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NetBioSIG2014-Keynote by Marian Walhout
- 1. Interspecies Systems Biology: Nutritional Regulatory Networks A.J. Marian Walhout
- 2. Interspecies Systems Biology Intraspecies Systems Biology Development Behavior Physiology Disease
- 3. Intraspecies systems biology - Networks Protein-protein interaction networks Gene regulatory networks Metabolic networks ?
- 4. Protein-protein interaction networks Gene regulatory networks Metabolic networks ? ? Intraspecies systems biology - Networks
- 7. Two effects of nutrition
- 8. ? How is metabolism regulated in response to diet? ?
- 9. ? ? How is metabolism regulated in response to diet?
- 10. 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
- 11. Does bacterial diet affect C. elegans physiology? Life history traits Development Fertility Lifespan (aging)
- 12. E. coli OP50 Standard Laboratory Diet E. coli HT115 Used for RNAi feeding Comamonas DA1877 Soil isolate Measure developmental time, reproduction and lifespan MacNeil et al., Cell 2013
- 13. The life cycle of the worm
- 14. Dietary changes in life-history traits O P50 H T115 A B C Totalnumberofoffspring D A1877 MacNeil et al Figure 1 0 50 100 150 200 250 300 O P50H T115D A1877 0 10 20 30 40 50 60 70 80 90 100 1 5 9 13 17 21 25 29 DA1877 OP50 HT115 0 10 20 30 40 50 60 70 80 90 100 Adult Late L4 Mid-late L4 Mid L4 Early L4 L3 Percenttotalanimals Percentsurvival Days after L4 Reproduction Comamonas: faster development, fewer offspring, shorter lifespan Development Lifespan OP50 = E. coli DA1877 = Comamonas MacNeil et al., Cell 2013
- 15. Dietary changes in gene expression Nutrigenomics Microarray expression profiling of worms fed different bacterial diets
- 16. 46 272368 Core downregulated CUB domain Lectin GST Metabolic enzyme Transporter Neuropeptide Worm specific Transcription Other Core upregulated n=41 n=46 A set of core diet-response genes MacNeil et al., Cell 2013 Are these changes transcriptional?
- 17. Core downregulated CUB domain Lectin GST Metabolic enzyme Transporter Neuropeptide Worm specific Transcription Other n=46 A proxy gene acdh-1 Repressed >300-fold on Comamonas diet MacNeil et al., Cell 2013
- 18. Transgenic C. elegans reporter strains GFPPacdh-1 E. coli OP50 Comamonas DA1877
- 19. 5 mM OP50HT115DA1877 A C exp x25 exp x25 B D DA1877 (long exp The dietary response is transcriptional Pacdh-1::GFP – 1.5 kb promoter A dietary sensor in living animals E. coli OP50 Comamonas DA1877 5 mM OP50HT115DA1877 exp x25 exp x25 B D DA1877 (long exp NGM OP50 MacNeil et al., Cell 2013
- 20. Starvation represses Pacdh-1 E. coli OP50 Starvation Does a Comamonas diet mimic starvation? MacNeil et al., Cell 2013
- 21. E. coli OP50 GFP ON Comamonas DA1877 GFP 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
- 22. The dietary and starvation responses are distinct DA1877 OP50* *Exposure time 1/20 DA1877 1/200 DA1877 1/500 DA1877 1/1000 DA1877 1/10 000 DA1877 1/100 000 DA1877 OP50 1/20 exposure 1/100 DA1877 Also accelerated development and similar gene expression changes under 1/1000 dilution
- 23. Small amounts of one diet can have dramatic effects on gene expression and physiology, even when mixed with another diet
- 24. Unhealthy?+ = Healthy?+ = Comamonas DA1877 produces dilutable compound to which worms respond Two interpretations with very different implications
- 25. Physiology – life history traits
- 26. ? ?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
- 27. C. elegans target of rapamycin (TOR) and insulin signaling pathways are not involved in the response to Comamonas MacNeil et al., Cell 2013
- 28. 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?
- 29. 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 35 Multiple NHRs (worms have 271!)
