributing Authors: William Li, Bedford Institute of Oceanography

ABSTRACT: The oceans support diverse microbial communities that play essential roles in energy and nutrient cycling. Genomics and metagenomics (genomics of natural assemblages of microbes) has uncovered many novel metabolic capabilities such organic carbon compound specialization, phototrophy, and the use of many ��;�� <��*�� information on metabolic potential of an organism and no information on whether metabolic pathways are active in the ocean or under what environmental conditions different metabolic pathways are operational. Analogous to metagenomics, metaproteomics is the study of the whole proteome of a microbial community and provides a direct assessment of in situ protein expression. Here we employed MS/MS-based metaproteomics to investigate the metabolic activities of microbes in the ocean. Through comparative metaproteomics we revealed the spatio-temporal structure of numerous metabolic activities in the ocean central to carbon, nitrogen, and sulfur cycling in the sea. By combining metaproteomics and metagenomics, we aim to identify interactions between microbial �� climate.

��'��'�� +��Amber Couzens Lunenfeld-Tanenbaum Research Institute Toronto, ON, Canada

Contributing Authors: Amber L. Couzens, James D. R. Knight, Dan Mao, Michelle J. Kean, Guoci Teo, Alexander Weiss, Wade H. Dunham, Zhen-Yuan Lin, Richard D. Bagshaw, Frank Sicheri, Tony Pawson, Jeffrey L. Wrana, Hyungwon Choi and Anne-Claude Gingras.

ABSTRACT: 9��*�� [�� 7��-way components led to tumor or overgrowth phenotypes. This signaling pathway is now also known to regulate cell growth and fate decisions in mammals and not surprisingly is dysregulated in many human cancers. Mech-anistically, Hippo signaling functions to sequester the transcriptional co-activators YAP1 and TAZ in the cytosol through creation of a 14-3-3 binding site or through creation of a phosphodegron leading to proteasome mediated degradation. In the absence of Hippo signaling, YAP1/TAZ enter the nucleus where they interact with transcrip-��;9��[`"�<� �� 7��acid, an inhibitor of the PP2A-like phosphatases, is known to activate the Hippo pathway. Here we present our ��7��"�� ;\#<��okadaic acid. This network is complemented by proximity-based biotinylation coupled to MS that allows detection �� 9�� |�� 599 novel interactions (Couzens et al. 2013 Science Signaling). Included in the novel interactions were several phosphatase complexes, notably association of PP6 and MST1 with MOB1 and MST1 with STRIPAK. Phosphory- �� *�� ;��proteins MOB1 and SLMAP) that mediate these phosphorylation-dependent interactions. Our current focus is on the mechanistic and functional basis of these associations.

16:15

16:55

Tuesday April 15


��5�?��Nicholas Brodie University of Victoria Victoria, BC, Canada

Contributing Authors:��+��7��=}�+��}��?�}��=��"��BC Proteomics Centre.

ABSTRACT: In amyotrophic lateral sclerosis (ALS) misfolding of the protein SOD1 results in toxic aggregation in motor neurons, leading to their degeneration and to symptoms of the disease. The structural nature of this mis-�� $\{��"�� '�� and crosslinking were employed. Native SOD1 is a homodimer, each monomer containing two Î²-sheets oriented perpendicular to the dimer interface and two metal binding sites for Cu2+ and Zn2+ on the opposite side. Dif-�� 7�� }=[+#��`�\��[9�� `�� 7��of them differential, involving all 10 lysines as well as the N-terminus. All four of these were located in the loop region of the metal binding sites in SOD1, including the crosslink K4-K69, which suggests that this region may �� [�� $*#"��+}�#"12C314N/13C315N reagent yielded complete coverage of all 10 lysines as well as the N-terminus, Y109, and 9��:��`��!��#�[`��[9��`�� exposed, further indicating that loss of metal binding leads to opening of the loop region. T56 was less exposed ��[9��'�� ¯°��³�� ´�� -ing T56 and bringing K69 closer to K4.

2��/�� /��#��E��*��'�Jason Serpa University of Victoria, Genome BC Proteomics Centre Victoria, BC, Canada

Contributing Authors:��#��?��+��7�?�}��=��

ABSTRACT: Zero-length crosslinking combined with MS is a valuable method for studying protein structure and ��"�� [��&��" �� 7�� 7�� 9�� "��$"�� ;}��`��<��!� ��;+}�#"*�%[�<� ;+��7�� `�<�� 7�� $"�� [�� 9��;��"9{9�#9['�{�##{��[�:'$��{<�� -��;{$��#�� &�� <�� 7��&��" �� 7��[}��+'}�+��described previously. The PCAS reagent will modify N-termini as well as amino acids K, Y, S, and T. The treatment ��\�$*��*��#��9��$��"��-��&�� "��;��<�which may be present in standard tryptic digests. The NTM DXMSMS Match Software was developed in-house �� $"�� 7��9�� 7��based on precursor mass and the combined light and heavy MS/MS spectra. The method described here -- a com-��$"�� !��""�� ":��"�� &��" �� 7��

17:10

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4��*�&��-��)��%��Neil Kelleher Director, Proteomics Center of Excellence Northwestern University ��':��#�

ABSTRACT: With the prospect of resolving whole protein molecules into their myriad proteoforms on a proteomic scale, the question of their quantitative analysis in discovery mode comes to the fore. In this brief �� 7�� 9��$�� and stringent scoring of abundance changes of whole protein forms <30 kDa in a complex proteome. The input is ~100-300 micrograms of total protein for each biological replicate and the outputs are graphical displays �� ;�� technical and biological variation). A key part of the pipeline is the hierarchical linear model that is tailored to the original design of the study. We apply this new pipeline to measure the proteoform-level effects of deleting �� ;��<��#��`�� _9��¶�� H4 and both H2B isoforms. Ultimately, this approach to label-free top down proteomics in discovery mode is a critical technical advance for testing the hypothesis that whole proteoforms can link more tightly to complex phenotypes in cell and disease biology than do peptides created in shotgun proteomics.

��2��4��'��Hanno Steen Pathology (Speaker Sponsored by AB Sciex), Boston Children’s Hospital and Harvard Medical School Boston, MA, USA

CONTRIBUTING AUTHORS Saima Ahmed, Department of Pathology, Boston Children’s Hospital and Harvard Medical School, Boston, MA, USA;Vivek Kadiyala, Shadeah Suleiman, Linda S. Lee & Peter A. Banks, from Center for Pancreatic Disease, Brigham and Women’s Hospital and Harvard Medical School, Boston, MA, USA;Darwin L. Conwell, Ohio State University Wexner Medical Center, Columbus, OH, USA (and CPD, Boston).

ABSTRACT: The emerging concept of data-independent acquisition routines, such as SWATH Acquisition, holds great promise for quantitative proteomics in general and MS-based biomarker discovery in particular. Using urine specimens from chronic pancreatitis patients and appropriate controls, we explored different data dependent and ��7��The currently available diagnostic modalities for chronic pancreatitis are unsatisfying as they only detect mod-erate to advanced disease, at which point only symptomatic treatment is possible. Thus, we hypothesized that �� }+��thereby possibly offering a superior alternative to current CP diagnosis modalities. We used an optimized urine ��7�� ¸�� +��-pal Component Analyses of the different datasets show clear separation of the diseased and control patients, and also indicate that urine proteome analysis can stratify chronic pancreatitis patients based on the disease stage, which would be a major breakthrough in chronic pancreatitis diagnostics. Further in-depth analysis of the data dependent and independent datasets revealed interesting commonalities and differences pointing to the strengths and limitations of the different acquisition routines.

Wednesday April 16

8:30 Invited

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9:10


��0��&��Juergen Kast BC Proteomics Network, University of British Columbia Vancouver, BC, Canada

CONTRIBUTING AUTHORS ��*��=��}� ��?��"�� =��Columbia; Ru Li, University of British Columbia; Chengcheng Zhang, University of British Columbia.

ABSTRACT: Atherosclerosis is the leading cause of cardiovascular diseases, the most common cause of death globally. Platelet-monocyte aggregrates have been established as early markers of disease, and aggregate for-�� Platelet-monocyte aggregates are a result of platelet activation, which also causes platelets to aggregate and form �� 7�� _�� "� �� -cytes, and endothelial cells, and established a suite of quantitative proteomic approaches to study the mechanisms underlying these interactions. This included phosphoproteomics and an MRM-based activation assay for 14 small �9+�� "��!��and glycoproteomics to determine changes in glycoprotein expression, as well as several physiological assays. Our studies have revealed a novel mechanism leading to increased adhesion and migration of monocytes: activat-ed platelets release soluble factors and vesicles that stimulate monocytes also without physical contact. We have ��9+�� to production of reactive oxygen species in monocytes, cysteine oxidation of ~40 proteins, and up-regulation of 49 ��|��¹��#��novel contact-independent mechanism, and produced potential targets for future therapeutic intervention, high-lighting the power of proteomics for the investigation of disease-relevant processes.

3��!��)��2��*�&��+��-��F�� &��;-�� =Eric Bonneil IRIC-Université de Montréal Montréal, PQ, Canada

CONTRIBUTING AUTHORS: Michael Belford1, Pierre Thibault2, 1�9��#�� 2 IRIC-Université de Montréal, Montreal, QC Canada.

INTRODUCTION: Quantitative proteomics with isobaric chemical tag is commonly used to compare protein abundance across different experimental conditions. However, this method is limited in terms of precision and accuracy when quantitative measurements are performed on reporter ions arising from mixed precursors, a situation often encountered in samples of increased complexity. Here, we report on the use high-Field Asymetric waveform ion Spectrometry (FAIMS) to reduce the occurrence of mixed MS/MS spectra and improve quantitative proteomics experiments using isobaric labeling reagents

METHODS EMPLOYED: Human monocyte TCL tryptic digest were spiked in yeast a yeast tryptic digest at various amounts, 9\9"�� &��":}"\#%\#��:9^"�� '\#��9��6.5 mm-radius FAIMS inner electrode was replaced by a 7.75 mm electrode leaving a 1.25 mm analytical gap. This electrode �� `��$2 as a carrier gas. Optimal transmission of peptide ions in the FAIMS interface was �� ;}�<��|�:%��

ABSTRACT: LC-MS/MS analyses of yeast tryptic digests revealed that the occurrence of isobaric ions is prevalent and up to ��\#%\#��!��9��:}"��'\#"\#%\#�� !�� 9��7�� "�� ;��'\#¤��|`��`��"��'\#<��9�� !�� peptides by reducing the extent of mixed MS/MS spectra. We also performed a systematic study on yeast tryptic digests spiked ��`¤`��`¤`��|�� !��"��ratio measurements leading to lower compression of TMT reporter ion ratios. These results indicates that the use of FAIMS ��

NOVEL ASPECTS: LC-FAIMS-MS/MS on an Orbitrap mass spectrometer provides enhance accuracy for quantitative proteomics using isobaric tag labeling

KEYWORDS: isobaric tag labeling, differential ion mobility, FAIMS, nano-LC-MSMS, HCD

9:50

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�� &��0��Liang Li Department of Chemistry, University of Alberta ��=��}��

ABSTRACT:Combining the proteome and metabolome information of a biological system can provide a more ho-listic insight into the biological processes of the system. However, there are several technical challenges in trying to integrate the proteome and metabolome results. In this presentation, the current challenges and potential solutions �� 5�� comprehensive and quantitative metabolome data from a biological system. A high-performance isotope labeling �� biological systems will be described. Some selected applications of combining proteomics and metabolomics for biological studies will be presented.

