TF Target Mapper: A BLAST search tool for the identification of Transcription Factor target genes

Department of Cell Biology, Erasmus Medical Center, Dr Molewaterplein 50, 3015GE Rotterdam, The Netherlands.
BMC Bioinformatics (Impact Factor: 2.58). 02/2006; 7(1):120. DOI: 10.1186/1471-2105-7-120
Source: DOAJ


In the current era of high throughput genomics a major challenge is the genome-wide identification of target genes for specific transcription factors. Chromatin immunoprecipitation (ChIP) allows the isolation of in vivo binding sites of transcription factors and provides a powerful tool for examining gene regulation. Crosslinked chromatin is immunoprecipitated with antibodies against specific transcription factors, thus enriching for sequences bound in vivo by these factors in the immunoprecipitated DNA. Cloning and sequencing the immunoprecipitated sequences allows identification of transcription factor target genes. Routinely, thousands of such sequenced clones are used in BLAST searches to map their exact location in the genome and the genes located in the vicinity. These genes represent potential targets of the transcription factor of interest. Such bioinformatics analysis is very laborious if performed manually and for this reason there is a need for developing bioinformatic tools to automate and facilitate it.
In order to facilitate this analysis we generated TF Target Mapper (Transcription Factor Target Mapper). TF Target Mapper is a BLAST search tool allowing rapid extraction of annotated information on genes around each hit. It combines sequence cleaning/filtering, pattern searching and BLAST searches with extraction of information on genes located around each BLAST hit and comparisons of the output list of genes or gene ontology IDs with user-implemented lists. We successfully applied and tested TF Target Mapper to analyse sequences bound in vivo by the transcription factor GATA-1. We show that TF Target Mapper efficiently extracted information on genes around ChIPed sequences, thus identifying known (e.g. alpha-globin and zeta-globin) and potentially novel GATA-1 gene targets.
TF Target Mapper is a very efficient BLAST search tool that allows the rapid extraction of annotated information on the genes around each hit. It can contribute to the comprehensive bioinformatic transcriptome/regulome analysis, by providing insight into the mechanisms of action of specific transcription factors, thus helping to elucidate the pathways these factors regulate.

Download full-text


Available from: John Strouboulis
  • Source
    • "The oligonucleotide probes used in EMSA were designed according to a computer-based search with the software Promoter Scan (PROSCAN Version 1.7 suite of programs developed by Dr. Dan Prestridge) and Transfac [11]encompassing putative binding sites in the CTNNAL1 promoter. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Adhesion molecules play important roles in airway hyperresponsiveness or airway inflammation. Our previous study indicated catenin alpha-like 1 (CTNNAL1), an alpha-catenin-related protein, was downregulated in asthma patients and animal model. In this study, we observed that the expression of CTNNAL1 was increased in lung tissue of the ozone-stressed Balb/c mice model and in acute ozone stressed human bronchial epithelial cells (HBEC). In order to identify the possible DNA-binding proteins regulating the transcription of CTNNAL1 gene in HBEC, we designed 8 oligo- nucleotide probes corresponding to various regions of the CTNNAL1 promoter in electrophoretic mobility shift assays (EMSA). We detected 5 putative transcription factors binding sites within CTNNAL1 promoter region that can recruit LEF-1, AP-2α and CREB respectively by EMSA and antibody supershift assay. Chromatin immunoprecipitation (ChIP) assay verified that AP-2 α and LEF-1 could be recruited to the CTNNAL1 promoter. Therefore we further analyzed the functions of putative AP-2 and LEF-1 sites within CTNNAL1 promoter by site-directed mutagenesis of those sites within pGL3/FR/luc. We observed a reduction in human CTNNAL1 promoter activity of mutants of both AP-2α and LEF-1 sites. Pre-treatment with ASOs targeting LEF-1and AP-2α yielded significant reduction of ozone-stress-induced CTNNAL1 expression. The activation of AP-2α and LEF-1, followed by CTNNAL1 expression, showed a correlation during a 16-hour time course. Our data suggest that a robust transcriptional CTNNAL1 up-regulation occurs during acute ozone-induced stress and is mediated at least in part by ozone-induced recruitments of LEF-1 and AP-2α to the human CTNNAL1 promoter.
    Preview · Article · Feb 2012 · PLoS ONE
  • Source
    • "In this way, a number of known and novel GATA-1 protein partners were identified [8]. We also tested the utility of the biotin tag and streptavidin binding in ChIP assays and provided preliminary evidence that it can be successfully applied in place of antibodies in ChIPs of GATA-1 target genes [7,13]. Subsequent work in other labs has provided further supporting evidence for the application of biotinylation tagging in ChIP and Chip-on-chip assays [14-16]. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Chromatin immunoprecipitation (ChIP) assays coupled to genome arrays (Chip-on-chip) or massive parallel sequencing (ChIP-seq) lead to the genome wide identification of binding sites of chromatin associated proteins. However, the highly variable quality of antibodies and the availability of epitopes in crosslinked chromatin can compromise genomic ChIP outcomes. Epitope tags have often been used as more reliable alternatives. In addition, we have employed protein in vivo biotinylation tagging as a very high affinity alternative to antibodies. In this paper we describe the optimization of biotinylation tagging for ChIP and its coupling to a known epitope tag in providing a reliable and efficient alternative to antibodies. Using the biotin tagged erythroid transcription factor GATA-1 as example, we describe several optimization steps for the application of the high affinity biotin streptavidin system in ChIP. We find that the omission of SDS during sonication, the use of fish skin gelatin as blocking agent and choice of streptavidin beads can lead to significantly improved ChIP enrichments and lower background compared to antibodies. We also show that the V5 epitope tag performs equally well under the conditions worked out for streptavidin ChIP and that it may suffer less from the effects of formaldehyde crosslinking. The combined use of the very high affinity biotin tag with the less sensitive to crosslinking V5 tag provides for a flexible ChIP platform with potential implications in ChIP sequencing outcomes.
    Full-text · Article · Mar 2009 · BMC Molecular Biology
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Identification of downstream target genes of stress-relating transcription factors (TFs) is desirable in understanding cellular responses to various environmental stimuli. However, this has long been a difficult work for both experimental and computational practices. In this research, we presented a novel computational strategy which combined the analysis of the transcription factor binding site (TFBS) contexts and machine learning approach. Using this strategy, we conducted a genome-wide investigation into novel direct target genes of dehydration responsive element binding proteins (DREBs), the members of AP2-EREBPs transcription factor super family which is reported to be responsive to various abiotic stresses in Arabidopsis. The genome-wide searching yielded in total 474 target gene candidates. With reference to the microarray data for abiotic stresses-inducible gene expression profile, 268 target gene candidates out of the total 474 genes predicted, were induced during the 24-h exposure to abiotic stresses. This takes about 57% of total predicted targets. Furthermore, GO annotations revealed that these target genes are likely involved in protein amino acid phosphorylation, protein binding and Endomembrane sorting system. The results suggested that the predicted target gene candidates were adequate to meet the essential biological principle of stress-resistance in plants.
    Full-text · Article · Nov 2008 · Plant Molecular Biology
Show more