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A Database of human biological pathways Steve Jupe - [email protected] 1 July 2011 www.reactome.org Rationale – Journal information Nature 407(6805):770-6.The Biochemistry of Apoptosis. “Caspase-8 is the key initiator caspase in the death-receptor pathway. Upon ligand binding, death receptors such as CD95 (Apo-1/Fas) aggregate and form membranebound signalling complexes (Box 3). These complexes then recruit, through adapter proteins, several molecules of procaspase-8, resulting in a high local concentration of zymogen. The induced proximity model posits that under these crowded conditions, the low intrinsic protease activity of procaspase-8 (ref. 20) is sufficient to allow the various proenzyme molecules to mutually cleave and activate each other (Box 2). A similar mechanism of action has been proposed to mediate the activation of several other caspases, including caspase-2 and the nematode caspase CED-3 (ref. 21).” How can I access the pathway described here and reuse it? 2 July 2011 www.reactome.org Rationale - Figures A picture paints a thousand words… but…. • Just pixels • Omits key details • Assumes • Fact or Hypothesis? Nature. 2000 Oct 12;407(6805):770-6. The biochemistry of apoptosis. 3 July 2011 www.reactome.org Reactome is… Free, online, open-source curated database of pathways and reactions in human biology Authored by expert biologists, maintained by Reactome editorial staff (curators) Mapped to cellular compartment 4 July 2011 www.reactome.org Reactome is… Extensively cross-referenced Tools for data analysis – Pathway Analysis, Expression Overlay, Species Comparison, Biomart… Used to infer orthologous events in 20 non-human species 5 July 2011 www.reactome.org Using model organism data to build pathways – Inferred pathway events PMID:5555 Direct evidence PMID:4444 Direct evidence human PMID:8976 mouse Indirect evidence PMID:1234 cow 6 July 2011 www.reactome.org Theory - Reactions Pathway steps = the “units” of Reactome = events in biology BINDING DEGRADATION DISSOCIATION DEPHOSPHORYLATION PHOSPHORYLATION CLASSIC TRANSPORT 7 July 2011 BIOCHEMICAL www.reactome.org Reaction Example 1: Enzymatic 8 July 2011 www.reactome.org Reaction Example 2: Transport Transport of Ca++ from platelet dense tubular system to cytoplasm REACT_945.4 9 July 2011 www.reactome.org Other Reaction Types Dimerization Binding Phosphorylation 10 July 2011 www.reactome.org Reactions Connect into Pathways CATALYST CATALYST CATALYST INPUT OUTPUT INPUT OUTPUT INPUT 11 July 2011 www.reactome.org OUTPUT Evidence Tracking – Inferred Reactions Direct evidence PMID:5555 PMID:4444 Human pathway PMID:8976 Indirect evidence mouse PMID:1234 cow 12 July 2011 www.reactome.org Data Expansion - Link-outs From Reactome • GO • Molecular Function • Compartment • Biological process • KEGG, ChEBI – small molecules • UniProt – proteins • Sequence dbs – Ensembl, OMIM, Entrez Gene, RefSeq, HapMap, UCSC, KEGG Gene • PubMed references – literature evidence for events 13 July 2011 www.reactome.org Species Selection 14 July 2011 www.reactome.org Data Expansion – Projecting to Other Species Human B A + ATP A -P + ADP Mouse B A A -P + ADP + ATP Drosophila A + ATP B No orthologue - Protein not inferred 15 July 2011 www.reactome.org Reaction not inferred Exportable Protein-Protein Interactions Inferred from complexes and reactions Interactions between proteins in the same complex, reaction, or adjoining reaction Lists available from Downloads See Readme document for more details 16 July 2011 www.reactome.org Coverage – Content, TOC And many more... 17 July 2011 www.reactome.org Planned Coverage – Editorial Calendar 18 July 2011 www.reactome.org Reactome Tools • Interactive Pathway Browser • Pathway Mapping and Over-representation • Expression overlay onto pathways • Molecular Interaction overlay • Biomart 19 July 2011 www.reactome.org Tutorial 20 July 2011 www.reactome.org Front Page http://www.reactome.org Sidebar Navigation bar 21 July 2011 Main text www.reactome.org Exercise 1 22 July 2011 www.reactome.org The Pathway Browser Species selector Diagram Key Sidebar Zoom/move toolbar Pathway Diagram Panel Details Panel (hidden) Thumbnail 23 July 2011 www.reactome.org Pathways tab – pathway hierarchy Pathway Reaction Black-box 24 July 2011 www.reactome.org Exercise 2 From the homepage, search for ‘Notch signaling’. Click on the top pathway hit. This will open it in the Pathway Browser. Ignoring the diagram for now, look at the Pathways tab on the left. 1. How many sub-pathways does this pathway have? 2. How many reactions are in the first of these sub-pathways? 3. What reaction follows Notch 2 precursor transport to Golgi? Hint: If it’s not visible, open the Details pane at the bottom of the page by clicking on the blue triangle. 25 July 2011 www.reactome.org The Pathway Browser - Pathway Diagrams Boxes are proteins, protein sets, mixed sets or complexes. Ovals are small molecules (or sets of) Green boxes are proteins or sets, blue are complexes. Regulation +ve -ve Input Reaction node Catalyst Outputs Compartment Transition 26 July 2011 Binding Dissociation Omitted www.reactome.org Uncertain Exercise 3 27 July 2011 www.reactome.org Navigating in the Pathway Browser I Home and Analyze buttons Click here Highlights Details here Click here to open pathway diagram... 28 July 2011 www.reactome.org Navigating in the Pathway Browser II Click here Highlights Zoom Details here 29 July 2011 www.reactome.org Exercise 4 30 July 2011 www.reactome.org The Details Panel 31 July 2011 www.reactome.org Exercise 5 32 July 2011 www.reactome.org Pathway Analysis 33 July 2011 www.reactome.org Pathway Analysis – Overrepresentation P-val Reveal next level ‘Top-level’ 2011 34 July www.reactome.org Exercise 6 Check this! 1. What is the most significantly over-represented top-level pathway for this dataset? 2. How many genes are in this pathway, and how many were represented in the dataset? 3. Why is the top-level pathway Chromosome Maintenance higher in the list than Signalling by Wnt when the latter has a more significant probability score? (Hint – use the Open All button) 4. Can you interpret these results in terms of the underlying biology? (Hint: good luck, there are many correct answers!) 35 July 2011 www.reactome.org Species Comparison I 36 July 2011 www.reactome.org Species Comparison II Yellow = human/rat Blue = human only Grey = not relevant Black = Complex 37 July 2011 www.reactome.org Exercise 7 38 July 2011 www.reactome.org Expression Analysis I 39 July 2011 www.reactome.org Expression Analysis II Step through Data columns 40 July 2011 www.reactome.org ‘Hot’ = high ‘Cold’ = low Exercise 8 41 July 2011 www.reactome.org Molecular Interaction Overlay 42 July 2011 www.reactome.org Exercise 9 43 July 2011 www.reactome.org BioMart – selecting your dataset 44 July 2011 www.reactome.org BioMart – filters 45 July 2011 www.reactome.org BioMart – attributes Check to get attribute 46 July 2011 www.reactome.org BioMart – results 47 July 2011 www.reactome.org Exercise 10 48 July 2011 www.reactome.org The End 49 July 2011 www.reactome.org