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Mathilde Triquigneaux Concordia University, CBAMS Montréal, PQ, Canada

CONTRIBUTING AUTHORS [Trine Dalsgaard,Department of Food Science, Aarhus University, Tjele Denmark;] [Leesa [��:��#�� =�� $�� '�� *� ��#��%$'*��{��9�� +��7��#?¨¦��#��:��9�!�� +�� $�� '�� *� ��#��%$'*��{��9�� +��7��#?¨¦�� {�� :��9�!�� +�� $�� '��-mental Health Sciences/NIH, Research Triangle Park, US;][Grith Mortensen, Department of Food Science, Aarhus University, 95 ��[��7?¨¦{�� \��:��9�!�� +�� $�� '�� *� ��Sciences/NIH, Research Triangle Park, US;]

ABSTRACT: �� !��'��of radical intermediates formed during these processes requires appropriate analytical techniques due to their instability and low concentration levels. In the presence of a spin trap, radicals can be trapped to form a persistent derivates further detected as a tag of the radical reaction. The combination of the so-called spin trapping technique with immunochemical methods and LC-MS(/MS) is a powerful strategy to elucidate radical mechanisms. For instance, oxidative changes in milk proteins have been investigated after enzymatically catalyzed reactions, metal-catalyzed reactions, and photosensitized reactions which induced changes in protein structure, functionality, �� 9��7��!�� 7��¯ª" �� !��!��&��5��:}"\#;%\#<��'��!��¯ª" �� generation, as well as DMPO trapped radicals and secondary oxidation products. The approach of using immuno-spin trapping in combination with MS is a promising tool box for the characterization of free radical generation in food proteins in more complex food systems. Platelet-monocyte interactions in atherosclerosis: combining proteomic approaches helps discover a novel mechanism

Wednesday April 16

11:25

10:45 Invited

Speaker


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Golfam Ghafourifar Université de Montréal Montréal, PQ, Canada

CONTRIBUTING AUTHORS: Brian Fleury, University of Montréal; Antoine Fleitz,University of Montréal; Karen C. Waldron, Université de Montréal

ABSTRACT: ��&�� "�� &�� &�� &��¤��&�"��"�� _�are using chymotrypsin (CT) as enzyme and glutaraldehyde (GA) as crosslinking agent to immobilize CT. The polymeric, spongy form of crosslinked GA-CT provides an insoluble particle that can digest dissolved substrate ;��"��<�� &��&��;'\�{<�� column. The GA-CT particles were used to digest BSA as an example of a large folded protein that needs denatur-ation prior to digestion, and casein-FITC as an example of a small labelled substrate to test enzyme activity in the ��"��'\�{��"}9�� - �� ;}�<��=#�� !�� &��"��!�� [��}�� "��;:'�<��;\#<��Immobilization conditions were investigated for different buffers, pHs, and reaction times. Fluorescence microsco-��!��"}9�� &��'\�{��9��"}9�'\�{��


��'��'�Wu Yang Jin (poster #44) University of British Columbia Vancouver, BC, Canada

�� *�� !��Robert Popp (poster #45) University of Victory, Genome BC Proteomics Centre Victoria, BC, Canada

3��&��-%�!� �� "�� *��(Xiaodong Wang (poster #46) University of Victory, Genome BC Proteomics Centre Victoria, BC, Canada

*��3��&�� )��Veronique Laforte (poster #40) McGill University Montréal, PQ, Canada

12:05

11:45

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��0��!��'��"��&��(��Olga Vitek Purdue University West Lafayette, IN, USA

ABSTRACT: This talk will overview the statistical methods and functionality implemented in MSstats, an open-��{��7�� \#��!��with complex designs, such as comparisons of multiple groups or time course comparisons. It handles quantita-tive shotgun DDA (data-dependent acquisition) experiments, targeted SRM (selected reaction monitoring), and SWATH/DIA (data independent acquisition) experiments. It can be used in conjunction with label-free experimental ��7��7�� &�� \#��-alities for data processing and quality control, model-based statistical analysis (including testing proteins and pep-tides for differential abundance, or estimating protein abundance on a relative scale), and model-based calculation of a sample size for a future experiment. MSstats is available stand-alone, or via graphical user interface as an external tool in the software framework Skyline.

%��*��Leonard Foster University of British Columbia Vancouver, BC, Canada

Contributing Authors:�$�� #��?�:��

ABSTRACT: Over the last decade mass spectrometric technologies have become an integral tool in the study of protein-protein interactions. This integration stems from the ability of MS technologies to now enable the monitor-ing of systems at near proteome wide levels. This depth of analysis, unfortunately, results in a tsunami of data pre-�� %�� {��- �� !��&�!� �� -�� ;#�}"+}+"#':�}<�¦`¨��=�� we demonstrated that the deconvolution of proteins into individual Gaussian curves and hierarchal clustering �� 9�� #�}"+}+"#':�}�� !��7�� &��"��!�� 7�� {�� `��|��}�{�\��9�� !�� #�}"+}+"#':�}��9�� day on standard workstation.

Wednesday April 16

14:25

13:45 Invited

Speaker


��3��)��0��&��&��&�� "��Mathieu Lavallée-Adam The Scripps Research Institute La Jolla, CA, USA

Contributing Authors: Navin Rauniyar, The Scripps Research Institute; Daniel McClatchy, The Scripps Research Institute; John R. Yates III, The Scripps Research Institute.

ABSTRACT: The extraction of relevant biological information from large proteomics datasets is a great challenge. 9��5�� "�� }�� &��quantitative proteomics datasets. We propose to use a statistical approach to computationally identify protein sets ;�� ;��<� ��<� �� "�� -�� 9��+#��"^�� "�� "��+#��"^�� protein-protein abundance dependencies and literature annotation biases to yield biologically meaningful results. It offers an alternative approach to classic GO enrichment analyses and allows the analysis of samples originating �� 7�� #��+#�� _�� +#��"^�� -�� _�� +#��"^�� "�� !��}�9{�� +#��"^�� 7�� !�� biological insights.


3I��J��3��I�2��&��Andy Kong (poster #51) University of Michigan Ann Arbor, MI, USA

��%��3(��Yassene Mohammed (poster #52) Bioinformatics, University of Victoria, Genome BC Proteomics Centre Victoria, BC, Canada

��&�� Alexandr Ignatchenko (poster #48) The University Health Network Toronto, ON, Canada

��&��&�� &�� &��Dr. Razvan Florin Albu (poster #50) University of British Columbia Vancouver, BC, Canada

Session: 6. BIOINFORMATICSB

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��"�� Alexey L. Nesvizhskii University of Michigan Ann Arbor, Michigan

Contributing authors: Chih-Chiang Tsou1,2, Dmitry Avtonomov2, Brett Larsen3, Monika Tucholska3, Anne-Claude Gin-gras3,4,

1Department of Computational Medicine and Bioinformatics, University of Michigan, Ann Arbor, Michigan, USA.

2Department of Pathology, University of Michigan, Ann Arbor, Michigan, USA. 3Lunenfeld-Tanenbaum Research Institute, Toronto, Canada 4Department of Molecular Genetics, University of Toronto, Toronto, Canada

ABSTRACT: Improvements in the scanning rates and the accuracy of mass measurement achieved in the latest generation of mass spectrometers (MS) have enabled several practical implementations of the so called data independent acquisition (DIA) strategy. Recent examples of DIA approaches include SWATH-MS, which uses a fast scanning Q-TOF instrument to systematically fragment the entire useful mass range in increments of a few tens of Daltons. A targeted data analysis approach that requires pre-existing spectral libraries (generated using convention data dependent data, DDA) is then used to extract accurate quantitative information from #_�9*"\#��*��#_�9*"\#�� 7�� [[��_��open source software tool that extracts signal features from SWATH MS1 and MS2 data and assembles them into pseudo-MS/MS spectra that are fully compatible with conventional database search engines and error rate ��;��º�9��9��"+��+�� <��_�� #_�9*"\#�� !-ity. We discuss the advantages of the untargeted strategy, but also its drawbacks observed in the case of low abundance peptides in samples with a large dynamic range of protein abundances. We also discuss the differ-ences between our untargeted approach and the existing targeted data extraction approaches, and propose a ³��"��´�� ['��9�� #_�9*"\#�� ['��^��!��+ ��_��\#��

�3�#��)0�/��*�&��-��)��Bin Ma Computer Science, University of Waterloo Waterloo, ON, Canada

Contributing Authors: Lei Xin, Bioinformatics Solutions Inc; Hao Lin, Bioinformatics Solutions Inc; Baozhen Shan, Bioin-formatics Solutions Inc; Zefeng Zhang, Bioinformatics Solutions Inc; Weiwu Chen, Bioinformatics Solutions Inc; Mingjie Xie, Bioinformatics Solutions Inc;

ABSTRACT: +�� }�� !�� -��9�� '�� +��#�^�� 9��integrates the following steps for the data analysis:

7�� +��\�� "�� -tide features.

5�� Retention time alignment. A dynamic programming algorithm was developed to correct the LC retention time differences in different samples.

@�� Peptide feature matching.

8�� +��\#%\#��

16:55

16:15 Invited

Speaker

Wednesday April 16


B�� Ratio calculation. Peptide ratios are calculated from peak areas, and averaged up to obtain the protein ratio

;�� #��+"�� +"�� change is due to the difference in the samples, instead of from experimental errors and software arti-facts.

9�� !� ��!�� !��7��reported by the software were all true positives.

��&9�� %�� %�� !��+��3(�� Christoph Borchers University of Victoria, Genome BC Proteomics Centre Victoria, BC, Canada

Contributing Authors: Jingxi Pan, UVIC-Genome BC Proteomics Centre, Victoria, BC Canada;Suping Zhang, UVIC-Ge-nome BC Proteomics Centre, Victoria, BC, Canada.

ABSTRACT: Conformational dynamics of proteins are crucial for their functions such as enzyme catalysis, target recognition and activation, and protein-ligand interactions. Amide hydrogen/deuterium exchange (HDX), moni-tored by mass spectrometry (MS), is now a popular technique for measuring protein structural dynamics. Although the proteolysis-based HDX-MS approach is currently the method of choice, it gives only peptide-level information and suffers from a high level of back exchange. By fragmenting intact proteins directly using electron-based dis-sociation, the “top-down” HDX approach thus has several advantages. However, all of the top-down strategies reported to date have been exclusively direct infusion-based, and thus can only be used with volatile buffers. This has meant that the “top-down” approach could not be used for studying proteins under physiological conditions -- the very conditions which are often very important for preserving a protein’s native structure and function. There-�� "��*[�� -tein samples. In this presentation, we show how top-down electron capture dissociation and subzero temperature *+:}��9��7��7"!�� has no limitations in terms of protein type and sample solution conditions. Close to single-residue level protein structural information can be generated, with complete sequence coverage. In addition, the use of this technique for protein-metal ion interaction studies will also be discussed.

17:15

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Isabelle Kelly CHU de Québec, Centre Proteomique, Université Laval Québec, PQ, Canada

Contributing Authors: André J Tremblay, Institute on Nutrition and Functional Foods, Laval University; Sylvie Bourassa, Proteomics Centre, CHU de Quebec Research Center; Benjamin Nehmé, Proteomics Centre, CHU de Quebec Research Center; Benoît Lamarche, Institute on Nutrition and Functional Foods, Laval University; Amélie Charest, Institute on Nutrition and Functional Foods, Laval University; Marie-Claude Lépine, Institute on Nutrition and Functional Foods, Laval University; Patrick Couture, Institute on Nutrition and Functional Foods, Laval University & Lipid Research Centre, CHUL Research Centre, Laval University.

ABSTRACT: Dyslipidemia is an important risk factor for cardiovascular disease. Human in vivo kinetic studies of apolipoproteins are conducted to elucidate mechanisms underlying metabolic variations in plasma and impact of drug treatment. However, the current approach using stable isotope-labelled tracers and GC-MS requires highly �� '�� \{\��7-�� "�� '��7�� primed-constant infusion of L-[5,5,5-D3]leucine for 12 h with participants in a constantly fed state. Blood samples �� :[:�� digestion, one peptide per protein containing one leucine was selected. The native and the isotopically labelled forms were monitored by scheduled MRM. The isotopic ratios were calculated after peak area integration for each protein in function of time. Linearity of the signals was evaluated as well as the isotopic contribution of the native precursor. The correlation between measurements made by MRM and GC-MS for apoB48 and apoB-100 showed R2 values > 0.98. The optimized MRM method was used to investigate the effects of sitagliptin therapy (DPP-4 inhibitor) on the kinetics of apolipoproteins in patients with type 2 diabetes. The isotopic ratios were determined ��`��=��="`��"'��}"'''��-imum sample preparation.

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Vincent Richard Concordia University Montréal, PQ, Canada

Contributing Authors: Adam Beach, Concordia University; Anna Leonov, Concordia University; Amanda Piano, Con-cordia University; Rachel Feldman, Concordia University; Michelle T. Burstein, Concordia University; Anthony Arlia-Ci-ommo, Concordia University; Veronika Svistkova, Concordia University; and Vladimir I. Titorenko, Concordia University.

ABSTRACT: Caloric restriction (CR) extends lifespan across species and improves health by delaying the onset of age-related diseases. All currently known anti-aging drugs: (1) mimic life-extending and health-improving effects of CR without restricting caloric and nutrient intake; and (2) target signaling pathways that are under the stringent control of calorie and/or nutrient availability. It was believed therefore that all longevity pathways are “adaptable” by nature because they modulate longevity only in response to certain changes in the nutrient and energy status of an organism. However, it is possible that that some longevity pathways could be constitutive or “housekeeping” because they control longevity irrespective of calorie and/or nutrient availability. Our high-throughput chemical �� ;:}�<�� }{�� -ing such housekeeping pathways. Using proteomics, lipidomics, metabolomics and cell biological approaches, we found that LCA delays aging by: (1) remodeling lipid metabolism - thereby preventing liponecrotic cell death caused by fatty acid accumulation; (2) remodeling the repertoire of mitochondrial membrane lipids - thereby el-evating the abundance of mitochondrial respiratory supercomplexes and altering the age-related dynamics of changes in mitochondrial reactive oxygen species; and (3) attenuating mitochondrial fragmentation - thereby sup-��"�� "��LCA exhibits a potent and selective anti-tumor effect in cultured human neuroblastoma, glioma, breast cancer and prostate cancer cells by activating both the intrinsic and extrinsic pathways of apoptotic death.

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Dennis Orton Dalhousie University Halifax, NS, Canada

Contributing Authors: Orton, D.J.; Doucette, A.A.; Huang, W.F.; MacLellan, D.L.

ABSTRACT: Congenital urinary tract obstruction (UTO) is a commonly noted disorder on prenatal ultrasound that has the potential to lead to permanent loss of renal function. Current postnatal management requires lengthy and �� ^�� !��obtained from rat urine following complete (severe) or partial (mild) UTO in neonatal rats compared to controls ��7��9��}��!�� &�� %��;:}"\#<�� #�� `�� `��#�� `�� partially obstructed groups, 99 when comparing the control and the completely obstructed groups, and 83 proteins �� exosomes originate from cells lining the urinary tract, thus changes in exosomal protein abundance provided en-��7��"�� +�� 7��!�� &�� experiencing UTO. This study provides diagnostic and prognostic insight following mild and severe UTO. Further validation of these results may therefore improve upon existing postnatal management strategies for this disease.

