NetBioSIG2014-Keynote by Marian Walhout
-
Upload
alexander-pico -
Category
Science
-
view
484 -
download
2
description
Transcript of NetBioSIG2014-Keynote by Marian Walhout
Interspecies Systems Biology:
Nutritional Regulatory Networks
A.J. Marian Walhout
Interspecies Systems Biology
Intraspecies Systems BiologyDevelopment
BehaviorPhysiology
Disease
Intraspecies systems biology - Networks
Protein-protein interaction networks
Gene regulatory networks Metabolic networks
?
Protein-protein interaction networks
Gene regulatory networks Metabolic networks
? ?
Intraspecies systems biology - Networks
Nutrition
Interspecies biology
Symbiosis
Pathogenesis
Interspecies systems biology
Symbiosis
Nutrition
Pathogenesis
Two effects of nutrition
?
How is metabolism regulated in response to diet?
?heterogeneous
heterogeneous
?
?
How is metabolism regulated in response to diet?
Bacteria
(diet/prey)
C. elegans
(predator)
Superworm: nematode C. elegans
• Hermaphrodite • ~3 days development• ~2 week life span• Lineage known - 959 cells• Transparent – GFP transgenics• RNAi, transgenics, mutants• Well annotated genome
• Feed individual bacterial species• Model for diet• Model for microbiota
Does bacterial diet affectC. elegans physiology?
Life history traits
DevelopmentFertility
Lifespan (aging)
E. coli OP50Standard Laboratory Diet
E. coli HT115Used for RNAi feeding
Comamonas DA1877Soil isolate
Measure developmental time, reproduction and lifespan
MacNeil et al., Cell 2013
The life cycle of the worm
Dietary changes in life-history traits
Reproduction
Comamonas: faster development, fewer offspring, shorter lifespan
Development Lifespan
OP50 = E. coliDA1877 = Comamonas
MacNeil et al., Cell 2013
Dietary changes in gene expression
Nutrigenomics
Microarray expression profiling of worms fed different bacterial diets
A set of core diet-response genes
MacNeil et al., Cell 2013
Are these changes transcriptional?
A proxy gene
acdh-1Repressed >300-fold on Comamonas diet
MacNeil et al., Cell 2013
Transgenic C. elegans reporter strains
GFPPacdh-1
E. coli OP50
Comamonas DA1877
The dietary response is transcriptionalPacdh-1::GFP – 1.5 kb promoter
A dietary sensor in living animals
E. coli OP50
Comamonas DA1877
MacNeil et al., Cell 2013
Starvation represses Pacdh-1
E. coli OP50
Starvation
Does a Comamonas diet mimic starvation?
MacNeil et al., Cell 2013
E. coli OP50GFP ON
Comamonas DA1877GFP OFF
Prediction: if Comamonas is nutrient poor, mixing it with E. coli should alleviate this
Comamonas DA1877+
E. coli OP50
GFP ?MacNeil et al., Cell 2013
The dietary and starvation responses are distinct
DA1877
OP50**Exposure time 1/20
DA18771/200
DA18771/500
DA18771/1000 DA1877
1/10 000 DA1877
1/100 000 DA1877
OP501/20 exposure
1/100 DA1877Also accelerated development and similar gene expression
changes under 1/1000 dilution
Small amounts of one diet can have dramatic effects on gene expression
and physiology, even when mixed withanother diet
Unhealthy?+ =
Healthy?+ =
Comamonas DA1877 produces dilutable compound to which worms respond
Two interpretations with very different implications
Physiology – life history traits
??
How does C. elegans respond toE. coli and Comamonas diets?
C. elegans genetics
Which bacterial molecules affect gene expression and life history traits?
Bacterial genetics
Interspecies systems biology
C. elegans target of rapamycin (TOR) and insulin signaling pathways
are not involved in the response to Comamonas
MacNeil et al., Cell 2013
What are the gene regulatory networks involved in the response to Comamonas?
The response to Comamonas is transcriptional
Comamonas makes a compound or signal
Transcription factors?
Signaling pathways?
What are the gene regulatory networks involved in the response to Comamonas?
Forward and reverse genetic screens
Activators
146
Worms cannot respond to Comamonas diet
Repressors
35Multiple NHRs
(worms have 271!)
