Exploring Interactions & Pathways

1. Exploring Interactions & Pathways Pablo PorrasMillán, IntAct

2. Session outline An example of an interaction database: IntActhands-on. A manually-curated pathway database:Reactome hands-on.

3. IntAct: The molecular interactions database at the EBI

4. IntAct – Home Page www.ebi.ac.uk/intact

5. IntAct webpage-based search

6. IntAct webpage-based search Choice of UniProtKB or Dasty View Details of interaction

7. IntAct: changing the layout

8. IntAct: download formats

9. Interaction detail in IntAct

10. Detailed participant information: Dasty view

11. IntAct: filtering results

12. IntAct: visualizing results as a network

13. IntAct: using lists

14. IntAct: browse menu

15. IntAct: advanced search ...

16. IntAct: MIQL syntax search

17. Reactome tutorial

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

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

20. Exercise 1 …see the Tutorial handouts

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

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

23. Exercise 2 …see the Tutorial handouts From the homepage, search for ‘PDGF 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 ‘Processing of classical PDGFs’? Hint: If it’s not visible, open the Details pane at the bottom of the page by clicking on the blue triangle.

24. 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

25. Exercise 3 …see the Tutorial handouts

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

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

28. Exercise 4 …see the Tutorial handouts

29. The Details Panel

30. Exercise 5 …see the Tutorial handouts • Find the reaction ‘Activated type I receptor phosphorylates SMAD2/3 directly’. What pathway does it belong to? • In which cellular compartment does this reaction take place? • What is the associated GO molecular function? • What references verify this reaction? • Is this reaction predicted to occur in Canisfamiliaris? In Saccharomyces cerevisiae?

31. Pathway Analysis

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

33. Exercise 6 Use data file from earlier exercises… Copy, and paste into submissions box… Check this! • What is the top ‘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 ‘Signal transduction’ higher in the list than ‘DNA repair’, when the latter has a more significant probability score? (Hint - use the Open All button)

34. Species Comparison I

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

36. Exercise 7 …see the Tutorial handouts • Launch Species Comparison and select the species Rattusnorvegicus. When the results are displayed, open the pathway ‘Membrane trafficking’. • Find VAMP7 (top right of diagram) - what colour is it and why? • Which other species are inferred to have this protein? Hint: You can answer this question without re-running Species comparison. • Find CHMP7 (top middle) - why is it blue? • Find AP-1 Complex (top middle, a bit to the right, below CHMP7) - Why is it black? How many proteins contribute to this object? Are they all predicted to exist in Rattusnorvegicus? • Why is ADP grey?

37. Expression Analysis I

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

39. Exercise 8 • Launch Expression Analysis and load the example dataset. Click ‘Analyse’. When the results are displayed, find the pathway ‘DNA replication’. • How many proteins are in this pathway? • How many had expression data? • Click on the View button to see this pathway in the Pathway Browser and then go to the ‘Regulation of DNA replication’ subpathway. Use the Experiment Browser toolbar to cycle through the timepoints. • Which protein has the greatest change of expression? • Find the complex ‘Orc4:Orc5:Orc3:Orc2:origin’ (top left of the diagram). Which component of the complex has the lowest expression at 24h? • What was the probe ID used to measure expression of this component?

40. Molecular Interaction Overlay

41. Exercise 9 …see the Tutorial handouts • Open the Pathway Diagram for ‘Netrin-1 Signaling’. • Find the protein PTPN11 (top left of the plasma membrane). Right click on it and select Display Interactors. How many are there? • How many times has the interaction between PTPN11 and A7MD36 (UniProtKb for GAB2) been documented? Hint: This detail is not in Reactome. • Find the protein SRC (just to the right of PTPN11). Display interactors for this protein. How many are there? Can you get a list of them? • What is the easiest way to remove interactors?

42. BioMart – selecting your dataset

43. BioMart – filters

44. BioMart – attributes Check to get attribute

45. BioMart – results

46. Exercise 10

47. The Reactome Pathways Portal (Beta) http://www.reactome.org/

48. Protein structures from PDBe

49. Chemical structures from ChEBI

50. Expression data from the Expression Atlas