- 30. Most repressors are metabolic genes, not transcription factors Watson, MacNeil et al., Cell 2013
- 33. Mutations in human homologs of repressors give rise to inborn metabolic diseases treated by dietary intervention
- 35. ? ? 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
- 36. Screen I: E. coli deletion collection Watson et al., Cell 2014 70 genes 7 genes
- 37. Screen II: Comamonas transposon mutagenesis 5 hits Lesley MacNeil Watson et al., Cell 2014
- 38. 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
- 39. A bacterial metabolic network implicated in the C. elegans dietary response Watson et al., Cell 2014
- 40. Metabolite screen: Supplement metabolites to C. elegans dietary sensor on either bacterial diet in increasing concentrations Watson et al., Cell 2014
- 41. Metabolites increasing GFP on Comamonas Watson et al., Cell 2014
- 42. Metabolites decreasing GFP on E. coli Watson et al., Cell 2014
- 43. Ado-Cbl and Me-Cbl are two forms of vitamin B12 Mutation in either C. elegans gene interferes with the response to Comamonas
- 44. Watson, MacNeil et al., Cell 2013 Vitamin B12 is in the C. elegans dietary response network
- 45. Is vitamin B12 the Comamonas dilutable molecule? Mass spectrometry of bacteria
- 46. Amy Caudy (U Toronto) Vitamin B12 levels in Comamonas are orders of magnitude higher than in E. coli Watson et al., Cell 2014
- 47. Vitamin B12 pathway status correlates perfectly with dietary sensor activity
- 48. Vitamin B12 pathway status correlates perfectly with dietary sensor activity
- 49. A metabolic network connecting the two vitamin B12 pathways Me-Cbl Ado-Cbl Propionyl-CoA Watson et al., Cell 2014
- 50. How does the gene regulatory network respond to Comamonas or E. coli?
- 51. B12 fails to repress GFP expression in propionyl-CoA breakdown mutants
- 52. Vitamin B12 is not sufficient to drive sensor repression Model: balance between Vitamin B12 and propionyl-CoA Chemical epistasis: add both B12 and propionic acid
- 53. Excess propionic acid is epistatic to Vitamin B12 Watson et al., Cell 2014
- 54. Excess propionic acid is epistatic to Vitamin B12 Watson et al., Cell 2014
- 55. So far all experiments with sensor Does vitamin B12 mimic broad Changes in C. elegans gene Expression elicited by Comamonas diet? qRT-PCR of 28 Comamonas-response genes
- 56. 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
- 57. Does vitamin B12 supplementation by Comamonas Explain its effects on C. elegans life history traits?
- 58. Vitamin B12 accelerates C. elegans development B12 reduces fertility B12 has no effect on lifespan Watson et al., Cell 2014
- 59. How does B12 accelerate development? Watson et al., Cell 2014
- 60. Vitamin B12 developmental acceleration occurs 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
- 61. Which metabolite drives developmental acceleration? Watson et al., Cell 2014
- 62. SAM production is important for developmental acceleration Watson et al., Cell 2014
- 63. What is the physiological role of the Other B12 pathway?
- 64. Metabolites increasing GFP on Comamonas Comamonas E. coli Watson et al., Cell 2014
- 65. Vitamin B12 mitigates propionic acid toxitiy Watson et al., Cell 2014
- 66. Functions of vitamin B12 development PA toxicity
- 68. Conclusions C. elegans and bacteria are a powerful interspecies system: Nutritional networks Effects of microbiota (SCFAs/vitamin B12)
- 69. Acknowledgements Lesley MacNeil Emma Watson Safak Yilmaz Ashlyn Ritter Jote Bulcha Juan Fuxman Bass Aurian Garcia Gonzales Gabrielle Giese Amy Holdorf Mike Hoy Rudi Kaushik Akihiro Mori Shaleen Shrestha Alex Tamburino Efsun Arda (now at Stanford) Julie Zhu (UMMS) Adam Rosebrock Amy Caudy (U Toronto) NIDDK NIGMS
Notas del editor
- Surprisingly, the majority of repressors were not TFs. Instead 24 of the 33 genes encode metabolic enzymes found in four specific pathways.
- One other interesting realization that we made was that most of the metabolic repressors found in our screens that could reverse the transcriptional diet response in C elegans have human orthologs associated with mendelian diseases. Here is a disease network with all the metabolic repressors from our screens connected to their human orthologs. These human orthologs are connected to their associated inborn metabolic disorders. Most of these orders are managed clinically by dietary intervention, and here on the right are dietary components that patients must avoid (red) or supplement (green), and as you can see these disorders have a lot of similarities in their dietary treatments.