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Prem Kumarathasan Analytical Biochemistry and Proteomics Lab, Health Canada Ottawa, ON, Canada

Contributing Authors:�$��\#�`��[��=�&��+�[`��}��\��"{��\#�`��[�� 5��\#�`�� -��\#��#��\�� +�[��9��+�[`��#��7��+�[`��{��+�[`��+��+�[��`'�� 9�!�� :��*#{=��*� ��}��$�� =��+��:��*#{=��*� ��}��$�

ABSTRACT: Inhalation of ambient particles is associated with episodic increases in cardiopulmonary morbidity and mortality. We aim to apply toxicoproteomic and toxicogenomic analyses in cell culture models to identify physicochemical characteristics of respirable particles that are related to biological reactivity, i.e. elucidation of potency determinants and adverse outcome pathways. The objective of our study was to verify the sensitivity of the proteome of human lung epithelial cells (A549) to the dose and to the physicochemical properties of particles. The cells were exposed for 24 hours to suspensions (0, 60, 140 and 200 Î¼g/cm2) of well characterized Ottawa ��+\`�� ;�*}"��*}�� <�� ;�*}�� <�� ;�*}�� <��reference particles TiO2 and SiO2. Cellular viability was determined (ATP, LDH, BrdU) and changes in cellular proteome were assessed by two-dimensional gel electrophoresis of whole cell extract, followed by MALDI-TOF-TOF-MS analysis of differentially expressed proteins. Gene expression was assessed by RT-PCR. Proteome and gene expression patterns included dose-dependent and particle-dependent changes, as well as dose x particle interactions (p<0.05, two-way ANOVA). Urban particles were relatively potent for activation of pathways related to cell necrosis and apoptosis, oxidative stress and antioxidant defense. A notable marker of urban particle toxicity ��"`" �7��;[$\`:<�� 9��response of A549 cells is sensitive to the dose and to the composition and physical aspect of the particles. Our study demonstrates that potency toward cellular pathways is related to the physicochemical properties of urban particles.

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Anja Rodenbrock Institute for Research in Immunology and Cancer, Université de Montréal Montréal, PQ, Canada

Contributing Authors: Granados, Diana-Paola1,2; Durette, Chantal1; Rodenbrock, Anja1; Caron-Lizotte, Olivier1; Côté, Caroline1; Laverdure, Jean-Philippe4; Lemieux, Sebastien1,5; Perreault, Claude1,2,6; and Thibault, Pierre1,3

1Institute for Research in Immunology and Cancer; Université de Montréal - Departments of: 2Medicine, 3Chemistry, 4Bioinfor-matics and Computer Science, and 5Operations Research: 6Division of Hematology, Hôpital Maisonneuve-Rosemont, Montréal, PQ, Canada.

ABSTRACT: *�� ;*}�<��`�� ;��<�� ;�*}9<��"��" �7��;��:<��an effective approach to eradicate leukemic cells. The curative effects of AHCT are mainly associated to immune cells that recognize or target tumor minor histocompatibility antigens (MiHAs), peptides presented by MHC-I mol-ecules and derived from polymorphic genomic regions that give rise to amino acid substitutions. Although approx-imately 200,000 subjects diagnosed with HCs have been cured with AHCT, the use of adoptive immunotherapy is relatively rudimentary and is hampered by the variable anti-HC activity of AHCT, and the risk of a devastating autoimmune response (graft vs. host disease, GVHD). The unpredictability of these two factors seriously limits the ��9��"*}�� -��"��\�*��9�� *�� \#"�� *:�"�£��¤�`"��\�*��=" ��-�� ;�� <��personalized protein databases using in silico translation of genome/transcriptome sequencing data. MHC-I pep-tides were isolated by mild acid elution followed by LC-MS/MS analysis. Peptides were searched against individual �� |�*:�"�£��¤�`"��\*}"'��\�*��9�� "�� "��-teomic analyses of MiHAs, opening up new avenues for leukemia immunotherapy.

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Mario Navarrete Manitoba Centre for Proteomics & Systems Biology Winnipeg, MB, Canada

Contributing Authors: Julie Ho, Manitoba Centre for Proteomics & Systems Biology, Department of Internal Medicine, Section of Biomedical Proteomics, University of Manitoba,Department of Internal Medicine, Section of Nephrology, University ��\��+��&&��\��}��+��»�#��=�� 7��\��}��+��-ics & Systems Biology, Department of Internal Medicine, Section of Biomedical Proteomics, University of Manitoba.John A Wilkins, Manitoba Centre for Proteomics & Systems Biology, Department of Internal Medicine, Section of Biomedical Pro-teomics, University of Manitoba.

ABSTRACT: Urine is a potential source of biomarkers for kidney-related diseases. However, analysis of the urine proteome is often hampered by high abundance proteins, many of which are in the higher molecular weight range. #�� 7�� 7�� -lecular weight suggesting that the selective removal of high molecular species (>50 kDa) could provide a method for enrichment of the former. In order to address this issue we examined the use of precipitation methods involv-ing the organic solvent acetonitrile as a readily applicable method for general sample processing. The optimal �� #[#�+�� 9�� "�and intra- individual level. Analysis on a Triple TOF 5600 mass spectrometer of the acetonitrile soluble proteins indicated that there was marked depletion of high molecular weight and high abundance protein species such as albumin and immunoglobulins. The MS/MS data revealed that the soluble fraction was enriched in lower molecular weight species; many of these proteins were not detected in either whole urine or the acetonitrile pellet fractions.

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These results suggest that the method may have general applicability and enrich low abundance species. We will present protocols and considerations for the standardized precipitation conditions for general application to the analysis of urine.marker discovery

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Zuhier Awan McGill University Montréal, PQ, Canada

Contributing Authors:�� [��=�� #��7�[��=�� '�� {� ��*��:��Krimbou, Sylvie Laboissiere, Jacques Genest.

ABSTRACT: =�}��{��$[¤�'��5�� ;�}#<��*[:�� }#��\�9*�[#¤�#��*[:�� "��control, stable CAD, and ACS subjects (n=10/group). HDL was isolated by ultracentrifugation and separated by `["� �� :}"\#%\#��{�#�:9#¤�_��|�*[:"�� -�� }�=�� -�� *[:"�� ;¸��<�� !�� "�� %��'�� "'�� }#�� ;#��<�� }��;}�<�� ;�� ?� �"��½¾¿��<�� :'#��[�� =}�`��=}�`��#{"='�� !�� *[:��}#�� "'��}�$}:�#'�$#¤�� -port that the HDL proteome differs between control, CAD and ACS patients. Increased abundance of SAA, C3, and ��*[:��}#��*[:�� which, in turn, might alter the protective effects of HDL on the atherosclerotic plaque.

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Pammy (Pik Shan) Lo Biomedical Engineering Department, McGill University Montréal, PQ, Canada

Contributing Authors:�*��:�?�#��=��?�+��:�?�[��7��+��=�� -neering Department, McGill University, Montréal, PQ, Canada.

ABSTRACT: Dynamic analysis of proteins in blood holds great potential for disease diagnosis and monitoring. Currently, experimental data on the changes of protein levels during the growth of tumors are lacking. Here, we ��"�� !��correlation with the tumor burden using an antibody colocalization microarray (ACM) on the snap chip. Human breast cancer cells with triple-negative phenotype were injected into 11 mice, along with three mice injected with Matrigel as controls. For 7 consecutive weeks, 15-40 μL of serum were collected from each mouse weekly, and the volume of the tumor was measured after injection of the cancer cells. 38 proteins detected in mice sera above the limits of detection of the assay were clustered in a heatmap. 10 proteins showed higher levels in the sera of mice injected with cancer cells than those in control mice, and among them the concentrations of 6 proteins (IL 8, TNF RI, uPA, uPAR, GCSF, and GM-CSF) increased continuously in direct correlation with the tumor volume. To validate that these proteins were indeed secreted from the tumor tissues in mice, we also measured tumor tissue lysates, cancer cell lysates and conditioned media. These results indicate that ACM can be used for dynamic analysis of multiple proteins in complex samples. The capability of measuring serial changes of proteins in animal models provides the foundation for tumor growth studies.

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Andre LeBlanc Université du Québec à Montréal (UQAM) Montréal, PQ, Canada

Contributing Authors: André Leblanc, UQAM; Tze Chieh Shiao, UQAM; René Roy, UQAM; Lekha Sleno, UQAM

ABSTRACT: Acetaminophen toxicity is the most common cause of acute liver failure in the United States. The hepatotoxicity of acetaminophen is caused by an electrophilic reactive metabolite of acetaminophen, N-acetyl p-benzoquinone imine or NAPQI, that forms covalent adducts with nucleophilic thiol groups in liver protein, causing ��#�� ;#�<��7�� $�+^'��-�� #�� 7�� $�+^'��*�� $�+^'��- �� '�� ;�� <�� -�� "�� 9��:}"\#%\#�� &��of all cysteine residues in standard albumin, yielding a surrogate peptide that is a positional isomer to the target ��$�+^'��'�� $�+^'"�� ;��!�� {#�<�� 7��levels of acetaminophen (from 75 mg/kg to 600 mg/kg) that includes low (non-toxic) doses of acetaminophen, showing the viability of this approach to directly monitor acetaminophen covalent binding from in vivo samples.

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Yunee Kim University of Toronto Toronto, ON, Canada

Contributing Authors: Yunee Kim, Department of Medical Biophysics, University of Toronto; Salvador Mejia, Princess Margaret Cancer Center; Vladimir Ignatchenko, Princess Margaret Cancer Center; Cindy Yao, Department of Medical Bio-��9��?�� $�� [��\�� \� �� } �=�� \�� #�� ?�{��:��:��9��}�� }��{��}��\�� #�� ?��Gramolini, Department of Physiology, University of Toronto; Dean Troyer, Department of Microbiology and Molecular Cell =�� \�� #�� ?�+�� =��'��}��{��'��=��"��+ ��?��#��[��\�� \� �� } �=�� \�� #�� ?�{��[��7��[��} ��\� �� +�� !�� 9��\�� #��}�� -na; Thomas Kislinger, Princess Margaret Cancer Center.

ABSTRACT: Current prostate cancer (PCa) prognostic factors stratify patients into risk groups, but are inaccu-rate in predicting outcome, resulting in over-treatment of many men with indolent disease. Furthermore, multiple biopsies are required, subjecting patients to associated risks. Therefore, a pressing need in PCa management is improved prognostic factors that enable follow-up of men with low-risk disease without repeat needle biopsies. To �� !� ��"�� "��!�� 7��!��;�+#<�� +}��7�� "�+#��-gressive and indolent PCa revealed differentially expressed proteins based on spectral counts [Kim, MCP 2012]. {�� `�|�� ;��<�� SRM-MS. For each candidate, a corresponding crude heavy isotope-labelled peptide was synthesized and used ��#{\"\#�� $!�� -�� +#"��#'["#{\"\#��`��peptides, and a diagnostic signature consisting of 27 peptides, were generated using machine learning algorithms. Future experiments will be dedicated to developing a single standardized and multiplexed SRM-MS assay used �� +#"�� +#��7��^��9��5�� "�� +}�� -�� "��

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Dylan Dieters-Castator Western University London, ON, Canada

Contributing Authors: [Dylan Dieters-Castator, University of Western Ontario; Gilles Lajoie, University of Western Ontario; Lynne-Marie Postovit, University of Alberta]

ABSTRACT: =��7��¤��!�� tumourigenesis in part through the recruitment of bone marrow populations including multipotent stromal cells (MSCs). Nodal, a cytokine and embryonic morphogen critical for development, has previously been shown to be a pivotal regulator of tumour vascularization in breast cancer, such that it is required for blood vessel recruitment and tumour growth in murine models and is positively correlated with high microvascular density in breast cancer patients. To better understand the role of Nodal- and MSC associated tumour vascularization, we investigated the effects of conditioned media (CM) derived from Nodal expressing breast cancer cells on MSC behaviours and characterized the Nodal-regulated breast cancer secretome by mass spectrometry. Results: MSCs were found to express Nodal, its receptor ALK4 and co-receptor Cripto by Western blotting although stimulation with recombinant ��$�� ;��$�� <�� #��{�`%��\#}��\�� rhNodal treatment. However, CM derived from Control (shC) compared to Nodal knockdown (shN) MDA-MB-231 �� \#}��#�� #[#"+��-��#':�}� �� }��$�\[�"\="��`�}\��`��;��in one or more biological replicates) with many implicated in breast cancer progression present. Select candidate ��!�� !�� _�� \#}��assayed.

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Miljan Kuljanin Western University London, ON, Canada

Contributing Authors: David Hess, Western University; Gilles A. Lajoie, Western University.

ABSTRACT: Diabetes is a metabolic disease that affects millions of people worldwide, and is projected to dras-tically increase in the up-coming years. Recently, multipotent Stromal cells (MSC) have surfaced as an attractive strategy to combat diabetes. MSC have the ability to differentiate into various cell types and more importantly have been shown to recruit to the injured pancreas and induce endogenous beta cell regeneration to improve islet ��*�� -ulation based on their aldehyde dehydrogenase activity (ALHD). Cells that retained high ALDH activity were grown �� 7��#':�}��¦`�}�`�$¨��¦`�}¨��lysine. The above approach was designed to answer two fundamental questions: 1) what factors are secreted by regenerative cell lines vs. non-regenerative cell lines, 2) what factors are being lost as the cells move through early to late passage. Fractionation of condition media using in-gel digestions had yielded over 1500 validated proteins, of which 770 are unique to each condition being evaluated. Regenerative cell lines have been shown to secret �� known secreted proteins or associated with extracellular matrix revealed some interesting results. These included: ��¯°"��7��-noic acid binding, SCD factors as well as proteins downstream of the Wnt pathway.

