June A Database of human biological pathways Steve Jupe -
June 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 membrane- bound 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?
June Nature Oct 12;407(6805): The biochemistry of apoptosis. Rationale - Figures A picture paints a thousand words… but…. Just pixels Omits key details Assumes Fact or Hypothesis?
June 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
June Extensively cross-referenced Tools for data analysis – Pathway Analysis, Expression Overlay, Species Comparison, Biomart… Used to infer orthologous events in 20 non-human species Reactome is…
June human PMID:5555PMID:4444 mouse cow Direct evidence Indirect evidence PMID:8976 PMID:1234 Using model organism data to build pathways – Inferred pathway events
June Theory - Reactions Pathway steps = the “units” of Reactome = events in biology TRANSPORT CLASSIC BIOCHEMICAL BINDING DISSOCIATION DEGRADATION PHOSPHORYLATION DEPHOSPHORYLATION
June Reaction Example 1: Enzymatic
June Reaction Example 2: Transport REACT_945.4 Transport of Ca++ from platelet dense tubular system to cytoplasm
June Other Reaction Types Binding Dimerization Phosphorylation
June Reactions Connect into Pathways OUTPUT INPUT CATALYST OUTPUT INPUT CATALYST INPUT OUTPUT CATALYST
June Human pathway PMID:5555PMID:4444 mouse cow Direct evidence Indirect evidence PMID:8976 PMID:1234 Evidence Tracking – Inferred Reactions
June 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
June Species Selection
June Data Expansion – Projecting to Other Species A + ATP A + ADP -P B Human A + ATP A + ADP -P B Mouse B A Drosophila Reaction not inferred No orthologue - Protein not inferred + ATP
June 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
June Coverage – Content, TOC And many more...
June Planned Coverage – Editorial Calendar
June Reactome Tools Interactive Pathway Browser Pathway Mapping and Over-representation Expression overlay onto pathways Molecular Interaction overlay Biomart
June Tutorial
June Front Page Sidebar Main text Navigation bar
June Exercise 1
June The Pathway Browser Species selector Diagram Key Sidebar Pathway Diagram Panel Details Panel (hidden) Zoom/move toolbar Thumbnail
June Pathways tab – pathway hierarchy Pathway Reaction Black-box
June 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.
June 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. Catalyst Input Outputs Compartment Reaction node Transition Binding Dissociation Omitted Uncertain Regulation +ve -ve
June Exercise 3
June Navigating in the Pathway Browser I Click here Highlights Details here Click here to open pathway diagram... Home and Analyze buttons
June Details here Navigating in the Pathway Browser II Click hereZoomHighlights
June Exercise 4
June The Details Panel
June Exercise 5
June Pathway Analysis
June Pathway Analysis – Overrepresentation ‘Top-level’ Reveal next level P-val
June 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!)
June Species Comparison I
June Species Comparison II Yellow = human/rat Blue = human only Grey = not relevant Black = Complex
June Exercise 7
June Expression Analysis I
June Expression Analysis II ‘Hot’ = high ‘Cold’ = low Step through Data columns
June Exercise 8
June Molecular Interaction Overlay
June Exercise 9
June BioMart – selecting your dataset
June BioMart – filters
June BioMart – attributes Check to get attribute
June BioMart – results
June Exercise 10
June The End