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G:Profiler

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angelauzelac10 edited this page Mar 25, 2023 · 5 revisions

Objective

  • become familiar with annotation source g:profiler

Duration

Expected: 1 hr

Actual: 1 hr

Task

Use this list of genes:genelist.txt as your query set and run a g:profiler enrichment analysis with the following parameters:

Data sources : Reactome, Go biological process, and Wiki pathways Multiple hypothesis testing - Benjamini hochberg

Procedure

  • copied and pasted the gene list from https://github.com/bcb420-2020/Student_Wiki/blob/master/genelist.txt
  • chose the specified parameters (why did we not choose no electronic annotations?)
  • ran query
  • chose Select Ensembl ID with most GO annotations
  • for those that had no GO annotations chose the top one because it wouldn't select it automatically
  • Clicked the Detailed Results tab to answer question 1
  • in Detailed Results tab, clicked on arrows next to stats heading which opened up more columns
    • T: number of genes in a term
    • Q: number of genes in my query
    • T intersect Q: number of genes in both
  • changed the term size slider to between 5 and 200 to answer 4

Results

1. What is the top term returned in each data source?

GO bio process: Immune System Process

Reactome: Immune System

WikiPathways: Allograft rejection

2. How many genes are in each of the above genesets returned? (hint, in the Detailed results tab of g:profiler results if you click on the arrows next to the stats heading you will be able to see the number of genes in a term, number of genes in your query and number of genes in your query that are also in your term)

Is geneset interchangeable with pathway? I think I should be looking at the number of genes in a term (T)?

GO bio process: Immune System Process

  • 2683

Reactome: Immune System

  • 2041

WikiPathways: Allograft rejection

  • 88

3. How many genes from our query are found in the above genesets?

I think I should be looking at the number of genes in my query that are also in my term (T intersect Q)?

GO bio process: Immune System Process

  • 290

Reactome: Immune System

  • 220

WikiPathways: Allograft rejection

  • 32

4. Change g:profiler settings so that you limit the size of the returned genesets. Make sure the returned genesets are between 5 and 200 genes in size. Did that change the results?

Yes the results changed. The top term name changed for GO BP and Reactome. They are now:

GO bio process: antigen processing and presentation

Reactome: Immunoregulatory interactions between a Lymphoid and a non-Lymphoid cell

Also, there are far less genes in each term/geneset, and less genes from our query that are in these genesets. These results go for all of the datasources.

5. Which of the 4 ovarian cancer expression subtypes do you think this list represents?

It is Immunoreactive ovarian cancer.

What is immonureactivity?

A measure of the immune reaction caused by an antigen (Immunoreactivity, n.d.).

Immunoreactivity is a word pathologists use to describe the results of a test called immunohistochemistry. This test is performed to see if the cells in a tissue sample make a protein of interest. Cells that make the protein may be described as reactive or positive. Cells that do not produce the protein may be described as non-reactive or negative (Immunoreactivity, n.d.).

Bonus: The top gene returned for this comparison is TFEC (ensembl gene id:ENSG00000105967). Is it found annotated in any of the pathways returned by g:profiler for our query? What terms is it associated with in g:profiler?

No annotations in any of the terms in any of the data sources after filtering.

Before filtering is associated with response to stimulus, response to stress, cellular response to stimulus - all in GO BP.

Conclusion and Outlook

  • g:profiler is a great annotation source to use because it uses many genesets like GO, Reactome, and KEGG
  • has a simple, user-friendly web interface where we can visualize GO, pathway, of transcription factor binding site enrichments
  • allows for multiple testing corrections
  • can interpret ranked gene lists, has efficient algorithm
  • updated its data regularly, and gets many annotations from Ensembl (Reimand et al., 2007)

References

Immunoreactivity - MyPathologyReport.ca. (n.d.). My Pathology Report. Retrieved March 11, 2023, from https://www.mypathologyreport.ca/pathology-dictionary/immunoreactivity/

Immunoreactivity. (n.d.). Wiktionary. Retrieved March 11, 2023, from https://en.wiktionary.org/wiki/immunoreactivity

Reimand, J., Kull, M., Peterson, H., Hansen, J., & Vilo, J. (2007). g:Profiler--a web-based toolset for functional profiling of gene lists from large-scale experiments. Nucleic acids research, 35(Web Server issue), W193–W200. https://doi.org/10.1093/nar/gkm226

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