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François Chartier CHU de Québec, Cancer Research Centre, Université Laval Québec, PQ, Canada

Contributing Authors: François J.-M. Chartier Centre de Recherche sur le CancerCentre de Recherche du Centre Hos-pitalier Universitaire (CHU) de Québec; Nicolas Bisson, Centre de Recherche sur le Cancer Centre de Recherche du Centre Hospitalier Universitaire (CHU) de Québec Département de Biologie Moléculaire, Biochimie Médicale et Pathologie, Universi-té Laval

ABSTRACT: Adaptor proteins serve as links between transmembrane receptors and signal transduction path-��9�� !�� _�� adaptor protein GRB2, despite having only 3 protein interaction domains, namely Src-Homology (SH) 2 and 3, interacts with an extensive array of proteins partners. This suggested that many distinct GRB2-containing com-plexes are assembled upon tyrosine kinase receptor (RTK) signaling. We sought to delineate the composition of ��!��{=�� "��;�+"\#<��GRB2-associated proteins as baits. In addition, we combined siRNA knockdown and successive AP-MS to better characterize the relationships within the GRB2 network. Finally, we performed size exclusion chromatography fol-lowing GRB2 AP to resolve individual GRB2-centric complexes using MS. Taken together, our data provide novel �� {9�� the assembly of large protein complexes.

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Andree-Anne Lacombe IRCM - Institut de Recherche Clinique de Montréal Montréal, PQ, Canada

Contributing Authors: Diane Forget, IRCM; Mathieu Lavallée-Adam, McGill Centre for Bioinformatics and School of Computer Science, McGill University; Philippe Cloutier, IRCM; Annie Bouchard, IRCM; Racha Al-Khoury, IRCM; Celia Jeron-imo, IRCM; Denis Faubert, IRCM; Jacques Archambault, IRCM; Mathieu Blanchette, McGill Centre for Bioinformatics and School of Computer Science, McGill University; Benoit Coulombe, IRCM.

ABSTRACT: RNA polymerase II (RNAPII), the 12-subunit enzyme that synthesizes all mRNAs and several non-coding RNAs in eukaryotes, plays a central role in cell function. Although multiple proteins are known to reg-ulate the activity of RNAPII during transcription, little is known about the machinery that controls the fate of the enzyme before or after transcription.The RNA polymerase II (RNAP II)-associated proteins (RPAP) 2 and RPAP4 ��{$�+�''�� mass spectrometry experiments. Here, we show that RPAP2 and RPAP4 are mainly cytoplasmic proteins that shuttle between the cytoplasm and the nucleus. This shuttling is tightly coupled with nuclear import of RNAP II, as RPAP2 and RPAP4 silencing provokes abnormal accumulation of RNAP II in the cytoplasmic space. Most notably, RPAP4 silencing provokes the retention of RPAP2 in the nucleus. Binding of RNAP II to RPAP2 is mediated by an N-terminal domain that contains a nuclear retention domain and binding of RPAP4 to RPAP2 occurs through a C-terminal domain that has a dominant cytoplasmic localization domain. Our results support a model in which RPAP2 enters the nucleus in association with RNAP II and returns to the cytoplasm in association with the GTPase RPAP4.

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Jacob Galan IRIC - Université de Montréal Montréal, PQ, Canada

Contributing Authors:�� '{'}"��]��\��]� ?�}�� :��?�� =�� ?��=�� ?�+��9�� #]��}��]��?��?�+�� +��{��!?��;� ��<�'��{��'�� Cancer (IRIC), Université de Montréal, Montréal, PQ, Canada.

ABSTRACT: 9��`�"�"�� !��-works of molecular interactions affecting various cellular processes such as cell growth, cell cycle progression, apoptosis and differentiation. Indeed, identifying these conserved 14-3-3 interactions across species (human and ��<�� -mental biological processes. While there are various bioinformatics tools available to study these interactions, a complete list using experimental approaches is greatly warranted. In this study, we used mass spectrometry-based �� `�"�"�� _� �� Rab11-interacting protein Rip11 and its human ortholog FIP5 as novel phospho-dependent 14-3-3 client proteins. _�� {��``��'+�� `�"�"�� "�� involved in 14-3-3 binding. Interestingly, we found that depletion of Rip11 in S2 cells resulted in a high frequency of failed cytokinesis, and a mutant Rip11 unable to bind 14-3-3 could not rescue this phenotype, suggesting a functional interplay between Rip11 and 14-3-3 during cytokinesis. Taken together, this novel and conserved Rip11/FIP5 interaction with 14-3-3 highlights the necessity for more protein-protein interaction screens across different species and deepens our understanding of the role of 14-3-3 during cell division.

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Wade Dunham Lunenfeld-Tanenbaum Research Institute Toronto, ON, Canada

Contributing Authors: Tamiko Nishimura; Marc Fabian; Anne-Claude Gingras.

ABSTRACT: ��7��;<{$�� 3’polyadenylation, and export to the cytosol for translation. Post-transcriptional regulation of mRNA populations is equally complex and can involve inhibition of mRNA translation, and initiation of mRNA deadenylation, decap-ping and decay. Many of the enzymes and co-factors that ‘silence’ mRNAs exist in macroscopic structures known as RNA granules (e.g. stress granules, SGs) and bodies (e.g. “processing” or P-bodies, PBs). Our knowledge of mRNA regulation in these cytoplasmic foci has been hampered by a poor understanding of their protein compo-�� *��conducted a proteomic analysis of these bodies using a recently-introduced in vivo biotinylation strategy, BioID. '��=��'[��;�� "��&��#��+=��<�� biotin ligase (BirA*), and expressed within a cell line to label the local protein environment of the BirA*-fusion with ��} � �� of neighboring proteins by MS. Using PB (DCP1A, PATL1) and SG (G3BP1, PABC1) markers we were able to identify known granule components (i.e. translation initiation factors associated with SG proteins, and components ��{$��+=��<�� to each type of compartment, shared between them, and not previously known to associate with PBs or SGs, thereby expanding understanding of RNA regulation in eukaryotes.

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Geoffrey Hesketh Lunenfeld-Tanenbaum Research Institute Toronto, ON, Canada

Contributing Authors: Matthew N Seaman, University of Cambridge; Anne-Claude Gingras, Lunenfeld-Tanenbaum Re-search Institute.

ABSTRACT: The evolution of eukaryotic cells relied upon the emergence of a diverse complement of membrane ��7�� 7�of eukaryotic life. The interaction of cells with their environment (through the proper positioning of cell surface proteins such as receptors, transporters, and adhesion proteins), and diversity in the structural and functional �� &�� 7��*��7�� -��!�� +#�� late onset Parkinson disease. VPS35, along with VPS26 and VPS29, is a core subunit of the retromer complex, which recycles a variety of functionally important cargo proteins out of endosomes. Importantly, a general role for a ��7��{��7��+#�"��Ankyrin Repeat Protein (VARP) as a novel interacting partner of the retromer complex. VARP itself interacts with a #$�{��;��\+|<�� {��9+�� -tion. Besides greatly expanding the retromer network, our observations raise new questions about the mechanistic details underpinning retromer function, and how this may contribute to neurodegenerative disease. We therefore propose to perform a detailed analysis of the retromer network interactome using in vivo biotinylation coupled to mass spectrometry (BioID-MS), and will present our preliminary results.

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Syed Shah Ryerson University Toronto, ON, Canada

Contributing Authors: Jean-Philippe Lambert, Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, Canada; Anne-Claude Gingras, Lunenfeld-Tanenbaum Research Institute & Department of Molecular Genetics, University of 9��9��}��?�{�� +�� [��=�� 7��9��}��?��#�� -ham, Department of Chemistry and Biology, Ryerson University, Toronto, Canada.

ABSTRACT: In humans, transport of histones H3/H4 from the cytoplasm to the nucleus occurs in a stepwise fashion and is mediated by protein factors including HSP90, NASP, Asf1 and Importin4.Cells can adopt two differ-ent pathways to assemble chromatin, namely replication dependent and replication independent pathways which are mediated respectively by “Chromatin Assembly Factor 1 (CAF1)” and “Histone Regulator A (HIRA)” protein �� !��[�� *�%*��DNA remains unclear. To address this, we have initiated a proteomic characterization of NASP, HIRA, CAF1 and *��`��9��9��excellent model system to study chromatin related processes providing that it features nuclear dualism in the form of a transcriptionally active macronucleus and a silent micronucleus. Our results indicate that NASP physically interacts with Asf1 and HSP90 and thus likely functions in H3/H4 transport suggesting an evolutionarily conserved nature of this pathway. We also show that the three protein CAF1 complex (Cac1,Cac2, Cac3) interacts with H3/*�� }��;}��<�� #�{%9*{��7��\#�� *��`��"��*��*��'�� *��*�9�complex is also shared by CAF1 complex as its Cac3 subunit. This suggest that Hat2/Cac3 exists in two separate pools of protein complexes and is capable of carrying functions that are divided between two separate proteins in higher eukaryotes such as humans.

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Christopher Go Samuel Lunenfeld Research Institute, Mt. Sinai Hospital Toronto, ON, Canada

Contributing Authors: Wade H Dunham, Lunenfeld-Tanenbaum Research Institute, Mt. Sinai Hospital; Andy Kong,De-��}�� \��=��\��?�¾��}��}��'��9��?=��Raught,Ontario Cancer Institute, UToronto;Alexey Nesvizhskii,Depart. of Computational Medicine and Bioinformatics, UMich-igan;Anne-Claude Gingras,Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital; Dept. of Molecular Genetics, Utoronto

ABSTRACT: A characteristic of eukaryotic life is compartmentalization, which occurs via classical membrane-bound organelles, but also through other subcellular structures and macromolecular complex recruitment. Aberrant sub-cellular localization has been implicated in numerous pathological conditions. Classically, the composition of sub-cellular structures has been determined through cellular fractionation. Additionally, co-localization of a protein with various markers can help reveal compartmental association for the protein under investigation. While powerful, there are limitations to these approaches. Notably, the composition of compartments not amenable to fractionation �� 7��-ing a few proteins at a time. My project aims to use BioID, coupled to mass spectrometry (MS), to provide a high resolution and quantitative proteomic view of the subcellular architecture in human cells. In BioID, a promiscuous biotin ligase (BirA) is fused to a bait protein and stably expressed in human cells to facilitate biotinylation of proteins ��!�� [�� of biotinylated components by MS.An initial set of 42 baits (classical subcellular compartment markers) were se- �� &�� 7�� '�� '�� {�;�� <��5��;�� <��7��;�{"�� <��\#��

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Mr. Guomin Liu Samuel Lunenfeld Research Institute, Mt. Sinai Hospital Toronto, ON, Canada

Contributing Authors: Guomin Liu, Dattatreya Mellacheruvu, Jian Ping Zhang, Zach Wright, James Knight, Mike Tyers, Brian Raught, Hyungwon Choi, Alexey Nesvizhskii and Anne-Claude Gingras

ABSTRACT: �� ;�+"\#<�� protein-protein interactions. In the last several years, we have developed tools to facilitate AP-MS analysis and �� "��*�� ProHits (prohitsms.com) is a laboratory information management system designed to store and analyze mass ��+��*�� !�� &��!��+"\#��9�� 7�� #��Analysis of INTeractome (SAINT) software. Different versions of SAINT, including the most recent SAINTexpress tool are integrated within ProHits. We also report on the integration of the Nested Cluster tool, and new software that enables visualization of clustered data within ProHits.Lastly, we report on two additional tools which are �� +��*��9��}��{��+��;}{�+��<��web-accessible resource to help researchers identify background proteins in AP-MS data. The CRAPome contains Â�� !�� ;�� <��own complete AP-MS experiments, using SAINT or simpler fold-change calculations. We have also developed a searchable web platform (prohits-web.lunenfeld.ca) enabling interrogation of our quantitative proteomics data.

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Francois-Michel Boisvert Assistant Professor, Université de Sherbrooke Sherbrooke, PQ, Canada

Contributing Authors: Romain Drissi; Marie-Line Dubois; Catherine Drouin; Francois-Michel Boisvert.

ABSTRACT: The MCM proteins, which act as a replicative helicase during DNA synthesis, are required for proces-sive DNA replication and are a target of S-phase checkpoints. A mass spectrometry based proteomics approach for measuring the subcellular distribution of the proteome, termed “spatial proteomics”, showed evidence that DNA damage alters the subcellular distribution of the MCM complex. Our results demonstrate that the MCM complex becomes strongly associated with nuclear structures following exposure to DNA damage. Using a quantitative proteomic approach to study the dynamics of protein interactions with the MCM complex during the cellular re-��[$�� MCM2-7 proteins, but also several proteins involved in chromatin organisation. Of particular interest was the DNA damage-dependent increase in the interaction between MCM2 and the histone chaperone ASF1. Moreover, len-tiviral shRNA mediated knock down of proteins from the MCM complex resulted in increased sensitivity to DNA damaging agent and a delay in DNA repair. These results provide valuable clues pointing towards a role for the MCM complex in chromatin remodelling in response to DNA damage.

