Tutorial:Extend Biological Process with regulatory interactions

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Extend biological process with regulatory interactions

In this tutorial we use four apps:

  • WikiPathways-App: search and download pathways from WikiPathways in pathway or network view (network = unique nodes, no pathway layout).
  • NetworkAnalyzer: find the hubs in a network by analyzing the node degree and betweenness properties in a network.
  • PathExplorer: find all path that go to or from a specific node.
  • CyTargetLinker: extend the network with regulatory interactions to better understand the regulation of a biological process.

Note: In this exercise we are going to use the human Fatty Acid Beta Oxidation pathway. We want to use the human ENCODE transcription factor data, so we chose to use the human instead of the rat pathway.

The original tutorial is available here.

Download pathway from WikiPathways

  • Go to Import → Network → Public Databases and select WikiPathways as the data source.
  • Search for "Fatty acid beta oxidation" in Only Homo sapiens (see figure below). Please select the Import as network option.
  • The resulting network contains 143 nodes and 198 edges (see figure below).

Fig 3a: WikiPathways webservice import Fig 3c: Pathway shown as network in Cytoscape

Analyzing the properties of the network

  • Go to Tools → Network Analyzer → Network Analysis → Analyze network. Analyze the network as an undirected network. You can have a closer look at the different properties, like node degree distribution or betweenness.
  • In the end you can visualize the properties on the network to identify hub nodes. Click on Visualize Parameters and select map node size to Degree and map node color to Betweenness (see left figure below).
  • The resulting network identifies the two genes encoding the subunits of the mitochondrial trifunctional protein (HADHA and HADHB), which catalyzes the last three steps of mitochondrial beta-oxidation of long chain fatty acids, as two of the hub proteins in the network (see right figure below).

Fig 3d: Visualize parameters Fig 3e: Find hub nodes in resulting network

Find all path in the network that lead to TCA cycle

  • In the pathway diagram on WikiPathways you can see that the pathway has several end points going into the TCA cycle. With the PathExplorer app, we are going to have a closer look what this means from a network perspective. Please be aware that the group and complex nodes are not ideal for this analysis (the WikiPathways App will be extended soon to support the visualization of a pathway without those group nodes).
  • Find the TCA Cycle node in the network and right click on the node. Select PathExplorer → Find paths to here (see left figure below).

The app highlights all paths within the network that result in this node. You can immediately see that nearly all path end up in the TCA cycle (see right figure below).

Fig 3f: Use PathExplorer to find paths to TCA cycle Fig 3g: Highlighted paths to TCA cycle

Extending the network with TF and microRNA regulators

  • Go to Apps → CyTargetLinker → Extend network. Select the network, choose GeneID as the attribute containing a biological identifier, specify the directory containing the RINs (they are available on the USB sticks) and choose "Add Regulators" as the direction.

Fig 3h: CyTargetLinker dialog

  • Select all RINs that are in the directory on the USB stick. The extension might take a while – check status in the status bar in the bottom left corner.

Fig 3i: RIN selection

  • CyTargetLinker extends the network with 1930 microRNA-target interactions from various databases and 58 transcription factor-target gene interactions from ENCODE.

Fig 3j: CyTargetLinker legend Fig 3k: Extended network

  • When changing the overlap threshold in the CyTargetLinker control panel (left side) to 2, only interactions supported by at least 2 RINs are shown. That allows us to e.g. identify two microRNAs that target several genes in the pathway, hsa-miR-124 and hsa-miR-506.

Fig 3l: Threshold overlap = 2