1. Gene function analysis Stem Cell Network Microarray Course, Unit 5 May 2007

2. Sections Introduction to Gene Ontology GOstat Example

3. Gene Ontology Michael Ashburner Annotate genes or proteins http://www.geneontology.org Started for Drosophila melanogaster (fly). Now expanded for all taxa

4. Gene Ontology Biological process A phenomenon marked by changes that lead to a particular result, mediated by one or more gene products. Molecular function Elemental activities, such as catalysis or binding, describing the actions of a gene product at the molecular level. A given gene product may exhibit one or more molecular functions. Cellular component The part of a cell of which a gene product is a component; for purpose of GO includes the extracellular environment of cells; a gene product may be a component of one or more parts of a cell; this term includes gene products that are parts of macromolecular complexes, by the definition that all members of a complex normally copurify under all except extreme conditions.

5. Gene Ontology http://www.geneontology.org/GO_nature_genetics_2000.pdf Biological process

6. Gene Ontology http://www.geneontology.org/GO_nature_genetics_2000.pdf

7. Gene Ontology http://www.geneontology.org/GO_nature_genetics_2000.pdf

8. Gene Ontology Evidence codes http://www.geneontology.org/GO.evidence.shtml IC: Inferred by Curator IDA: Inferred from Direct Assay IEA: Inferred from Electronic Annotation IEP: Inferred from Expression Pattern (2006) IGC: Inferred from Genomic Context (2007) IGI: Inferred from Genetic Interaction IMP: Inferred from Mutant Phenotype IPI: Inferred from Physical Interaction ISS: Inferred from Sequence or Structural Similarity NAS: Non-traceable Author Statement (2006) ND: No biological Data available RCA: inferred from Reviewed Computational Analysis TAS: Traceable Author Statement NR: Not Recorded (2006)

9. Gene Ontology Stats. May 29 th 2007. biological_process: 13,553 terms (10,894 in 2006; 9,277 in 2005) cellular_component: 1,966 terms (1,815; 1,512) molecular_function: 7,609 terms (7,927; 6,957), Total: 23,128 terms (20,636; 17,746)

10. Gene Ontology Stats. May 29 th 2007. Mouse Genome Informatics (The Jackson Laboratory http://www.informatics.jax.org/) biological_process: 14,200 genes, 42,675 annotations (3.0 kw/gene) [13,329 genes, 33,783 annotations (2.5 kw/gene) in 2006] cellular_component: 14,713 genes, 31,330 annotations (2.1 kw/gene) [13,547 genes, 26,515 annotations (2.0 kw/gene)] molecular_function: 15,553 genes, 50,343 annotations (3.2 kw/gene) [14,056 genes, 40,806 annotations (2.9 kw/gene)] 8.3 terms per gene [ 7.5 in 2006]

11. Databases using Gene Ontology NetAffx (Affymetrix probe annotations) Flybase (sequences) was the first SGD (yeast) MGI (mouse) InterPro (Protein sequences) ProDom (Protein domains) Entrez Gene (gene information)

12. GOstat Find statistically overrepresented properties within a group of genes as selected by......typically, analysis of a DNA microarray experiment http://gostat.wehi.edu.au/ Beissbarth & Speed (2004) Bioinformatics, 20: 1464-1465.

13. gene A gene B gene C gene D gene E XXXX Total set of genes 2,000 of 5,000 are X Not significant YYYY Total set of genes 4 of 5000 are Y Very significant Do it for all Gene Ontology terms Take into account the structure of the ontology Sort by p-values GOstat

14. Contigency Table genes with GO in group total genes in group selected genes (e.g. differentially expressed) reference group (e.g. all genes on array) 51 176 467 9180 p-value 8e-52 Chi-square Test (Fisher's Exact Test for small values) Probability of obtaining those values from a random distribution.

15. Web tool

17. Output

18. Example We will study the function of a set of genes selected via StemBase http://www.stembase.ca/ (see corresponding Unit for more info on using StemBase) http://gostat.wehi.edu.au/

19. 1. Select a set of genes Objective: Genes correlated to Lgals3bp (lectin, galactoside-binding, soluble, 3 binding protein) A galectin, a beta-galactoside-binding protein implicated in modulating cell- cell and cell-matrix interactions

20. 1. Select a set of genes

25. 2. Run in GOstat

26. Calcium ion binding mannosyl-oligosaccharide mannosidase activity

27. 2. Run in GOstat http://www.geneontology.org/amigo

28. 2. Run in GOstat Calcium ion binding mannosyl-oligosaccharide mannosidase activity

29. 2. Run in GOstat

30. 3. Examine expression MAN2A11448647_at MAN1A1417111_at Lgals3bp1448380_at

31. 3. Examine expression

33. To know more Gene Ontology. http://www.geneontology.org/GO.doc.shtm GOstat http://gostat.wehi.edu.au Beissbarth & Speed (2004) Bioinformatics, 20: 1464-1465. StemBase. http://www.stembase.ca See corresponding Unit in this course.