using pathway-tools for phenotype- directed curation jeremy zucker broad institute of mit and...
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Using Pathway-tools for phenotype-directed curation
Jeremy Zucker
Broad Institute of MIT and Harvard
Boston University
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GENOME SEQUENCE TO METABOLIC RECONSTRUCTION
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Genome sequence to metabolic modelPathways Literature
Nutrient media(Vogels)
NeurosporaCyc
Elements Metadata
Complexes
Reactions
Transporters
Biomass composition
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EFICAz2 predicts enzymes
…
Dec
isio
n tr
ee
Databases
HMMs
FDR
SVM
9934 protein sequences
1993 enzymes1770 reactions
BMC Bioinformatics 2009, 10:107
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Protein Complex editor182 reactions with isozymes or complexes
56 complexes experimentally validated through literature search
• 2-oxoisovalerate alpha subunit• 2-oxoisovalerate beta subunit
…• fatty acid synthase betasubunit dehydratase• fatty acid synthase alpha subunit reductase
Identify multiple genes of reaction
Allow curator to validate potential complexes
2-oxoisovalerate complex
Present all possible combinations of
complexes
Fatty acid synthasecomplex
…
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Transport inference parser (TIP)9934 free-text Protein annotations
442 transport reactions
• MFS glucose transporter• ATP synthase
…• sucrose transporter
Filter proteins for transporters
Infer multimeric complex
Infer substrate
Infer energy-coupling mechanism
…
Bioinformatics (2008) 24 (13): i259-i267.
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Pathologic predicts pathways1770 enzyme-catalyzedreactions
893 reactions assigned to 265 Pathways
… …
X = #rxns in metacyc pwy
Y = #rxns with enzyme evidence
Z = #unique rxns in pwy
P(X|Y|Z) = prob of pwy in Neurospora
Science 293:2040-4, 2001.
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Literature curation validates pathways
…
523 citations associated with 265 pathways 56 complexes524 genes
…
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PHENOTYPE-DIRECTED CURATION
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Metabolic Model+
Steady State Assumption
=
Predict Rate of Biomass
Production
Nutrient Uptake
Biom
ass
Com
pone
nts
Knockout Phenotype Prediction
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Knockout Phenotype PredictionNutrient Uptake
Biom
ass
Com
pone
nts
• Simulate KO by removing reaction
• Are any biomass components blocked when applying FBA?– No: non-essential
gene– Yes: essential gene
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Why is it useful?
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Phenotype-directed curation
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461 curated minimal media viable and essential genes
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Minimal media gene essentiality training set
Predicted
Growth No-growth
Observed Growth 103 (TP) 0 (FN)
No-growth 2 (FP) 46 (TN)
Sensitivity TP/(TP+FN) 100%
Specificity TN/(TN+FP) 95.8%
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105 auxotroph supplemental rescue phenotypes
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Supplemental rescue training set
Predicted
Growth No-growth
Observed Growth 74 (TP) 3 (FN)
Sensitivity TP/(TP+FN) 96.1%
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Final metabolic reconstruction224 Pathways 523 Citations
Nutrient media(Vogels)
NeurosporaCyc
Elements Metadata
56 Complexes
1650 Reactions
442 Transporters
Biomass composition
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Independent test set validation
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Minimal media gene essentiality test set
Predicted
Growth No-growth
Observed Growth 274 (TP) 20 (FN)
No-growth 1 (FP) 15 (TN)
Viable predicted/observed (sensitivity): 93.2%Essential predicted/observed (specificity): 93.8%
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Comparison of model organisms under minimal media
Yeast 4.01 E.coli iJO13662 Neurospora
Viable Predicted/ Observed
691/772=89.5% 1079/1159=91.2% 274/294=93.2%
Essential Predicted/Observed
49/152=32.3% 168/207=81.2% 15/16=93.8%
[1] Dobson et al. BMC Systems Biology 2010, 4:145[2] Molecular Systems Biology 7 Article number: 535
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Supplemental rescue test set
Predicted
Growth No-growth
Observed Growth 23 (TP) 5 (FN)
Rescues predicted/observed (sensitivity): 82.1%
trp-1,3 rescued by tryptophannic-3 rescued by nicotinamidecys-11 rescued by sulfate, thiosulfite, methionine, cysteine
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MULTIPLE KNOCKOUTS
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Synthetic Lethality
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Pathway interpretation
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Synthetic lethal rescues computationally predicted
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Pathway interpretation
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Phenotype arrays
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AcknowledgementsBoston University
James Galagan
Jonathan Dreyfuss
Andrew Krueger
OHSU
Heather Hood
Texas A&M
Joe Sturino
Matt Sachs
UC RiversideKathy BorkovichJacqueline Servin
University of LeedsAlan Radford
SRIPeter KarpMario Latendresse
Broad InstituteChristian Stolte
Citizen ScientistLinda Ocasio