Exploring Interactions & Pathways
Pablo Porras Millán,
IntAct
Session outline
1. An example of an interaction database: IntAct hands-on.
2. A manually-curated pathway database: Reactome hands-on.
EMBL-EBI
IntAct:
The molecular interactions database
at the EBI
www.ebi.ac.uk/intact
IntAct – Home Page
EMBL-EBI
IntAct webpage-based search
EMBL-EBI
IntAct webpage-based search
Details of
interaction
Choice of UniProtKB
or Dasty View
EMBL-EBI
IntAct: changing the layout
EMBL-EBI
IntAct: download formats
EMBL-EBI
Interaction detail in IntAct
EMBL-EBI
Detailed participant information:
Dasty view
EMBL-EBI
IntAct: filtering results
EMBL-EBI
IntAct: visualizing results
as a network
EMBL-EBI
IntAct: using lists
EMBL-EBI
IntAct: browse menu
EMBL-EBI
IntAct: advanced search
...
EMBL-EBI
IntAct: MIQL syntax search
EMBL-EBI
Reactome tutorial
Reactome Tools
• Interactive Pathway Browser
• Pathway Mapping and Over-representation
• Expression overlay onto pathways
• Molecular Interaction overlay
• Biomart
EMBL-EBI
Front Page
http://www.reactome.org
Sidebar
Navigation bar
Main
text
EMBL-EBI
Exercise 1
…see the Tutorial handouts
EMBL-EBI
The Pathway Browser
Species selector
Diagram Key
Sidebar
Zoom/move
toolbar
Pathway Diagram Panel
Details Panel (hidden)
Thumbnail
EMBL-EBI
Pathways tab – pathway hierarchy
Pathway
Reaction
Black-box
EMBL-EBI
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.
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.
EMBL-EBI
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
EMBL-EBI
Exercise 3
…see the Tutorial handouts
EMBL-EBI
Navigating in the Pathway Browser I
Home and Analyze buttons
Click here
Highlights
Details here
Click here to open pathway diagram...
EMBL-EBI
Navigating in the Pathway Browser II
Click here
Zoom
Highlights
Details here
EMBL-EBI
Exercise 4
…see the Tutorial handouts
EMBL-EBI
The Details Panel
EMBL-EBI
Exercise 5
…see the Tutorial handouts
1. Find the reaction ‘Activated type I receptor phosphorylates SMAD2/3
directly’. What pathway does it belong to?
2. In which cellular compartment does this reaction take place?
3. What is the associated GO molecular function?
4. What references verify this reaction?
5. Is this reaction predicted to occur in Canis familiaris? In
Saccharomyces cerevisiae?
EMBL-EBI
Pathway Analysis
EMBL-EBI
Pathway Analysis – Overrepresentation
P-val
Reveal next level
‘Top-level’
EMBL-EBI
Exercise 6
Use data file from earlier
exercises…
Copy, and paste into submissions
box…
Check this!
1. What is the top ‘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 ‘Signal transduction’ higher in the list
than ‘DNA repair’, when the latter has a more significant probability
score? (Hint - use the Open All button)
EMBL-EBI
Species Comparison I
EMBL-EBI
Species Comparison II
Yellow = human/rat
Blue = human only
Grey = not relevant
Black = Complex
EMBL-EBI
Exercise 7
…see the Tutorial handouts
Launch Species Comparison and select the species Rattus
norvegicus. When the results are displayed, open the pathway
‘Membrane trafficking’.
1. Find VAMP7 (top right of diagram) - what colour is it and why?
2. Which other species are inferred to have this protein? Hint: You
can answer this question without re-running Species comparison.
3. Find CHMP7 (top middle) - why is it blue?
4. 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 Rattus norvegicus?
5. Why is ADP grey?
EMBL-EBI
Expression Analysis I
EMBL-EBI
Expression Analysis II
‘Hot’ = high
‘Cold’ = low
Step through
Data columns
EMBL-EBI
Exercise 8
Launch Expression Analysis and load the example dataset. Click
‘Analyse’. When the results are displayed, find the pathway ‘DNA
replication’.
1. How many proteins are in this pathway?
2. 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.
3. Which protein has the greatest change of expression?
4. Find the complex ‘Orc4:Orc5:Orc3:Orc2:origin’ (top left of the
diagram). Which component of the complex has the lowest
expression at 24h?
5. What was the probe ID used to measure expression of this
component?
EMBL-EBI
Molecular Interaction Overlay
EMBL-EBI
Exercise 9
…see the Tutorial handouts
Open the Pathway Diagram for ‘Netrin-1 Signaling’.
1. Find the protein PTPN11 (top left of the plasma membrane). Right
click on it and select Display Interactors. How many are there?
2. How many times has the interaction between PTPN11 and
A7MD36 (UniProtKb for GAB2) been documented? Hint: This
detail is not in Reactome.
3. 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?
4. What is the easiest way to remove interactors?
EMBL-EBI
BioMart – selecting your dataset
EMBL-EBI
BioMart – filters
EMBL-EBI
BioMart – attributes
Check to get
attribute
EMBL-EBI
BioMart – results
EMBL-EBI
Exercise 10
EMBL-EBI
The Reactome Pathways Portal (Beta)
EMBL-EBI
Protein structures from PDBe
EMBL-EBI
Chemical structures from ChEBI
EMBL-EBI
Expression data from the Expression Atlas
EMBL-EBI
Interaction Data from IntAct
EMBL-EBI
More about Reactome
Reactome User's Guide
http://wiki.reactome.org/index.php/Usersguide
EMBL-EBI
Websites & Tools
• IntAct
– www.ebi.ac.uk/intact
• PSICQUIC view
– www.ebi.ac.uk/Tools/webservices/psicquic/view
• Reactome
– www.reactome.org
EMBL-EBI
Acknowledgements
Group leader
Supported by the European Community's
Seventh Framework Programme
(FP7/2007-2013) under grant agreement for
Affinomics (FP7-241481).
Henning Hermjakob
Coordinator
Sandra
Orchard
Curation team
Margaret
Duesbury
Birgit
Meldal
Steve Jupe
Antonio
Fabregat
Bijay Jassal
Phani
Garapati
David Croft
Developing team
Rafael
Jiménez
Marine
Dumousseau
Noemí
del Toro
EMBL-EBI