GOToolBox: functional analysis of gene datasets based on Gene Ontology. David Martin, Christine Brun, Elisabeth Remy, Pierre Mouren, Denis Thieffry, Bernard Jacq To cite this version: David Martin, Christine Brun, Elisabeth Remy, Pierre Mouren, Denis Thieffry, et al.. GOToolBox: functional analysis of gene datasets based on Gene Ontology.. Genome Biology, BioMed Central, 2004, 5, pp.R101. 10.1186/gb-2004-5-12-r101. inserm-00095249 HAL Id: inserm-00095249 https://www.hal.inserm.fr/inserm-00095249 Submitted on 15 Sep 2006 HAL is a multi-disciplinary open access L’archive ouverte pluridisciplinaire HAL, est archive for the deposit and dissemination of sci- destinée au dépôt et à la diffusion de documents entific research documents, whether they are pub- scientifiques de niveau recherche, publiés ou non, lished or not. The documents may come from émanant des établissements d’enseignement et de teaching and research institutions in France or recherche français ou étrangers, des laboratoires abroad, or from public or private research centers. publics ou privés. Software2004MartinetVolume al. 5, Issue 12, Article R101 Open Access GOToolBox: functional analysis of gene datasets based on Gene comment Ontology David Martin*, Christine Brun*, Elisabeth Remy†, Pierre Mouren*, Denis Thieffry* and Bernard Jacq* Addresses: *Laboratoire de Génétique et Physiologie du Développement, IBDM, CNRS/INSERM/Université de la Méditerranée, Parc Scientifique de Luminy, case 907, 13288 Marseille, France. †Institut de Mathématiques de Luminy, Parc Scientifique de Luminy, 13288 Marseille, France. reviews Correspondence: David Martin. E-mail: [email protected] Published: 26 November 2004 Received: 13 April 2004 Revised: 31 August 2004 Genome Biology 2004, 5:R101 Accepted: 25 October 2004 The electronic version of this article is the complete one and can be found online at http://genomebiology.com/2004/5/12/R101 reports © 2004 Martin et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. GOToolBox:<p>Toolsto find genes are functionalwith presented similar analysis to annotations.</p> identify of geGenene datasets Ontology based terms on that Gene are Ontology over- or under-represented in a dataset, to cluster genes by function and deposited research Abstract We have developed methods and tools based on the Gene Ontology (GO) resource allowing the identification of statistically over- or under-represented terms in a gene dataset; the clustering of functionally related genes within a set; and the retrieval of genes sharing annotations with a query gene. GO annotations can also be constrained to a slim hierarchy or a given level of the ontology. The source codes are available upon request, and distributed under the GPL license. refereed research refereed Rationale the description of some aspects of gene function which are Since complete genome sequences have become available, the specific to few lineages only. Within each of these ontologies, amount of annotated genes has increased dramatically. These the terms are organized in a hierarchical way, according to advances have allowed the systematic comparison of the gene parent-child relationships in a directed acyclic graph (DAG). content of different organisms, leading to the conclusion that This allows a progressive functional description, matching organisms share the majority of their genes with only rela- the current level of experimental characterization of the cor- tively few species-specific genes. On this basis, one can responding gene product. The hierarchical organization of interactions develop strategies to infer gene annotations from model spe- the gene ontology is particularly well adapted to computa- cies to less experimentally tractable organisms. However, tional processing and is used for the functional annotations of such functional inferences require the definition of species- gene products of several model organisms such as budding independent annotation policies. yeast [2], Drosophila [3], mouse [4], nematode [5] and Ara- bidopsis [6]. More recently, GO annotations for human genes In this regard, the Gene Ontology consortium [1] has been have been proposed in the context of the GOA project [7]. created to develop a unified view of gene functional annota- tions for different model organisms. Three structured vocab- In parallel, the recent development of new high-throughput information ularies (or ontologies) have been proposed, which allow the methods has generated an enormous amount of functional description of molecular functions, biological processes and data and has motivated the development of dedicated analy- cellular locations of any gene product, respectively. Whereas sis tools. For instance, one might wonder whether genes the majority of GO terms are common to several organisms, detected as being coexpressed in a DNA chip experiment are some of them are specific to a few organisms only, enabling related in terms of molecular or cellular