A Database of human biological pathways Steve Jupe

1. A Database of human biological pathways Steve Jupe

2. 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?

3. 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.

4. 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

5. 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

6. Using model organism data to build pathways – Inferred pathway events Direct evidence PMID:5555 PMID:4444 Direct evidence human PMID:8976 mouse Indirect evidence PMID:1234 cow

7. BINDING DEGRADATION DISSOCIATION DEPHOSPHORYLATION PHOSPHORYLATION CLASSIC BIOCHEMICAL TRANSPORT Theory - Reactions Pathway steps = the “units” of Reactome = events in biology

8. Reaction Example 1: Enzymatic

9. Reaction Example 2: Transport Transport of Ca++ from platelet dense tubular system to cytoplasm REACT_945.4

10. Other Reaction Types Dimerization Binding Phosphorylation

11. Reactions Connect into Pathways CATALYST CATALYST CATALYST INPUT OUTPUT INPUT OUTPUT OUTPUT INPUT

12. Evidence Tracking – Inferred Reactions Direct evidence PMID:5555 PMID:4444 Human pathway Indirect evidence PMID:8976 mouse PMID:1234 cow

13. 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

14. Species Selection

15. Data Expansion – Projecting to Other Species Human B A A + ATP -P + ADP Mouse B A A + ATP + ADP -P Drosophila Reaction not inferred B + ATP A No orthologue - Protein not inferred

16. 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

17. Coverage – Content, TOC And many more...

18. Planned Coverage – Editorial Calendar

19. Reactome Tools • Interactive Pathway Browser • Pathway Mapping and Over-representation • Expression overlay onto pathways • Molecular Interaction overlay • Biomart

20. Tutorial

21. Front Page http://www.reactome.org Navigation bar Sidebar Main text

22. Exercise 1

23. Zoom/move toolbar Thumbnail Species selector The Pathway Browser Diagram Key Sidebar Pathway Diagram Panel Details Panel (hidden)

24. Pathways tab – pathway hierarchy Pathway Reaction Black-box

25. 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. How many sub-pathways does this pathway have? How many reactions are in the first of these sub-pathways? 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.

26. 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 Binding Dissociation Omitted Uncertain

27. Exercise 3

28. Navigating in the Pathway Browser I Home and Analyze buttons Click here Highlights Details here Click here to open pathway diagram...

29. Navigating in the Pathway Browser II Highlights Details here Click here Zoom

30. Exercise 4

31. The Details Panel

32. Exercise 5

33. Pathway Analysis

34. Pathway Analysis – Overrepresentation ‘Top-level’ Reveal next level P-val

35. Exercise 6 Check this! What is the most significantly over-represented top-level pathway for this dataset? How many genes are in this pathway, and how many were represented in the dataset? 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) Can you interpret these results in terms of the underlying biology? (Hint: good luck, there are many correct answers!)

36. Species Comparison I

37. Species Comparison II Yellow = human/rat Blue = human only Grey = not relevant Black = Complex

38. Exercise 7

39. Expression Analysis I

40. ‘Hot’ = high ‘Cold’ = low Expression Analysis II Step through Data columns

41. Exercise 8

42. Molecular Interaction Overlay

43. Exercise 9

44. BioMart – selecting your dataset

45. BioMart – filters

46. BioMart – attributes Check to get attribute

47. BioMart – results

48. Exercise 10

49. The End