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Dohee Lee University Health Network, University of Toronto Toronto, ON, Canada

Contributing Authors:�¦��9��}��=��{��?+��\��}��}��¨

ABSTRACT: �� `�;��`<��" �7��;�=:<�� :�7��Ufm1 has been proposed to be covalently conjugated to substrate proteins to affect their function, in an unknown ��9��`��5��;�`<��`�;��<��`�;��<��9��`�� `�� !��Ufm1 has been reported in ischemic heart and type 2 diabetes, its biological roles are not understood. Using both �� ;��+"\#<��=��'["\#�� `��_��`��-��"��"�� 7��!�� (ODR-4) and the Ufm1 protease Ufsp2. ODR-4 is required for localization of odorant receptors to the cilia of olfac-tory neurons in Caenorhabditis elegan. Cilia are hair-like structures located on the surface of cells, and play many �� ;��<��"��(i.e. taste). Defects in cilia can lead to ciliopathies, a collection of human diseases. We are now in the process of conducting a second round of BioID in ciliated and non-ciliated cells. We are also characterizing the intracellular localization of all Ufm1 system components under the same conditions.

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Ms. Deborah Ng University of Toronto Toronto, ON, Canada

Contributing Authors:��}��?�=��{��

ABSTRACT: The ubiquitin system plays many critical roles in normal cellular physiology, and deregulation of this pathway has been implicated in many human diseases. Conjugation of ubiquitin to a substrate is carried out via a

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��"��;�`<��5��;��<�� ;��<��_�� "��"��7�� ;��Â��<�� -jority of these enzymes have not been elucidated. Using a novel technique (BioID) in which protein interactors are �� &��*�� `��"�� _�� -�� `�� `��"��`�� 9�� '�� }9:*�;}"�� -� ��"`" �7�� <�� !��=��*��=��'[��}9:*�� (HLB). The HLB is a nuclear organelle associated with replication-dependent histone gene clusters and contains factors required for histone mRNA processing. We are currently characterizing the role of CTLH in this process.

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Christina Bell IRIC- Université de Montréal Montréal, PQ, Canada

Contributing Authors: Christina Bell, Institute for Research in Immunology and Cancer, Université de Montréal; Guang-hou Shui, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, China; Markus Wenk, Department of Biological Sciences, National University of Singapore, Singapore; Michel Desjardins, Department of Pathology and Cell Biology, Université de Montréal; Pierre Thibault, Institute for Research in Immunology and Cancer, Université de Montréal.

ABSTRACT: Macrophages play an important role in immunity as professional phagocytes. They participate to the clearance of microorganisms through sequential events that include sequestration in phagosomes, degrada-tion by hydrolytic enzymes and processing and presentation of antigens via MHC complexes at the cell surface. These steps involve a complex remodeling of phagosomes through interactions with various organelles, a process ��+��7��'�$"��-tions and regulate the ability of macrophages to elicit a sustained immune response. We used a system biology approach that combined cell fractionation, quantitative proteomics and comprehensive lipidomics analyses of phagosomes to characterize the dynamic association of proteins and lipids during the maturation of resting and IFN-gamma activated macrophages. Quantitative proteomics analyses of phagosome proteins during maturation �� '�$"�� `��|�� $�� Lanosterol synthase were modulated during phagosome maturation and upon IFN-gamma activation. Lipidomics analyses performed on phagosomes obtained from different maturation stages enabled the acquisition of lipid pro-� ��`�� comprised a total of 172 lipid species. In phagosomes from control macrophages, the abundance of cholesterol �� '��'�$"��activated macrophages cholesterol species remained constant. This unique approach, combining proteomics and ��

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Peter Kubiniok Institute for Research in Immunology and Cancer Montréal, PQ, Canada

Contributing Authors: Peter Kubiniok, Institure for Research in Immunology and Cancer (IRIC) Université de Montréal; ��'{'}��]��\��]� ?��}��"��'{'}��]��\��]� ?�\�7�9��'{'}�Université de Montréal; Steve Michnik , Departement of Biochemistry Université de Montréal; Pierre Thibault, IRIC Université de Montréal.

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ABSTRACT: �� -�� 7�� scale, and only limited information is available on corresponding activation pathways. To further understand the �� #��;�+*��<�� 7�� ;#':�}<��^"�!��9�� ;Â��`��<��!�� } �� `�� -�� 9�� of transcription factors including those from the NOT complex that exhibited reversible changes in phosphorylation �� 7��'�� } ��7�� localization and the function of septins during mitosis and that were differentially phosphorylated upon changes of ��9�� 7��[$�� ;��\��\��`<�� and cell cycle progression during heat and cold shock.

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Afnan Abu-Thuraia IRCM - Institut de Recherche Clinique de Montréal Montréal, PQ, Canada

Contributing Authors: Rosemarie Gauthier; Jean-Francois Côte.

ABSTRACT: Metastasis is a process that is often a fatal step during breast cancer progression. To develop strat-egies to reduce breast tumor cells dissemination, it is important that we identify novel modulators of the metastatic process and fully grasp the cellular mechanismsemployed by breast cancer cells to migrate out of the tissue and reach secondary organs. The objective of our study is to unravel the contribution of the receptor tyrosine kinase Axl in the progression of breast cancer to metastasis by characterizing signalling pathways triggered by Axl inducing � ��_�� :��"��function of this RTK. Methods and Results:The methods used in this study contain co-immunoprecipitation as-��{��\��_�� ! ��! �7��=�� be phosphorylated by Axl on 2 tyrosine residues, Y713 and Y717. In addition, Rac activation and cell invasion by �! ��\[�"\="��`�� !�� 9��|`��}�� { ��¤� �� ! ��5��7��{��:\�%[�}�`�� '�� |`��|`|�� -gration, a plausible mechanism would be that it serves as a binding site for pY-binding proteins.

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Philippe Cloutier IRCM - Institut de Recherche Clinique de Montréal Montréal, PQ, Canada

Contributing Authors: Anaïs Aulas, Centre de recherche du centre hospitalier universitaire de Montréal;Mathieu La-vallée-Adam, McGill Centre for Bioinformatics;Mathieu Blanchette, McGill Centre for Bioinformatics;Christine Vande Velde, Centre de recherche du centre hospitalier universitaire de Montréal;Benoit Coulombe, Institut de recherches cliniques de Montréal.

ABSTRACT: A number of degenerative neuromuscular diseases are associated with abnormal protein folding and complex assembly, often through dysfunction of molecular chaperones. Developing strategies for reverting chaperone dysfunction or stimulating normal chaperone activity is awaited for the design of novel therapeutics.

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A family of lysine methyltransferases that preferentially target and regulate molecular chaperones was recently �� }+��\�99:�`[��*��|��\�99:�`�� '��}+�� `��\�99:�`[�is stimulated by addition of UBX cofactor ASPSCR1, which directly interacts with the methyltransferase. This stim-ulatory effect is lost with VCP mutants (R155H, R159G and R191Q) known to cause Inclusion Body Myopathy with Paget’s disease of bone and Fronto-temporal Dementia (IBMPFD) and/or familial Amyotrophic Lateral Sclerosis ;�:#<��:��`��_� 7��=��}+��9+��%[`�� that methylation of this site negatively impacts ATPase activity. In the case of Hsp70, methylation of lysine 561 by \�99:�`�� '��having implications for the development of therapeutics for degenerative neuromuscular disorders such as ALS and IBMPFD, the discovery of a family of chaperone-targeting methyltransferases led us to propose the existence �� ³��´�� proper folding and assembly of protein complexes that make up the human proteome.

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Marie Cargnello IRIC - Université de Montréal Montréal, PQ, Canada

Contributing Authors: Marie Cargnello, Institute for Research in Immunology and Cancer (IRIC), Université de Montréal, Montréal, PQ, CanadaJoseph Tcherkezian, Institute for Research in Immunology and Cancer (IRIC), Université de Mon-��]� ��\��]� ��+^��}��{��}��$�� {��#��]��9�� 9�� :��*�� [��} �=�� 9�� =�� \��#�� *��\�� #�� Boston, MA, USAGenevieve Lavoie, Institute for Research in Immunology and Cancer (IRIC), Université de Montréal, Mon-tréal, PQ, CanadaSteven P. Gygi, Department of Cell Biology, Taplin Biological Mass Spectrometry Facility, Harvard Medical School, Boston, MA, USAPhilippe P. Roux, Institute for Research in Immunology and Cancer (IRIC), Department of Patholo-gy and Cell Biology, Faculty of Medicine, Université de Montréal, Montréal, PQ, Canada.

ABSTRACT: The mammalian target of rapamycin (mTOR) promotes cell growth and proliferation by promoting mRNA translation and increasing the protein synthetic capacity of the cell. Although mTOR globally promotes �� {$��"��'��;�7��<�� -�� {$�� 9�� 7�� -edge, we performed a quantitative proteomic screen to identify proteins that associate with the mRNA 5’ cap in an 9�{"��9��LysC-digested peptides were labeled with 6-plex isobaric mass tags (TMT6) and subjected to liquid chromatog-��"�� ;:}"\#%\#<�� -latory factors, including the putative RNA-binding protein LARP1 (La-related protein 1). Our results indicate that LARP1 associates with actively translating ribosomes via PABP and that LARP1 stimulates the translation of mR-NAs containing a 5’ terminal oligopyrimidine (TOP) motif, encoding for components of the translational machinery. We found that LARP1 associates with the mTOR complex 1 (mTORC1) and is required for global protein synthesis as well as cell growth and proliferation. Together, these data reveal important molecular mechanisms involved in TOP mRNA translation and implicate LARP1 as an important regulator of cell growth and proliferation.

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Cunjie Zhang Lunenfeld-Tanenbaum Research Institute Toronto, ON, Canada

Contributing Authors: Yong Zheng, Lunenfeld-Tanenbaum Research Institute, Mt. Sinai, Toronto, ON; Suya Liu, AB-#}'��9��$?�� :�� "9��{��'��\��#��9��$?�:��9�� :��-feld-Tanenbaum Research Institute, Mt. Sinai, Toronto, ON; Tony Pawson(Deceased), Lunenfeld-Tanenbaum Research Institute, Mt. Sinai, Toronto, ON; Karen Colwill, Lunenfeld-Tanenbaum Research Institute, Mt. Sinai, Toronto, ON; Lunen-feld-Tanenbaum Research Institute, Mt. Sinai, Toronto, ON.

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ABSTRACT: Due to its involvement in many crucial biological functions, phosphorylation is one of the most im-�� "�� 9��¹|��[��parent ion is the hallmark used by search engines to identify a phosphorylation site. However, sulphonation (SO3, +79.956 Da) is almost isobaric with phosphorylation and if it is present in a sample, it may be reported incorrectly �� +�� threonine-containing peptides have been reported.Here, we report that sulphonation can also occur on cysteine �� _��"��"��#��`�� 7� ��9��!� ��=#��-fers (e.g., NP40 lysis buffer, Urea, Tris-HCl, all +/- Na2SO3), exposed to different cysteine-modifying reagents (+/-DTT and iodoacetamide), and sulphonation was measured on QTRAP 5500 or TripleTOF5600 instruments. The levels of cysteine sulphonation were dependent on both the buffer type and the cysteine-modifying reagents. Recognizing this issue, we are now modifying our sample preparation procedures. In addition, we are also em-ploying protocols to distinguish phosphorylation and sulphonation by mass accuracy and peptide fragmentation ��

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Julian Saba �� San Jose, CA, USA

Contributing Authors: Ryan Jacobs; Jason Fong; Greg Botting; Ryan Bomgarden; John Rogers; Rosa Viner; Michael Blank; Julian Saba; Neelu Puri.

ABSTRACT: _�� 7�� ;9�'�<� �� {�� ;��{<� ��$��"#� �} �:��}��;$#}:}<�� 9�'��9��9�'��$#}:}��-lotinib-resistant cell lines were established. To identify differences in protein expression between parental and ��"�� %�� !�� 9\9��\#�analysis were utilized. Parental and H358 erlotinib-resistant NSCLC cell lines were treated with 10ÂμM erlotinib in ��+��&�� 9\9�� !�reagents plus two 13C/15N isotope variants of the TMT6-127 and TMT6-129 reagents. The peptides were then �� &��":}�� 9�\#��\#��+��[��%��+�� %�� performed by Protein Center. Using HCD MS2 or MS3 fragmentation, more than 1500 protein groups were identi-�� +��!�� showed a noticeable decompression using the multinotch MS3 method with many more proteins exhibiting a �� 9��*�� 7��!��[$�� -tion/transcription factors and replication licensing proteins, apoptosis inhibitors, and some Ras associated signal transduction elements.

��"��+@�B�+>

Mr. Nebiyu Abshiru IRIC - Université de Montréal Montréal, PQ, Canada

Contributing Authors:�� }��":�&��\��]� ?�{�� &��\��]� ?�}�� Pomies,University of Montréal; Alain Verreault, University of Montréal; Pierre Thibault, University of Montréal.

