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Cytoscape Tips

mir-29 Family Network

Additional notes when using Cytoscape with the GOlorize and BiNGO plugins. These notes assume you are already familiar with using Cytoscape. If you haven’t used this resource yet instructions are available at http://manual.cytoscape.org/en/stable/ or attend a training course (available at EMBL-EBI and UCL)

What data is in the PSICQUIC EBI-GOA-miRNA file

Key information listed in the Cytoscape Network view ‘Edge Table’

The primary interaction type is listed as ‘physical association’ if there is experimental evidence that the microRNA binds to a mRNA. The primary interaction type is listed as ‘association’ if the microRNA is predicted to bind an mRNA and there is experimental evidence that the mRNA levels are altered by the presence or absence of the microRNA.

Key information listed in the Cytoscape Network view ‘Node Table’

The ‘name’ lists the ENSG identifiers for the gene targets of the microRNA.

How to overlay GO terms onto a network created using the using the PSICQUIC EBI-GOA-miRNA file

The GO gene association file associates GO terms with UniProt identifiers (or Model Organism Identifiers). It is therefore necessary to edit this column so that the ENSG identifiers in the ‘name’ column are changed to the equivalent UniProt Identifiers.

  1. Select all nodes and view annotation rows in ‘Node Table’.
  2. In the column ‘name’ are listed the ENSG identifiers for all the genes listed as associated with the microRNAs. 
  3. UniProt has a very good mapping tool to enable you to download the UniProt identifiers. 
  4. Edit the Node table, replacing all the ENSG identifiers with their equivalent UniProt identifiers.
  5. Now you can use BiNGO to analyse the network.

How to edit the protein IDs to remove isoform information to fully exploit the GO annotations available in the isoform gene association file

The GO gene association isoform file associates GO terms with UniProt identifiers (or Model Organism Identifiers). The isoform information included in column 17, but this information is not interpreted by BinGO. Therefore, BinGO only overlays GO terms onto the canonical non-isoform IDs. It is therefore necessary to edit the column in the network table to remove isoform information (eg -1, -2) so that the identifiers are simple UniProt Identifiers.

  1. Before merging your networks
  2. Identify all isoforms in the network 
  3. Select all nodes and view annotation rows in ‘Node Table’.
  4. In the column which lists the uniprot identifiers remove all '-#' information 
  5. Merge your networks
  6. Now you can use BiNGO to analyse the network
  7. Note that now the specific isoform variants in the network have been removed.

How to include all relevant GO annotation data in a BiNGO analysis

Download gene association files from EBI ftp site

Use Firefox browser to download files from http://www.ebi.ac.uk/GOA/downloads

There are now multiple files for each species, you need to make sure you choose the right ones to use to fully exploit the annotation data. For example there are 4 files for human: protein, isoforms, RNA and complex. If you are only looking at protein networks make sure you include both the protein and isoform files in your analysis or you will miss a lot of annotations.

Decide what GO annotation data you want to include in the BiNGO analysis and copy the relevant data into one file.

Cytoscape plugins GOlorize and BinGO will not be able to use the downloaded files with their current names for files released since June 2016 which coincides with release 158 and above. A typical file name format that will work looks like this:

gene_association.goa_human.164

where 164 is the release number.

In general , the file and its extension must follow these rules:

1. the expression "gene_association" must exist in the file name.

2. the expression "goa_human" must exist in the file extension.

Other combinations can work too. However, we have not tried them all and we can only advise that the above combination will work.

Download the ontology file go-basic.obo from the GOC ftp site

download files from http://geneontology.org/page/download-ontology

Right click to save page as txt file

How to create a GO term mapping file to map GO term IDs to GO term names

This file is very useful when working with BiNGO enrichment analysis data, if you have not managed to save the enrichment file during the analysis.

When you save the enrichment results (eg by copying the data in the output table and pasting into an excel spreadsheet) the ‘Description’ (GO term name) does not copy across.

Therefore, it can be useful to generate a flat file of the ontology and to use the VLOOKUP option in excel to import the GO term names.

1. A file of GO term Ids and equivalent GO term names can be generated by copying the GO terms from:

http://www.ebi.ac.uk/QuickGO/GSearch?format=termlist&what=Process

http://www.ebi.ac.uk/QuickGO/GSearch?format=termlist&what=Function

http://www.ebi.ac.uk/QuickGO/GSearch?format=termlist&what=Component

2. Save data to an Excel spreadsheet. 

3. Select column A and go to ‘Data’ Menu. 

4. Select the icon 'Text to Columns', and ‘Fixed width’, Next. 

5. Drag the cursor where you want to split (e.g. in front of the GO term name), ‘Next’, 'Finish'. This will produce 2 columns in Excel - one of GO IDs and one of GO names. 

6. Remove the space after each GO identifier using find>replace option in Excel. 

7. The BiNGO output table just has the GO IDs, not GO:#######. So either add GO:000… to each GO ID in your Cytoscape table, or, edit the mapping file to reduce the GO IDs to just numbers, using find>replace GO:000000, then find>replace GO:00000, etc. This file can then be used in future for other analyses.