Steven Hallam University of British Columbia SRI International 2013 Overview Through the looking glass Environmental PathwayGenome Databases MetaPathways Pipeline Development 2 Metabolism ID: 1047313
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1. Analysis of environmental genomes using Pathway ToolsSteven Hallam | University of British ColumbiaSRI International, 2013
2. OverviewThrough the looking glass…Environmental Pathway/Genome DatabasesMetaPathways Pipeline Development2
3. MetabolismMetabolism, or the synthesis and decomposition of chemicals in a cell can be organized into pathways represented by graphs.3Vertex = chemical [substrate, product]Edge = enzyme
4. Cellular PathwaysOur genetic and biochemical understanding of metabolism is based largely on the study of complete pathways within cells.4Genome Management Information System, Oak Ridge National Laboratory
5. Distributed PathwaysHowever, microbial communities form distributed metabolic pathways directing matter and energy exchange.5
6. Community MetabolismThe goal is to predict and compare distributed pathways to better understand biogeochemical cycling and community metabolism in the environment.6
7. 7Predicting Community MetabolismPlurality SequencingSingle-Cell SequencingFragment Recruitment, SOM, PCAEnvironmental PGDB (ePGDB)with Taxonomic BinningSimulated ePGDB
8. MetagenomeDistributed PathwaysFrom Genomes to Biomes8Biogeochemical Cycles “The regulation of the pools and fluxes in biogeochemical cycles have their origins in the genetic inventory of individual microbes, and the regulation of these genes within the organism is determined by the environment. As such, one can look at the microbial food web as a collection of genomes whose expression and replication is coordinated through complex feedback loops at the organismal, population, and ecosystem level. “Chisholm Falkowski et al., (2008) Science 320, 1034-1038
9. Foundational QuestionsWhat is the taxonomic and functional structure of the ecosystem? How does this structure change in response to environmental perturbation? What are the ecological consequences of this change? What are relevant units of selection, conservation or utilization for ecological genomic resources?9
10. OverviewThrough the looking glass…Environmental Pathway/Genome DatabasesMetaPathways Pipeline Development10
11. Gene ProductsGenes/ORFsGenomic Map11Gene ProductsGenes/ORFsGenomic MapPathologic*CompundsReactionsGenomic MapGenes/ORFsGene ProductsPathwaysPGDBOrganismsPathwaysReactionsCompoundsInference of Metabolic Pathways* Integrates genome and pathway data to identify putative metabolic networksPGDB Navigator
12. Pathway/Genome Navigator*http://ecocyc.org/META/new-image?type=PATHWAY&object=GLYCOLYSISPGDB*HomepagePathway ViewerEvidence GlyphPathway InformationGene InformationMetaboliteEnzyme FoundUnique Enzyme12
13. Gene ProductsGenes/ORFsGenomic Map13Gene ProductsGenes/ORFsGenomic MapPathologic*CompoundsReactionsGenomic MapGenes/ORFsGene ProductsPathwaysePGDBPathwaysReactionsCompounds* Integrates genome and pathway data to identify putative distributed metabolic networks???Environmental PGDB
14. 14ePGDB Navigation
15. 15http://engcyc.org/
16. OverviewThrough the looking glass…Environmental Pathway/Genome DatabasesMetaPathways Pipeline Development16
17. 17MetaPathwaysA modular pipeline for constructing Pathway/Genome Databases from environmental sequence information MetaPathways currently supports four “data products” including i) GenBank submission, ii) LCA, iii) MLTreeMap, and iv) ePGDBs with associated feature summary tables and GFF files MetaPathways externalizes compute-intensive processes onto a user defined cluster using Sun Grid Engine or the Amazon elastic cloud
18. 18MetaPathwaysePGDBs facilitate pathway-centric exploration of environmental sequence information using Pathway Tools and the MetaCyc web interfaceProvides inference-based approach to metabolic reconstruction based on explicit computational rules to predict presence or absence of distributed metabolic networksMetaPathways can be used with multi-molecular data sets (DNA, RNA or protein) sourced from cultured isolates, single-cells and natural or human engineered ecosystemshttp://www.github.com/hallamlab/MetaPathways http://hallam.microbiology.ubc.ca/MetaPathways
19. 19ePGDB Navigation
20. 20ePGDB Validation
21. 21EcoCyc PathwaysThe number of E. coli pathways identified using the MetaCyc blast database decreases with increasing blast score ratio (BSR) cut-off while the others stay relatively constant. From this an optimal BSR between 0.4-0.6 can be inferred.