ABSTRACT: *�� "�� " ��*��*�� '�� "�� tryptic digestion. This approach generates tryptic peptides bearing more than one internal lysines that are either acetylated or propionylated. The same peptide can be acetylated on different lysines thereby forming positional

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��9�� {+":}%\#��9��"��acetylation in these peptides is a major challenge. Our method combines high resolution LC-MS/MS data with a �� "�� \#%\#�features that are diagnostic of each co-eluting peptide. These features were then used to deconvolute composite spectra generated from synthetic peptides mixtures composed of acetylated positional isomers. We further tested our approach using tryptic digests of histones extracted from human cells. K562 cells were treated for 1, 6 and 24h with DMSO as control or the HDAC inhibitors MS-275, SAHA and JNJ-26481585. Our results show that all the inhibitors cause global increases in acetylation of H4K5, H4K8, H4K12 and H4K16. Moreover, compared to cells treated with MS-275, we observed a striking increase in H4K5 and H4K12 acetylation six hours after treatment of K562 cells with JNJ-26481585. Diacetylation of H4K5 and H4K12 is a highly conserved pattern of acetylation in newly synthesized H4 molecules. Therefore, our results raise the exciting possibility that JNJ-26481585 inhibits HDACs that deacetylates new H4 molecules following their deposition into chromatin.

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Megan Lewicki University Health Network, University of Toronto Toronto, ON, Canada

Contributing Authors: Megan Lewicki, Tharan Srikumar; Brian Raught.

ABSTRACT: 9�� " �7��;#�\�<�� 5�-gated to target proteins to affect their functions in a number of ways. Interestingly, while environmental stresses effect the rapid transcriptional activation of stress response genes (and inhibit the transcription of others), in many cases this is accompanied by a dramatic burst of SUMOylation. How this SUMO burst is linked to stress-mediated changes in transcription was not understood. To this end, we: (i) characterized SUMO stress response (SSR) ki-��#�\��##{�� ?�;��<��#�\��to hyperosmotic shock, and; (iii) conducted RNA sequencing in the presence and absence of a functional SUMO system. Surprisingly, we found that while proteins involved in transcriptional control and chromatin remodeling are the primary SUMO stress response targets, cells in which the SUMO system is inactivated retain a fully functional �� '�� !�� de-repressed in the absence of SUMO system function. Together, our data suggest that SUMOylation is in fact not required for acute transcriptional control, but is instead required to preserve transcriptional repression via the maintenance of proper higher order chromatin structure.

Q��38��&��

Natalie Marshall University of British Columbia Vancouver, BC, Canada

Contributing Authors: Theo Klein [3]; Nikolay Stoynov [4]; Leonard J. Foster [4,5]; Christopher M. Overall [5,6]; B. Brett Finlay [1,2,5]; [1] Department of Microbiology & Immunology, UBC[2] Michael Smith Laboratories, UBC[3] Department of Oral Biological and Medical Sciences, UBC[4] Centre for High-Throughput Biology, UBC[5] Department of Biochemistry & Molecu-lar Biology, UBC[6] Centre for Blood Research, University of British Columbia.

ABSTRACT: �� ;�+�}<�� [��-��+�}��'''��;9�##<��³� �� ´��5�� intestinal cells. T3SS effector proteins subvert human cellular processes and cause disease. Characterizing the functions of T3SS unique effectors remains a challenge. Understanding human cellular proteome dynamics during infection could elucidate T3SS effector functions. This study aims to characterize T3SS-mediated alterations in the ��;$<"�� $"�� 7��+�}�9�##��$ }%[��\�� may reveal how these and other global pathways are affected by T3SS effectors. Cultured human HeLa cells were

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��`�� "��+�}�� 9�##��$"�� of infected human cells were quantitatively compared using terminal amine isotopic labeling of substrates (TAILS) �� {�� ¤�_� ��`��$"�� `��`��[{��+� ��9�##"�� 7��9�##"��$�� -skeletal rearrangements, etc. A novel T3SS-mediated proteolytic event in annexin A2, which interacts with two T3SS effectors, may provide further details on how the T3SS mediates host cytoskeletal rearrangements during �� 7��+�}��

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Adam Rabalski Western University, Department of Biochemistry London, ON, Canada

Contributing Authors:�¦[��_��:�� [��=��_��?¨¦�� :�5�� Dept. of Biochemistry, Western University]

ABSTRACT: CK2 is a constitutively active serine/threonine protein kinase overexpressed in many forms of cancer and is an emerging therapeutic target. Previous global phosphoproteomic studies implicate CK2 in the DNA Dam-age Response (DDR), as CK2 motifs are considerably represented in phosphoproteomes perturbed by DNA dou-ble strand breaks; however, these studies do not experimentally modulate CK2 to directly test whether CK2 does indeed regulate pathways linked to the DDR. To overcome this limitation, the goal of this work is the development ��"��7�� &��}��"��9�� &��#��}��!��;#}<�� 9�� -peptides from cell extracts. We also investigated the utility of porous graphitic carbon (PGC) as an alternative to }"`�9�#+�� }"`�9�� }��;�#%9"!"!"[%�<��#��#�� !��}"`�9��+�}�� phosphopeptide enrichment and mass spectrometry analysis yielded complementary sets of phosphopeptides. {��#}�� can be further enhanced. Future experiments will utilize Stable Isotope Labeling by Amino Acids in Cell Culture ;#':�}<�� }��"��!��!��stress. Overall, it is anticipated that this study will advance the knowledge of the CK2-dependent phosphopro-teome within the context of genotoxic stress which may aid the design of combinatorial cancer therapy strategies.

�� I3*-�33��

Andrew Crowell Dalhousie University Halifax, NS, Canada

Contributing Authors: Dr. Alan A. Doucette, Dalhousie University, Halifax, NS, Canada.

ABSTRACT: ��:�� &"�� ;9��Anal. Chem. 2008, p. 1568). By modifying the composition of the polyacrylamide gel ‘column’, as well as the col- �� *��5�� &��:�� ;:\_+<�� ;��7[�<�The LMWP of human plasma is recognized as a valuable source of candidate biomarkers. These proteins show increased permeability from tissue (Drake, Proteom Clin. Appl. 2007 p. 758). Unfortunately, MS analysis of these and other low abundance plasma proteins is challenged by the masking effect of high abundance proteins. Re-duction of these abundant proteins has been achieved through a number of strategies including immunodepletion, �� &"��;�� <�� plagued by loss of target proteins owing to a “piggyback” effect onto the abundant carrier proteins (e.g. albumin)

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To minimize loss of the LMWP during high abundance protein depletion, sodium dodecyl sulphate (SDS) is added to the plasma, thus ensuring complete denaturation and solubilize of the proteome. With SDS solubilization as �� :�� prefractionating the LMWP into 12 discrete fractions. MS analysis can be achieved following depletion of the SDS-containing fractions through solvent-based protein precipitation.

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Monika Tucholska Lunenfeld-Tanenbaum Research Institute Toronto, ON, Canada

Contributing Authors: Jean-Philippe Lambert; Christopher Go; Anne-Claude Gingras.

ABSTRACT: Chromatin—an essential element of gene regulation—has a complex organization where the basic structural unit is the nucleosome, consisting of two copies of each core histone around which 147 base pairs of DNA are wrapped. Proteins physically embedded in chromatin such as histones are poorly solubilized in conven-�� ;�+<�� "�� _�� of histone binding partners (Lambert et al., R8��, in press). Here, we investigate whether an alternative approach to map protein-protein interactions, in vivo biotinylation coupled to mass spectrometry (BioID-MS) that is compatible with harsh lysis conditions, can also be employed for the interrogation of chromatin interactomes. =��"��&��+"\#��=��'["\#��histone H2B (125 and 117, respectively); however, there was a surprisingly low overlap between both approaches �� _��+"\#��=��'["\#�� -lowing GO term analysis, it appears that the AP-MS approach favours proteins with DNA binding domains while BioID-MS favours protein complexes that are chromatin regulators and involved in transcription regulation. As ��+"\#��=��'[��

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Nicholas Brodie University of Victoria Victoria, BC, Canada

Contributing Authors:��+��7��=}�+��}��?�}��=��"��BC Proteomics Centre.

ABSTRACT: In amyotrophic lateral sclerosis (ALS) misfolding of the protein SOD1 results in toxic aggregation in motor neurons, leading to their degeneration and to symptoms of the disease. The structural nature of this mis-�� $\{��"�� '�� and crosslinking were employed. Native SOD1 is a homodimer, each monomer containing two Î²-sheets oriented perpendicular to the dimer interface and two metal binding sites for Cu2+ and Zn2+ on the opposite side. Dif-�� 7�� }=[+#��`�\��[9�� `�� 7��of them differential, involving all 10 lysines as well as the N-terminus. All four of these were located in the loop region of the metal binding sites in SOD1, including the crosslink K4-K69, which suggests that this region may �� [�� $*#"��+}�#"12C314N/13C315N reagent yielded complete coverage of all 10 lysines as well as the N-terminus, Y109, and T56. :��`��!��#�[`��[9��`�� !-posed, further indicating that loss of metal binding leads to opening of the loop region. T56 was less exposed upon �[9��'�� ¯°��³�� ´�� 9��and bringing K69 closer to K4.

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2��/�� /��#�� E��*��'�

Jason Serpa University of Victoria, Genome BC Proteomics Centre Victoria, BC, Canada

Contributing Authors:��#��?��+��7�?�}��=��

ABSTRACT: Zero-length crosslinking combined with MS is a valuable method for studying protein structure and ��"�� [��&��" �� 7�� 7�� 9�� "��$"�� ;}��`��<��!� ��;+}�#"*�%[�<� ;+��7�� `�<�� 7�� $"�� [��9��;��"9{9�#9['�{�##{��[�:'$��{<�� -��;{$��#�� &�� <�� 7��&��" �� 7��[}��+'}�+��described previously. The PCAS reagent will modify N-termini as well as amino acids K, Y, S, and T. The treatment ��\�$*��*��#��9��$��"��-��&�� "��;��<�which may be present in standard tryptic digests. The NTM DXMSMS Match Software was developed in-house �� $"�� 7��9�� 7��based on precursor mass and the combined light and heavy MS/MS spectra. The method described here -- a com-��$"�� !��""�� ":��"�� &��" �� 7��

��6��

Andrew Crowell Dalhousie University Halifax, NS, Canada

Contributing Authors: [Andrew Crowell, Dalhousie University;] [Samantha Rudolph, Dalhousie University;] [Alan Dou-cette, Dalhousie University]

ABSTRACT: Bottom up analysis depends on successful manipulation of the sample ahead of MS analysis. For dilute protein samples, or samples containing contaminants or solubilizing additives (eg SDS), a preconcentration/ �� } �� !�� ease of automation. Unfortunately, SDS depletion is ineffective with most chromatographic approaches. By con-trast, organic solvent-assisted protein precipitation is extremely effective at SDS-removal (Botelho, J. Proteome Res, 2010, p2863). Despite its effectiveness, protein precipitation remains a cumbersome approach. Quantitative protein recovery is possible (Crowell, Anal. Chim. Acta, 2013, p48), though it relies on the pipetting skills of the individual. It is therefore desirable to create a robust protein precipitation protocol ahead of enzymatic digestion, making SDS-containing protein samples amenable to bottom-up MS analysis.The ProTrap XG is a disposable two ��#[#"�� a plug to the base of the device. With plug removed, solvent and SDS are removed by centrifugation. With plug at-tached, the captured protein pellet is resolubilized and subject to tryptic digestion. Subsequent cleanup is achieved �� %� ��spin cartridge in a centrifuge. The device is evaluated with standards and complex proteome mixtures, noting ��;:}%��<�� #[#��;�� <��\#�� "#[#��

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*��3��&�� )��

Veronique Laforte, Biomedical Engineering Department, McGill University Montréal, PQ, Canada

Contributing Authors:�¦#��=��\�� »��^��'��}��=�� -gineering Department, McGill University, Montréal, Canada; Veronique Laforte, McGill University & Genome Quebec Innova-��}��[��$�� »�$��=�� \�� \��]� ��}��-��?�+�7�#��:��\�� »��^��'��}��=�� [��\�� \��]� ��}��?�*��:��\�� »��^��'��}��=�� Department, McGill University, Montréal, Canada; David Juncker, McGill University & Genome Quebec Innovation Center and [��$�� »�$��=�� \�� \��]� ��}��¨

ABSTRACT: ��5�� quill pins, because they have open channels that allow buffers to be in contact with the atmosphere. This evapo-ration during printing leads to a change in antibody concentration, and liquid viscosity, which changes the amount of antibody being printed on the surface. Glycerol has been used in the past to limit evaporation of printing buffers, however it also known to inhibit binding of antibodies on the surface. In this study, we compared 14 chemicals, �� _�� ;� �� !�� 1000Da, ethylene glycol, 1,3-butanediol, 2,3-butanediol and betaine) in pairwise combinations, spotted on 4 differ-ent functionalized glass surfaces. We found that excellent spot morphology, antibody density and antibody activity �� ;�� <?��"�� \�� ̀ ��-yl sulfoxide on Nexterion Slide H, a thin hydrogel slide manufactured by Schott. Overall, the quality of spots using those two buffers was greatly increased, which is essential for immunoassays carried on microarrays. Decreasing the variability in antibody printing is critical for the reliability required in clinical studies.

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Andy Ng Biomedical Engineering Department, McGill University Montréal, PQ, Canada

Contributing Authors:�*��:�?��\��&��?��$�?�[��7��+��=�� Department, McGill University, Montréal, PQ, Canada.

