March A Database of human biological pathways Steve Jupe -

March Background Cells contain thousands of molecules, –proteins and small chemical compounds They work together to perform tasks. –provide energy –produce materials –respond to the environment. Processes require coordinated efforts –like a factory production line. –Biologists refer to a chain of connected molecular events as a pathway. ELLS 2011

March Different types of pathways Metabolic networks – provide the energy and materials Signaling networks – sense the outside, coordinate activities within and between cells (Gene) regulatory networks – control processes, set limits, control the molecular composition of cells ELLS 2011

March The scale of pathways Early pathways were pieced together by studying biochemical reactions of individual proteins, measuring the activity, learning the substrates and products, joining them to the next enzyme in the pathway. New technologies allow simultaneous measurement of tens of thousands of different molecules. This reveals that many biological pathways are interconnected, more complicated than the typical textbook examples, with fluid boundaries. They can work together or against each other (feedback and regulation). Scientists who study or interpret these large scale interactions sometimes refer to the field as ‘systems biology’ When multiple biological pathways interact = a biological network. When all possible interactions are added, pathways become extremely difficult to understand Biologists often work with ‘canonical’ pathways, those that represent the well-understood part of the entire network. ELLS 2011

March 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?

March 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?

March 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

March Extensively cross-referenced Tools for data analysis – Pathway Analysis, Expression Overlay, Species Comparison, Biomart… Used to infer orthologous events in 20 other species Reactome is…

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

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

March Reaction Example 1: Enzymatic

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

March Other Reaction Types Binding Dimerization Phosphorylation

March Reactions Connect into Pathways OUTPUT INPUT CATALYST OUTPUT INPUT CATALYST INPUT OUTPUT CATALYST

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

March Species Selection

March 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

March Exportable Protein-Protein Interactions Inferred from complexes and reactions (more on this later) Interactions between proteins in the same complex, reaction, or adjoining reaction Lists available from Downloads See Readme document for more details

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

March Planned Coverage – Editorial Calendar

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

March Tutorial Tutorial handouts in your info packages…

March Front Page Sidebar Main text Navigation bar

March Exercise 1 …see the Tutorial handouts

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

March Pathways tab – pathway hierarchy Pathway Reaction Black-box

March Exercise 2 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. 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 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. …see the Tutorial handouts

March 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

March Exercise 3 …see the Tutorial handouts

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

March Details here Navigating in the Pathway Browser II Click hereZoomHighlights

March Exercise 4 …see the Tutorial handouts

March The Details Panel

March Exercise 5 …see the Tutorial handouts

March Pathway Analysis

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

March Exercise 6 Check this! …see the Tutorial handouts

March Species Comparison I

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

March Exercise 7 …see the Tutorial handouts

March Expression Analysis I

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

March Exercise 8

March Molecular Interaction Overlay

March Exercise 9 …see the Tutorial handouts

March New Look Browser

March End

March BioMart – selecting your dataset

March BioMart – filters

March BioMart – attributes Check to get attribute

March BioMart – results

March Exercise 10