DISCOVERYspace Tutorial

Scenario 2:

I have installed the SAGE plugin, and I have a cancer library and a normal library loaded. I want to look for possible drug targets. I would like to find upregulated putative secreted proteins.

Remember, there are multiple ways of manipulating biological data and using the application to get the answer you're looking for. Below is one method of answering the question:

Step 1: Retrieve Genes For Upregulated SAGE Tags

Bring up the 2D Contour Plot of your gene expression profile. In the application menu bar, click Tools > Expression Comparison > Select Upregulated. Change the Contour Plot to "Drag Mode" (the third button on the Contour Plot's toolbar). Right drag from the Contour Plot to the work area. A new Discovery Viewer will appear with a list of the tags.

Step 2: Map The Tags To Genes

Lets find out what genes those tags represent. Click the "Genes" button on the KDOM toolbar, and right-drag the "Human Unigene" option onto the tags displayed in the Discovery Viewer. The genes will appear in a new column in the Discovery Viewer.

Step 3: Isolate the Genes

Let's remove the genes and put them in their own Discovery Viewer. Right-drag the "Human Unigene" column header onto the work area. A new Discovery Viewer will appear listing all of the Human Unigene entries.

Step 4: Find The Proteins These Genes Represent

We need to know what proteins are translated from these genes. Click the "Proteins" button on the KDOM toolbar, and right-drag the "SWISSPROT" option onto the genes displayed in the Discovery Viewer. The proteins will appear in a new column in the Discovery Viewer.

Step 5: Isolate the Proteins

Let's remove the proteins and put them in their own Discovery Viewer. Right-drag the "SwissPROT" column header onto the work area. A new Discovery Viewer will appear listing all the SwissPROT entries.

Step 6: Get Subcellular Locatization Information

We need to know which of these proteins are probably secreted. PSORT is an algorithm that analyzes the protein sequence for motifs that reveal subcellular locatization. Click the "Functional Domains" button on the KDOM toolbar and right-drag the "PSORT" option onto the SwissPROT proteins displayed in the Discovery Viewer. The subcellular localization information for each protein will be displayed in a new column titled "PSORT".

Step 7: Get the PSORT Entry for Secreted Proteins

We could always just peruse the linked PSORT entries and look for proteins that have an appropriate relationship to a secretion-related entry. But if our list is long, this could take a while. So...

We need the PSORT entry that represents secreted proteins. If we click on the "Functional Domains" button on the KDOM toolbar and click the "PSORT" options, a dialog appears. We can search for "secret" in the "annotation" field. This is a good time to remind you to be savvy when doing keyword searches. We don't know if the annotation will use the word "secrete", "secreted", or "secretory". In this case, it's "vesicles of secretory system" - the only entry that will be found. Any protein that localizes to the vesicles of the secretory system is likely to be a secreted protein -- right? Right. This found entry will appear in a new Discovery Viewer. We can now right-drag the PSORT entry onto the Discovery Viewer showing the proteins. A popup will appear that will have an option to "Select Data With Link To...". Click this option, and any proteins that have a relationship to the PSORT entry being dragged will be selected.

You could now drag the selected entries to isolate the proteins of interest, and carry on with further interrogation.

POP QUIZ: Try using the knowledge from these scenarios to light up the found proteins on the gene expression plot.