ABSTRACT: Multiplex protein analysis has great potential for application in disease diagnosis, monitoring and drug screening. Microarrays have been developed to allow for thousands of miniaturized assays to be performed in parallel on a single chip, however they require expensive spotters for microarray fabrication. As an alternative, chip-to-chip transfer technology has been developed to deliver an array of pre-spotted reagents from one chip to the assay chip, thus avoiding the requirement of a spotter during the assay. However, this multiplexing capability is still relatively low due to the limited array density achievable in the transfer process, mainly due to inaccuracies in chip alignment. Here, we present a novel microarray-to-microarray transfer method by sequentially transferring reagents onto the assay slide using the snap chip. The array has a density of 625 spots cm-2, corresponding to more than 5000 spots on a single slide, an amount that has not been achieved before. The alignment accuracy is characterized and a user-friendly snapping system for aligning and snapping the two slides is presented. The multiplexing capability of this transfer method is demonstrated through a sandwich immunoassay quantifying 50 proteins simultaneously, which is the largest one-pot multiplex sandwich assay on a slide to date. The assay achieved limits of detection in the pg/mL range for 35 of the protein targets. We also conducted a 4-plex enzyme inhibition assay and obtained results comparable to bulk volume assays. This work demonstrates that the snap chip is a powerful, easy-to-use and versatile tool for high-throughput protein analysis.

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*5F�5�%�+�I+��3/��%�Q�%7��/%�45�7� �!5�!!�7�2��/I��-*�%43��//3! -5!��7�!3��52%

Gina Zhou McGill University Montréal, PQ, Canada

Contributing Authors: Sebastien Bergeron; David Juncker.

ABSTRACT: =��7��¤��&�" ��7�� 7��!��scanners limits their adoption in clinical practice. Methods: Here, we introduce a multiplexed silver enhanced sand-��;#�$#'�<��9��7�� 7��}}:��}}:�� 7��':"`¯°��':"��':"`��':"`��¯°"$�� #��G-CSF, MMP3, SPARC, and TNF-RII that have been linked to cancer progression. The accumulation of silver �� &��7��scanner or a smartphone and open-source data analysis software. Results: Up to 16 samples and 14 proteins �� ;:�[<��%:�� #�$#'��7�� ;}}:��"}#��':"`�<�� the remaining proteins. The concentrations of the proteins measured in diluted serum using the two methods were also found to be in good agreement (R2> 0.9).Conclusion: A low-cost easy-to-use 14-plex sandwich assay was �� -��#�$#'�� "��multiplexed immunoassays for diagnostics.

*��"��+!.� �� . � ��&��6�)�5%�"�%��

Julian Saba �� San Jose, CA, USA

Contributing Authors:�\�� #�7�� 7��#��9��\�� \��}��{��Hühmer, Julian Saba.

ABSTRACT: DIA is an emerging high throughput quantitative technique for quantifying large number of proteins of interest in the biological complex samples. A novel mass spectrometer, Orbitrap Fusion enables parallel acqui-sition of Orbitrap SIM and rapid ion trap MS/MS, maximizing instrument duty cycle. Here we developed a high ��['��7��*{%�\�#'\�� ;��<�;_�#'\<��cover all precursor ions of 400—1000 m/z. In parallel with each SIM scan, 17 sequential ion trap MS/MS with 12 amu isolation windows are acquired to cover the associated 200 amu SIM mass range. With this WiSIM-DIA ap-proach, quantitative information for all precursor ions detected in three sequential SIM scans is recorded in a single run. Plus, all MS/MS fragment information over the mass range of 400—1000 m/z is recorded. The targeted data !�� '}��;}`�¹}`�<��#�� and matching fragment ion distribution with that stored in the spectral library. The quantitative performances (LOD/LOQ, dynamic range, reproducibility and accuracy) and throughput of this new approach were evaluated using various samples. This new data independent acquisition (WiSIM-DIA) approach enabled accurate and reproduc-ible quantitative results for any precursor ions detected in SIM mode without prior knowledge on Orbitrap Fusion MS.

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��'��'�

Wu Yang Jin University of British Columbia Vancouver, BC, Canada

Contributing Authors: Wu Yang Jin, UBC; Xuelai Fan, UBC; Jie Lu, UBC; Yu Tian Wang, UBC.

ABSTRACT: Despite the massive socioeconomic toll of the ischemic stroke, there is currently no effective neu-�� {��"��7��`�;[�+�`<��7��mediator of ischemic cell death. Following an ischemic insult, DAPK1 activates and binds to the GluN2B subunit of the NMDA Receptor (NMDAR) and enhances NMDAR-mediated currents and excitotoxicity. DAPK1 is also involved in mediating cell-death through non NMDAR-mediated oxidative stress. In an effort to reduce ischemic brain injury, we fashioned a blood brain barrier and cell membrane permeant peptide TAT-GluN2BCTM which, �� \}�� 7��7��;��-tive) DAPK1 through chaperone-mediated autophagy. A single intravenous injection of 10mg/kg TAT-GluN2BCTM ;�� 9�9"� �$�=<�`�� 7��7��[�+�`�� by the insult, including the ipsilateral striatum and nearby cortex as visualized with hematoxylin and eosin stain-��\�� 7��7��[�+�`��9�9"� �$�=}9\��substantial reduction of the infarct area and fewer number of degenerating neurons in comparison with that pro-vided by uncoupling DAPK1 from the GluN2B receptor signaling complex with TAT-GluN2B. Together, these data point to DAPK1 as a key target for the intervention of neurotoxicity and illustrate peptide-mediated degradation of death-promoting proteins as a new avenue to develop therapeutics for stroke.

�� *��!��

Robert Popp University of Victoria, Genome BC Proteomics Centre Victoria, BC, Canada

Contributing Authors: Robert Popp, David Malmstrom, Alex G. Camenzind, Andrew G. Chambers, J Grace van der Gugten, Daniel T. Holmes, Christoph H. Borchers.

ABSTRACT: The renin angiotensin aldosterone system (RAAS) is crucial for the regulation of blood pressure. Dysfunctions of the RAAS can lead to severe diseases related to hypertension. A well-established biomarker for the diagnosis of primary aldosteronism, a form of secondary hypertension, is plasma renin activity (PRA). It is commonly determined by radioimmunoassay (RIA), which has the disadvantages of using radioisotopes and the possibility of cross-reactivity. To overcome these limitations we have developed a mass spectrometric PRA assay based on immuno-MALDI (iMALDI). Magnetic beads conjugated with anti-Ang I antibodies are incubated with stable isotope-labeled Ang I analogues (Ang I SIS) and plasma (incubated at 37 ÂºC for Ang I generation, or kept on ice as a blank), washed and spotted directly onto a MALDI target. Addition of acidic MALDI matrix elutes the captured peptides from the beads.In order to develop a robust and high throughput assay suitable for clinical im-plementation we automated the sample preparation on an Agilent Bravo liquid handling robot followed by analysis ��=��7��\��!�\�:['"9��'�� {'�"�� ;{��<��"��"��`��}�� 9��evaluate the automated iMALDI PRA assay, PRA values for 200 patient samples, selected from the routine primary aldosteronism screening program at St. Paul’s Hospital (Vancouver), will be analyzed and compared to an LC-MS/MS approach.

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3��&��-%�!� � ��"�� *��(

Xiaodong Wang University of Victoria, Genome BC Proteomics Centre Victoria, BC, Canada

Contributing Authors: Xiaodong Wang,University of Victoria-Genome BC Proteomics Centre, Victoria, Canada;Jun Han,University of Victoria-Genome BC Proteomics Centre, Victoria, Canada;Juncong Yang,University of Victoria-Genome BC Proteomics Centre, Victoria, Canada;Jingxi Pan,University of Victoria-Genome BC Proteomics Centre, Victoria, Can-ada;Christoph Borchers,University of Victoria-Genome BC Proteomics Centre, Victoria, Canada & Uvic Dept of Biochemistry and Microbiology, Victoria, Canada.

ABSTRACT: Over the past few years, more than a dozen of new matrices have been introduced for tissue im-aging of lipids by MALDI-MS in the positive-ion detection mode, yet very few have been shown to be suitable for negative-ion MS imaging. Herein, we explored the use of quercetin for the imaging of lipids in both rat brain and ��"��9'}{"\#��{�� "�� '��the use of 2-MBT and 9-AA as the matrices detected only 103 and 87 lipids, respectively. 9-AA was found not to be very suitable for imaging on an Apex Qe 12-Telsa quadrupole-FTICR instrument because of its partial matrix sublimation in the high-vacuum source. In addition, the reconstituted ion maps showed that many lipids that were weakly-detected with 2-MBT or 9-AA displayed clearer spatial distribution patterns. Cardiolipins, gangliosides and glycosphingolipids were detected with much higher ion intensities when quercetin was used. For prostate �� Â�� distributions between the cancerous and the non-cancerous cell regions based on T-tests (p<0.05). This work demonstrated that quercetin performed better than other commonly-used matrices for the imaging of lipids by negative-ion MALDI-MS, and showed the value of ultrahigh-resolution FTICR-MS in combination with quercetin for better detection of potential cancer biomarkers.

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Karen Colwill Mount Sinai Hospital Toronto, ON, Canada

Contributing Authors: [Adrian Pasculescu, Lunenfeld-Tanenbaum Research Institute,Mount Sinai Hospital, Toronto; ��\��#��9�� [��7��:��[��7?+��}��! �9�� [��7��:��Denmark; Yong Zheng, Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto; Marina Olhovsky, Lun-enfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto; Ruijun Tian, Lunenfeld-Tanenbaum Research Insti-tute, Mount Sinai Hospital, Toronto; Jonathan So, Institute of Medical Science, University of Toronto, Toronto; Rachel D. Vanderlaan, Department of Cardiac Surgery, University of Toronto, Toronto; Tony Pawson, Lunenfeld-Tanenbaum Research Institute,Mount Sinai Hospital, Toronto (deceased); Rune Linding,Technical University of Denmark, Lyngy, Denmark; Karen Colwill, Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto]

ABSTRACT: A major challenge in mass spectrometry and other large-scale applications is how to handle, inte-grate, and model the data that is produced. Given the speed at which technology advances and the need to keep pace with biological experiments, we designed a computational platform, CoreFlow, which provides programmers with a framework to manage data in real-time. It allows users to upload data into a relational database (MySQL), and to create custom scripts in high-level languages such as R, Python, or Perl for processing, correcting and �� }�� &�� 5��"�� 7�� between related tasks, and enables the generation of summary reports as well as publication-quality images. As a result, the gap between experimental and computational components of a typical large-scale biology project is reduced, decreasing the time between data generation, analysis and manuscript writing. CoreFlow is now avail-�� "��7��;��<�� "��!�� which include corrections for incomplete isotopic labeling of peptides (SILAC) or arginine-to-proline conversion, and modeling of multiple/selected reaction monitoring (MRM/SRM) results.

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Alexandr Ignatchenko The University Health Network Toronto, ON, Canada

Contributing Authors: Vladimir Ignatchenko, Princess Margaret Cancer Center, University Health Network, Toronto, ON, Canada; Stephenie D. Prokopec, Informatics and Bio-computing Program, Ontario Institute for Cancer Research, Toronto, ON, Canada; Ankit Sinha, Princess Margaret Cancer Center, University Health Network, Toronto, ON, Canada, University of Toronto, Department of Pharmacology & Toxicology, Toronto, ON, Canada; Paul C. Boutros, Informatics and Bio-computing Program, Ontario Institute for Cancer Research, Toronto, ON, Canada, University of Toronto, Department of Medical Biophys-ics, Toronto, ON, Canada, University of Toronto, Department of Pharmacology & Toxicology, Toronto, ON, Canada; Thomas Kislinger, Princess Margaret Cancer Center, University Health Network, Toronto, ON, Canada, University of Toronto, Depart-ment of Pharmacology & Toxicology, Toronto, ON, Canada.

ABSTRACT: In recent years, open source software (OSS) has become a popular option in mass-spectrometry based shotgun proteomics. Protein sequence database-based search algorithms such as X!Tandem, OMSSA, \��\�� }��#�� -gorithms like SpectraST are gaining popularity because of increased speed and higher sensitivity for the analysis of complex biological mixtures. However, these approaches are often limited by the number of validated peptide �� ;+#\<�� +��integration of multiple search algorithms. Here, we introduce a new graphical user interface-based proteomic pipeline developed on Python that integrates PSM from multiple search algorithms. The interface will allow users to customize various parameters, such as selection of search algorithms, false discovery rate (FDR) thresholds and the structure of the generated reports. Resulting analyses are stored in an object-relational database man-agement system (ORDBMS), that will allow retrospective tracing of data while preserving integrity. FDR calculation for peptides will be based on a target-decoy approach and the post-PSM peptide-protein groupings are performed in accordance with the rules of parsimony. The user can obtained reports for desired algorithm combinations with �� [{��_�� 7�� -ments by other developers. Currently, our software is being tested on large sets of proteomics data and has shown ��

Q��0��R��-M�&��&��"��8

Vladimir Ignatchenko Ontario Cancer Institute, University Health Network, University of Toronto Toronto, ON, Canada

Contributing Authors: Alexandr Ignatchenko, Princess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada; Ankit Sinha, Princess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada, Department of Medical Biophysics, University of Toronto, Toronto, ON, Canada; Paul Boutros, Department of Medical Biophysics, University of Toronto, Toronto, ON, Canada, Informatics and Bio-computing Program, Ontario Institute for Cancer Research, Toronto, ON, Canada, University of Toronto, Department of Pharmacology & Toxicology, Toronto, ON, Canada; Thomas Kislinger, Princess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada, Department of Medical Biophysics, University of Toronto, Toronto, ON, Canada.