22. 22MetaSim Pathways
23. 23Synthetic EcologyThe pathway (S-adenosyl-L-methionine cycle II) was identified by Pathway Tools in the simulated metagenome based on the combined contribution of two genomes (a + b).
24. 24Infering Trophic InteractionsAn ePGDB constructed for the Mealybug symbionts Tremblaya princeps and Moranella endobia predicted interpathway complementarity in essential amino acid biosynthetic pathways.McCutcheon, J.P. and von Dohlen, C.D. “An interdependent metabolic patchwork in the nested symbiosis of mealybugs.” Current Biology, 2011, DOI: 10.1016/j.cub.2011.06.051
25. 25c1988-2012Hawaii Ocean Time Series (HOT)DeLong et al. Community Genomics Among Stratified Assemblages in the Ocean’s Interior. (2006) Science 311T. Danhorn, C. R. Young, E. F. Delong, Comparison of large-insert, small-insert and pyrosequencing libraries for metagenomic analysis, ISME J (2012), doi:10.1038/ismej.2012.35.
26. 26Environmental Sequence InformationHOT Sample Depth (m)DescriptionInformationSequencing PlatformNumber of SequencesAverage Sequence Length Protein Coding SequencesAnnotated Coding SequencesMetaCyc ReactionsMetaCyc Pathways25upper euphoticDNARoche 454623559257405613214149413886475upper euphoticDNARoche 4546736742444306892225724052854110chlorophyll maxDNARoche 4544731662703360351657754133860500mesopelagic DNARoche 454995747276714743361193446494925upper euphoticRNARoche 45456182124823440485781343372375upper euphoticRNARoche 454557718239203359668553208669110chlorophyll maxRNARoche 454398436228135107369122549532500mesopelagic RNARoche 454479661266207465714003034641ePGDBs were generated for environmental sequence information (DNA and RNA) sourced from the HOT water column.
27. 27Core PathwaysTop 50
28. 28Cellular OverviewComparison of DNA (Blue) and RNA +DNA (Red) pathway predictions
29. 29Comparison of genetic potential and gene expression data in photic and dark ocean watersPathway Partitioning
30. 30Diagnostic Pathways
31. 31For each depth interval, a small number of cryptic pathways were predicted in RNA that were not predicted in DNA data setsThese pathways showed depth distributions consistent with niche-partitioning between sunlit and dark ocean watersCryptic Pathways
32. 32Known HazardsMissing ATP citrate lyase indicates false positive for rTCA
33. Things to Keep in Mind…Pathologic cannot predict pathways not present in MetaCycEvidence for short pathways is hard to interpretFalse positives due to shared enzymes in multiple pathways or incorrect annotations create hazardsCurrently no taxonomic assignment or coverage information is mapped onto identified pathwaysLimited functional validation for pathways in metagenomes33
34. “One gene is many hypotheses”Anonymous34
35. Joint Genome InstituteSusannah TringeTijana Glavina del Rio Institute for Ocean SciencesMarie RobertRobin BrownUniversity of British ColumbiaPacific Northwest National LaboratoryAngela NorbeckLjiljana Pasa-TolicHeather Brewer35Sam KheirandishKishori KonwarKeith MewisAntoine PageMelanie ScofieldYoung SongNicole SukdeoJody WrightElena ZaikovaMaya BhatiaMonica Torres BeltranAnnie CoxEvan DurnoDiane FairlyEsther GeisAlyse HawleyAria HahnNiels HansenSRIPeter KarpTomer Altman