function. In practical Genome Biology 2004, 5:R101 R101.2 Genome Biology 2004, Volume 5, Issue 12, Article R101 Martin et al. http://genomebiology.com/2004/5/12/R101 available for download on the GOToolBox server for one Gene name/ID list Gene name/ID User input week. This file contains also the counts of terms within a ref- erence gene dataset (genome or user-defined), and can then Dataset GO- Program creation Family be used as an input for the GO-Stats and GO-Proxy programs Result described below. Associated terms Functionally and parents related genes Ontology options An optional tool, GO-Diet, allows either the reduction of the term dataset to a slim GO hierarchy (either one proposed by GO-Diet the GO consortium or a user-defined one) or the restriction of Terms sorted GO-Stats by relevance the considered terms to a chosen depth of the ontology. It is Slimmed GO also possible to filter terms based on the way these have been annotation set Genes clustered assigned to the gene products (evidence code). This tool is GO-Proxy by function useful to decrease the number of GO terms associated with a gene dataset, thereby facilitating the analysis of the results of FlowchartFigure 1 of the GOToolBox programs programs described below, particularly when the input gene Flowchart of the GOToolBox programs. list and/or the number of associated GO terms is large. Note that the GO-Diet program can generate a gene-term associa- tion file in the TLF format, allowing the use of GO terms as terms, we address here the following generic questions. First, gene labels with the TreeDyn tree drawing program [9]. The are there statistically over- or under-represented GO terms GO-Diet options are proposed in the Dataset-Creation form. associated with a given gene set, compared to the distribution of these terms among the annotations of the complete GO term statistics genome? Second, among a particular gene set, are there Frequencies of terms within the dataset are calculated and closely functionally related gene subsets? And third, are there compared with reference frequencies (for example with genes having GO similarities with a given probe gene? genomic frequencies or with the frequencies of these terms in the complete list of genes spotted on an array). This proce- To formulate such questions properly in a well defined math- dure allows the delineation of enrichments or depletions of ematical framework, we have developed a set of methods and specific terms in the dataset. The probability of obtaining by tools, collectively called GOToolBox, to process the GO anno- chance a number k of annotated genes for a given term among tations for any model organism for which they are available a dataset of size n, knowing that the reference dataset con- (Figure 1). tains m such annotated genes out of N genes, is then calcu- lated. This test follows the hypergeometric distribution All the programs are written in PERL and use the CGI and described in Equation 1: DBI modules. All the ontology data and the gene-GO terms associations are taken from the GO consortium website. m Nm− These data are structured in a PostGreSQL relational data- k nk− base, which is updated monthly. Statistics are calculated P(rX{}= k= 1) N using the R statistical programming environment. The web n implementation of the GOToolBox is accessible at [8]. where the random variable X represents the number of genes within a given gene subset, annotated with a given GO term. Features Implemented in the GO-Stats tool, this formula permits the In this section, we describe the five main functionalities of the automatic ranking of all annotation terms, as well as the eval- GOToolBox suite. Two of them (GO-Proxy and GO-Family) uation of the significance of their occurrences within the data- are not encompassed by any other GO-processing tool cur- set. An illustration of such an approach is given in 'Mining rently available (see also 'Comparison of the GOToolBox with biological data'. A typical GO-Stats output is presented in Fig- other GO-based analysis programs'). ure 2. Dataset creation GO-based gene clustering The first step in analyzing gene datasets consists in retrieving, The goal of the GO-Proxy tool is to group together function- for each individual gene of the dataset, all the corresponding ally related genes on the basis of their GO terms. The rationale GO terms and their parent terms using the Dataset creation sustaining our method is that the more genes have common program. The genomic frequency of each GO term associated GO terms, and the less they have specific associated terms, with genes present in the dataset is then calculated. The the more likely they are to be functionally related. For any two resulting information is structured and stored in a data file, genes of the gene set, the program calculates an annotation- Genome Biology 2004, 5:R101 http://genomebiology.com/2004/5/12/R101 Genome Biology 2004, Volume 5, Issue 12, Article R101 Martin et al.