ABSTRACT: Venn diagrams are a graphical representation of the overlap and differences between multiple data sets and are often used in proteomics and genomics experiments for qualitative illustration of differences between individual experiments. All currently existing tools for the generation of Venn diagrams fall into one of the follow-��!�� 7��{�� 7��&�� *�� shortcomings, termed Venn Diagram Interactive Software (VennDIS), a JavaFX based solution for producing us-�"��&�� 9��['#�� &��9�� way comparisons and modify attributes, such as font, style and position of the labels, background color, sizes and

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shape outline color. It is platform independent (i.e. can be run on any modern operating systems such as Windows, :��!��\��#<�� 7��9��\:�� !��"��

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Razvan Florin Albu University of British Columbia Vancouver, BC, Canada

Contributing Authors:��_��=}?��}��=}?��=}?�9�� \��=}�

ABSTRACT: In order to avoid the formation of potentially harmful protein aggregates, it is critically important for cells to eliminate misfolded proteins in a timely manner. However, some proteins escape quality control mecha-nisms and can form persistent aggregates that are the staple of many neurodegenerative pathologies, including Parkinson’s and Alzheimer’s disease. Aided by the advent of mass spectrometry-based proteomics, the protein composition of these aggregates has begun to be characterized in recent years. However, the reasons why some proteins appear to be more aggregation-prone than others remain poorly understood. We are investigating which features distinguish proteins found predominantly in the insoluble fraction from the rest of the proteome. Because of the pathological effects associated to protein aggregation within the central nervous system, we have focused on brain tissue, using mice as a model organism. We are employing quantitative mass spectrometry-based pro-�� &�� "�� analysis to determine enrichment of any standout features, such as amino acid composition, length, disordered regions or particular primary sequence motifs. Our preliminary results, comparing soluble and insoluble proteins in �� 9��next step will be to compare brain tissue harvested from mice at an old age to tissue taken from younger, adult animals. Using this system, we hope to identify which proteins become more insoluble with the onset of old age, and which features distinguish them from predominantly soluble ones.

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Andy Kong University of Michigan Ann Arbor, MI, USA

Contributing Authors: Alexey Nesvizhskii, University of Michigan.

ABSTRACT: Advances in the speed of modern mass spectrometers, along with growing interest in proteogenom-��"�� _�� popular database search tools can be executed on compute clusters, the associated infrastructure and operational ��*�� inexpensive graphical processing units (GPUs), which offers cluster level performance on a personal workstation. Previous efforts in GPU-accelerated database search have focused on the similarity scoring step, causing bot-tlenecks to arise in peptide generation. To combat this issue, we designed a novel peptide generation scheme �� +�"�� "�� "��"��!��_�� +�"�� +�"�� [#º��search tool that integrates accelerated peptide generation and scoring. Using an AMD Radeon 7950 video card, ��!�� }+�� -tion, with greater speedups anticipated as we increase CPU-GPU concurrency. Additionally, we examined several fundamental questions in the performance of database search tools, such as the relative computational complex-ities of different spectral similarity functions, the relative computational complexities of peptide generation versus ��[#º�� routine proteomics analysis while enabling the exploration of novel questions in proteomics that were previously untenable due to the computational costs involved.


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Yassene Mohammed University of Victoria, Genome BC Proteomics Centre Victoria, BC, Canada

Contributing Authors: Yassene Mohammed, Dominik Domanski, Angela M. Jackson, Derek S. Smith, Andre M. Deelder, Magnus Palmblad, Christoph H. Borchers.

ABSTRACT: Multiple Reaction Monitoring (MRM) is a powerful method for targeted proteomics. Designing an MRM assay is a multistep process that starts by choosing the most appropriate peptides to represent the target protein. Selecting ideal peptides involves careful decisions, which can become error-prone as the reviewed in-��_�� 7�� `��=��#� ��15 R, and 2 XPath building blocks, i.e. processors. These processors enforce 30 selection rules on the compiled information about each target protein, its tryptic peptides, and the suitability of these peptides for MRM. The infor-�� +��=��$}='��#$+��!+�#��+�� +{'[��+\�_��-�� }��³��´�� \{\�experiment based on the uniqueness of the peptide in the targeted proteome, its physiochemical properties, and �� 7�� error, by considering all of the isoforms of the protein, and by taking into account the protein’s post-translational �� "�� This resulted in faster and more standardized peptide selection -- our software running in a single thread selected peptides for 49 proteins in less than 1 hour, compared to 8 proteins per day done manually.


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Floor layout Hilton Hotel


List of participants

FIRST NAME LAST NAME COMPANY email

Nebiyu Abshiru Université de Montréal [email protected]

Afnan Abu-Thuraia IRCM [email protected]

Razvan Florin Albu University of British Columbia [email protected]

Laurie Allan BRUKER [Diamond Sponsor] [email protected]

Kanwal Ashraf York University [email protected]

Zuhier Awan McGill University [email protected]

Mohan Babu University of Regina [email protected]

Christina Bell Institute for Research in Immunology and Cancer [email protected]

Nicolas Bisson Cancer Research Centre, Laval University [email protected]

Francois-Michel Boisvert Universite de Sherbrooke [email protected]

Christoph Borchers UVic - Genome BC Proteomics Centre [email protected]

Marguerite Boulos IRCM [email protected]

Sylvie BourassaProteomics/CHU de Quebec Research Center/

Université [email protected]

Nicholas Brodie University of Victoria [email protected]

John Burke University of Victoria [email protected]

Irene Campoy-Moncayo [email protected]

Johnny Cardenas AB SCIEX [Diamond Sponsor] [email protected]

Marie Cargnello IRIC-Université de Montréal [email protected]

Cara Cesario Thermo Scientific [Diamond Sponsor] [email protected]

Josee Champagne IRCM [email protected]

Francois Chartier [email protected]

Philippe Cloutier IRCM [email protected]

Karen Colwill Mount Sinai Hospital [email protected]

Shannon Cornett BRUKER [Diamond Sponsor] [email protected]

Emilie Cossette IRIC-Université de Montréal [email protected]

Judith Cotton-Montpetit IRCM [email protected]

Jasmin Coulombe-Huntington IRIC-Université de Montréal [email protected]

Mathieu Courcelles IRIC-Université de Montréal [email protected]



Amber Couzens Lunenfeld-Tanenbaum Research Institute, Mt Sinai [email protected]

Andrew Crowell Dalhousie University [email protected]

Nikki Curtis Cambridge Isotope Laboratories, Inc. [email protected]

Dylan Dieters-Castator University of Western Ontario [email protected]

Alan Doucette Dalhousie University [email protected]

Maroun El Khoury Thermo Scientific [Diamond Sponsor] [email protected]

Daniele Fabris University of Albany [email protected]

Denis Faubert IRCM [email protected]

Matthew W. Forbes University of Toronto [email protected]

Diane Forget IRCM [email protected]

Leonard Foster University of British Columbia [email protected]

Jennifer Frahm Thermo Scientific [Diamond Sponsor] [email protected]

Jacob Galan IRIC-Université de Montréal [email protected]

Karen Gambaro McGill University [email protected]

Marie-Soleil Gauthier IRCM [email protected]

Scott Gerber Dartmouth University [email protected]

Golfam Ghafourifar Université de Montréal [email protected]

Anne-Claude Gingras Lunenfeld-Tanenbaum Research Institute, Mt Sinai [email protected]

Christopher GoSamuel Lunenfeld Research Institute, Mt. Sinai

[email protected]

Makan Golizeh Université du Quebec a Montréal (UQAM) [email protected]

Marilyn Goudreault Lunenfeld-Tanenbaum Research Institute, Mt Sinai [email protected]

Diana Paola Granados IRIC-Université de Montréal [email protected]

Gerry Guilfoy Cambridge Isotope Laboratories, Inc. [email protected]

Sam M. Hanash The University of Texas, MD Anderson Cancer Center [email protected]

Geoffrey Hesketh Lunenfeld-Tanenbaum Research Institute, Mt Sinai [email protected]

Ruth Huttenhain University of California [email protected]

Alexandr Ignatchenko The University Health Network [email protected]

Vladimir Ignatchenko [email protected]



Wu Yang Jin University of British Columbia [email protected]

Evgeny Kanshin IRIC-Université de Montréal [email protected]

Jim Kapron BRUKER [Diamond Sponsor] [email protected]

Juergen Kast University of British Columbia [email protected]

Neil Kelleher Northwestern University [email protected]

Isabelle Kelly Centre Proteomique CHU de Quebec [email protected]

Brenda Kesler Thermo Scientific [Diamond Sponsor] [email protected]

Jacquet Kevin CRCHUQ [email protected]

Yunee Kim University of Toronto [email protected]

Don Kirkpatrick Genentech [email protected]

Thomas Kislinger Princess Margaret Cancer Center [email protected]

John Klassen University of Alberta [email protected]

Andy Kong University of Michigan [email protected]

Mathew Kramar [email protected]

Gary Kruppa BRUKER [Diamond Sponsor] [email protected]

Peter Kubiniok Institute for research in Immunology and Cancer [email protected]

Miljan Kuljanin Western University [email protected]

Prem Kumarathasan Health Canada [email protected]

Andree-Anne Lacombe IRCM [email protected]

Veronique Laforte McGill University [email protected]

Jean-Philippe Lambert Lunenfeld-Tanenbaum Research Institute, Mt Sinai [email protected]

Mathieu Lavallee-Adam The Scripps Research Institute [email protected]

Andre LeBlanc UQAM [email protected]

Dohee Lee [email protected]

Megan Lewicki University of Toronto [email protected]

Liang Li University of Alberta [email protected]

David LitchfieldUniversity of Western Ontario,Department of

[email protected]



Guomin LiuSamuel Lunenfeld Research Institute, Mt. Sinai

[email protected]

Pammy (Pik Shan) Lo McGill University [email protected]

Bin Ma University of Waterloo [email protected]

Natalie Marshall University of British Columbia [email protected]

Janice Mayne University of Ottawa [email protected]

Thibault Mayor University of British Columbia [email protected]

Xavier Misonne AB SCIEX [Diamond Sponsor] [email protected]

Yassene Mohammed University of Victoria-Genome BC Proteomics Centre [email protected]

Mike Moran University of Toronto [email protected]

Tom Moy AB SCIEX [Diamond Sponsor] [email protected]

Patrick MurphyHarvard Medical School (Invited Speaker-Thermo

Scientific)[email protected]

Mario NavarreteManitoba Centre for Proteomics and Systems

[email protected]

Alexey Nesvizhskii University of Michigan, Dept. of Pathology [email protected]

Andy Ng McGill University [email protected]

Deborah Ng [email protected]

Marlene Oeffinger Institut de recherches cliniques de Montréal [email protected]

Dennis Orton Dalhousie University [email protected]

Gladys Perroquin [email protected]

Joelle Perusse IRCM [email protected]

Evgeniy Petrotchenko UVic - Genome BC Proteomics Centre [email protected]

Christian Poitras Institut de recherches cliniques de Montréal [email protected]

Robert Popp UVic - Genome BC Proteomics Centre [email protected]

Paul Poupart Agilent Technologies Canada Inc. [email protected]

Lawrence Puente Ottawa Hospital Research Institute [email protected]

Adam Rabalski UWO Dept. of Biochemistry [email protected]

Vincent Richard Concordia University [email protected]

Anja Rodenbrock Université de Montréal [email protected]



Justine Rousseau IRCM [email protected]

Philippe Roux IRIC-Université de Montréal [email protected]

Julian Saba Thermo Scientific [Diamond Sponsor] [email protected]

Payman Samavarchi-Tehrani Lunenfeld-Tanenbaum Research Institute, Mt Sinai [email protected]

Jason Serpa UVic - Genome BC Proteomics Centre [email protected]

Syed Shah Ryerson University [email protected]

Bay Sheldrick Thermo Scientific [Diamond Sponsor] [email protected]

Romain Simon [email protected]

Brigitte Simons AB SCIEX [Diamond Sponsor] [email protected]

K.W. Michael Siu University of Windsor and York University [email protected]

Tharan Srikumar University of Toronto [email protected]

Nicole St-Denis Lunenfeld-Tanenbaum Research Institute, Mt Sinai [email protected]

Hanno Steen Children’s Hospital of Boston [email protected]

Sylvain Tessier IRCM [email protected]

Pierre Thibault Université de Montréal [email protected]

Mathilde Triquigneaux Concordia University [email protected]

Monika Tucholska Lunenfeld-Tanenbaum Research Institute, Mt Sinai [email protected]

Olga Vitek Purdue University [email protected]

Karen Waldron University of Montreal, Department of Chemistry [email protected]

David Walsh Concordia University [email protected]

Xiaodong Wang UVic - Genome BC Proteomics Centre [email protected]

Shoshana Wodak Hospital for Sick Children [email protected]

Haiyuan Yu Cornell University [email protected]

Cunjie Zhang Lunenfeld-Tanenbaum Research Institute, Mt Sinai [email protected]

Gina ZhouMcGill University /Quebec Genome Innovation

[email protected]


NOTES

Diamond

Silver

Bronze

Iron

Gold

BCProteomicsNetwork

We thank all of our sponsors

THE SCIENCE OF WHAT’S POSSIBLE.™

UVic Genome BCPROTEOMICS CENTRE

CNPN 2014 Symposium��+��

April 14-16, 2014Hilton Bonaventure, Montreal, Quebec.

IN HEALTH AND DISEASE - CNPNProteome Dynamics in Health and Diseases 3 Contents Workshops 5 Short...

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Transcript of IN HEALTH AND DISEASE - CNPNProteome Dynamics in Health and Diseases 3 Contents Workshops 5 Short...