Endocytosis, Midwest 2011

7/27/2019 Endocytosis, Midwest 2011

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2011 National Academies Northstar Institute for Undergraduate Education in Biology

Endocytosis: the Dynamic Nature

of the Endomembrane System

Notes to Instructors are italicized on slides

and in notes


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Context1 class session in Introductory Biology for majors

Taught in a large lecture hall

Pre-existing information (from previous lectures)

Cell parts (including cytoskeleton)

Membrane structure

Exocytosis

Reading before class:Textbook section on endocytosis

How Viruses HijackEndocyticMachinery


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Goal: Understand the role of endocytosis incell physiology

Outcomes Explain steps in endocytosis using the concepts of

molecular recognition and sorting

Explain the function of endocytosis in cell metabolism Predict how perturbations of endocytic processes

affects cells Explain how the endocytic process may be hijacked by

pathogens Analyze and interpret experimental data Connect graphical data with visual representations


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Assessments

Identify and order the steps of endocytosis Use concepts of molecular recognition and sorting to

explain specificity in the process Interpret data about how a perturbation affects

endocytosis and predict consequences for cells Given information about how a drug affects endocytosis,

analyze experimental data to determine how a pathogenenters a cell

Predict how drug treatment might prevent a pathogenfrom reaching the cytosol


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7/14/11

New material is presented in multiple waysincluding extensive use of animations.

Interactions include everyone in the class.

The unit incorporates a variety of learningactivities and student interactions.

Material incorporates diverse examples that willreach different audiences.

Diversity


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Class plan (single class session)

Before class: Students complete assigned reading

1. Think-pair-share activity (10 min)

Use a diagram to identify and order steps

in the endocytic pathway

2. Lecture (20 min)

Key concepts: molecular recognition and sorting

3. Data analysis and predictions (20 min)Brainstorming and clicker questions

Homework assignment


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How do cells eat?

Membrane transporters & channels

Endocytosis


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Pathogens use the endocytic pathway to get into cells

e.g.: flu virus

Salmonella ?


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Think-Pair-Share: Endocytosis

A) THINK INDIVIDUALLY (4 minutes)Read the terms on next slideChoose endocytosis termsDraw a labeled diagram using those terms

B) PAIR (2 minutes)Discuss your diagram with your neighborModify your diagram as needed

C) SHARE (2 minutes)We will use one stellar example in lectureModify your diagram as we discuss


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Choose appropriate terms andcreate your own labeled diagram

CargoClathrinCoated pit

Coated vesicleEndosomeERGolgi

LysosomeMitochondriaNaked vesiclePlasma membraneReceptor


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Lecture starts

Mini-lecture + student input to cover theconcepts on the next few slides and videos

Remind students to add to their diagram:

Phagocytosis, pinocytosis

differences and similarities


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Types of Endocytosis

1. Multiple pathways ofendocytosis

2. Molecular recognition between cell (receptor)

and cargo (internalized object)

Biology (2002) Campbell et al.


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Receptor-mediated Endocytosis

Animated Video: Key Points

1. Recognition of SPECIFIC cargo

2. Multiple steps + compartments

3. Consequences for cargo

http://www.youtube.com/watch?v=lNKKg6hIBcM

Molecular Biology of the Cell (2002) Alberts et al.
http://www.youtube.com/watch?v=lNKKg6hIBcMhttp://www.youtube.com/watch?v=lNKKg6hIBcMhttp://www.youtube.com/watch?v=lNKKg6hIBcM


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Lecture continues

Coverstepsclathrin

acidification-2 steps

Concepts:Molecular recognition

Sorting

cargo-receptor

vesicle-vesicle

Why do we need this? Molecular Biology of the Cell (2002) Alberts et al.


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Phagocytosis

Animated Video: Key Points

1. Similarities to receptor-mediated endocytosis

Molecular recognition

Sorting

2. Differences

No clathrin

http://www.youtube.com/watch?v=7VQU28itVVw

Animation: McGraw-Hill
http://www.youtube.com/watch?v=7VQU28itVVwhttp://www.youtube.com/watch?v=7VQU28itVVw


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Phagocytosis

Molecular recognition between cell and bacterium

Lysosomes fuse with phagosome:

Sorting: vesicles specific places

Molecular recognition: lysosomes phagosomes

What would happen if lysosomes fused withany (all) cellular membranes?


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Exercise:Assess understanding of endocytosis by interpretingreal data

A study used a drug that blocks endocytosis to determinehow viruses enter/infect cells.

Viruses:

Ebola virusVesicular stomatitis virus (VSV)Human Immunodeficiency Virus (HIV)

Drug: Chlorpromazine (Drug C)

Adapted from: Bhattacharyya et al (2010)


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Drug C: effect on normal cell endocytosis

Images show clathrin localization inside cells.

nucleus

nucleus

Clathrin coated

pits or vesicles

Clathrin clumps

C



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Brainstorm in pairs (2 min):

What steps of endocytosis might be affected by Drug C?

Drug C: effect on normal cell endocytosis

nucleus

nucleus

Clathrin coated

pits or vesicles

Clathrin clumps

C


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Drug C

Clathrin clumps= non-functional

?

?

Adapted from Genes & Dev. 1998. 12: 3663-3674


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%o

fcellsi

nfected

Drug C

Ebola

Drug C: effect on viral entry into cells

Test cells received Drug C + virus. Untreated cells received only virus. Control cells received no virus. 3 viruses: Ebola, VSV and HIV.

Infection is when virus reached cytosol



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%o

fcellsi

nfected

Drug C

Ebola




Clicker Q1 (1 min): Whichvirus(es) enter the cell byclathrin-mediated endocytosis?

A. Ebola onlyB. VSV onlyC. HIV only

D. VSV and HIV


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%o

fcellsinfected

Drug C

Ebola




Clicker Q2 (1 min): Whichvirus(es) require receptors to

enter the cell?

A. Ebola onlyB. VSV onlyC. HIV onlyD. Not enough information


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Clicker Q3 (1 min.): Bafilomycin blocks endosomeacidification. Which cell diagram represents whereEbola will be located after treatment with this drug?(Small spots = virus.)

A B C D

V C

VC

PREDICT THE OUTCOME


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Drug C

Clathrin clumps= non-functional

BafilomycinBlocks endosome

acidification?

?


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Q1: What is the most likely method by which VSVgenters the cell?A. Clathrin-mediated endocytosisB. phagocytosisC. pinocytosisD. not enough information

Q2: Justify your answer to Q1

Q3: Based on experiments with Chlorpromazine(discussed in class) and Bafilomycin, whatmethod do you think HIV uses to enter the cell?

Q4: What would happen to a normal cell treated

with Bafilomycin, i.e. what are the side effectsof this drug?

%o

fcellsinfected

Bafilomycin

This experiment is like the one in class, except

cells were treated with Bafilomycin, whichinhibits endosome acidification.

HomeworkBafilomycin: effect on viral entry into cells

Ebola

Endocytosis, Midwest 2011

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