Most repressors are metabolicgenes, not transcription factors
Watson, MacNeil et al., Cell 2013
BCAA breakdown
TCA cycle
Glycine cleavage
Methionine/SAM
cycle
Mutations in human homologs of repressors give rise to inborn metabolic diseases treated by dietary intervention
? ?How does C. elegans respond to
E. coli and Comamonas diets?C. elegans genetics
Which bacterial molecules affect gene expression and life history traits?
Bacterial genetics
Interspecies systems biology
Screen I: E. coli deletion collection
Watson et al., Cell 2014
70 genes
7 genes
Screen II: Comamonas transposon mutagenesis
5 hits Lesley MacNeil
Watson et al., Cell 2014
From mutants to metabolites
Sequence Comamonas genome – identify genes
Map E. coli and Comamonas mutant genes onto metabolic network using KEGG
Annotate biological processes/pathways
Annotate candidate metabolites – mutant gene leads to buildup of precursor and lack of product
A bacterial metabolic network implicated in the C. elegans dietary response
Watson et al., Cell 2014
Metabolite screen:
Supplement metabolites to C. elegans dietary sensoron either bacterial diet in increasing concentrations
Watson et al., Cell 2014
Metabolites increasing GFP on Comamonas
Watson et al., Cell 2014
Metabolites decreasing GFP on E. coli
Watson et al., Cell 2014
Ado-Cbl and Me-Cbl are two forms of vitamin B12
Mutation in either C. elegans gene
interferes with the response to Comamonas
Watson, MacNeil et al., Cell 2013
Vitamin B12 is in the C. elegans dietary response network
Is vitamin B12 theComamonas dilutable
molecule?
Mass spectrometry of bacteria
Amy Caudy (U Toronto)
Vitamin B12 levels in Comamonas are orders of magnitude higher than in E. coli
Watson et al., Cell 2014
Vitamin B12 pathway status correlatesperfectly with dietary sensor activity
Vitamin B12 pathway status correlatesperfectly with dietary sensor activity
A metabolic network connecting the two vitamin B12 pathways
Me-Cbl
Ado-Cbl
Propionyl-CoA
Watson et al., Cell 2014
How does the gene regulatory network respond to Comamonas or E. coli?
B12 fails to repress GFP expression in propionyl-CoA breakdown mutants
Vitamin B12 is not sufficient todrive sensor repression
Model: balance betweenVitamin B12 and propionyl-CoA
Chemical epistasis: add both B12 and propionic acid
Excess propionic acid is epistatic toVitamin B12
Watson et al., Cell 2014
Excess propionic acid is epistatic toVitamin B12
Watson et al., Cell 2014
So far all experiments with sensor
Does vitamin B12 mimic broadChanges in C. elegans gene
Expression elicited by Comamonas diet?
qRT-PCR of 28 Comamonas-response genes
Vitamin B12 mimics Comamonas-induced changes in C. elegans gene expression
Two types of B12-regulated genes
Type 1: responsive to propionic acid
Type 2: not responsive to propionic acid
Watson et al., Cell 2014
Does vitamin B12 supplementation by Comamonas Explain its effects on C. elegans life history traits?
Vitamin B12 accelerates C. elegans development
B12 reduces fertilityB12 has no effect on lifespan
Watson et al., Cell 2014
How does B12 accelerate development?
Watson et al., Cell 2014
Vitamin B12 developmental accelerationoccurs via methionine/SAM cycle
Mutants in propionic acid breakdown are accelerated by B12
Mutants in methionine/SAM cycle are not accelerated by B12
Watson et al., Cell 2014
Which metabolite drivesdevelopmental acceleration?
Watson et al., Cell 2014
SAM production is important for developmental acceleration
Watson et al., Cell 2014
What is the physiological role of the Other B12 pathway?
Metabolites increasing GFP on Comamonas
Comamonas E. coli
Watson et al., Cell 2014
Vitamin B12 mitigates propionicacid toxitiy
Watson et al., Cell 2014
Functions of vitamin B12
development
PA toxicity
Vitamin B12
Propionic acid
Conclusions
C. elegans and bacteria are a powerful interspecies system:
Nutritional networks
Effects of microbiota (SCFAs/vitamin B12)
Acknowledgements
Lesley MacNeilEmma Watson
Safak YilmazAshlyn RitterJote BulchaJuan Fuxman BassAurian Garcia GonzalesGabrielle GieseAmy HoldorfMike HoyRudi KaushikAkihiro MoriShaleen ShresthaAlex Tamburino
Efsun Arda (now at Stanford)
Julie Zhu (UMMS)
Adam RosebrockAmy Caudy (U Toronto)
NIDDKNIGMS