title: Genome-wide discovery of functional transcription factor binding sites by comparative genomics: the case of Stat3.
abstract: The identification of direct targets of transcription factors is a key problem in the study of gene regulatory networks. However, the use of high throughput experimental methods, such as ChIP-chip and ChIP-sequencing, is limited by their high cost and strong dependence on cellular type and context. We developed a computational method for the genome-wide identification of functional transcription factor binding sites based on positional weight matrices, comparative genomics, and gene expression profiling. The method was applied to Stat3, a transcription factor playing crucial roles in inflammation, immunity and oncogenesis, and able to induce distinct subsets of target genes in different cell types or conditions. A newly generated positional weight matrix enabled us to assign affinity scores of high specificity, as measured by EMSA competition assays. Phylogenetic conservation with 7 vertebrate species was used to select the binding sites most likely to be functional. Validation was carried out on predicted sites within genes identified as differentially expressed in the presence or absence of Stat3 by microarray analysis. Twelve of the fourteen sites tested were bound by Stat3 in vivo, as assessed by Chromatin Immunoprecipitation, allowing us to identify 9 Stat3 transcriptional targets. Given its high validation rate, and the availability of large transcription factor-dependent gene expression datasets obtained under diverse experimental conditions, our approach appears to be a valid alternative to high-throughput experimental assays for the discovery of novel direct targets of transcription factors.
authors.affiliation: Molecular Biotechnology Center and Department of Genetics, Biology and Biochemistry and Department of Clinical and Biological Sciences, University of Turin, Via Nizza 52, 10126 Turin, Italy.
authors.forename: Francesco
authors.initials: F
authors.lastname: Vallania
authors.forename: Davide
authors.initials: D
authors.lastname: Schiavone
authors.forename: Sarah
authors.initials: S
authors.lastname: Dewilde
authors.forename: Emanuela
authors.initials: E
authors.lastname: Pupo
authors.forename: Serge
authors.initials: S
authors.lastname: Garbay
authors.forename: Raffaele
authors.initials: R
authors.lastname: Calogero
authors.forename: Marco
authors.initials: M
authors.lastname: Pontoglio
authors.forename: Paolo
authors.initials: P
authors.lastname: Provero
authors.forename: Valeria
authors.initials: V
authors.lastname: Poli
mesh_terms: D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D024202:Electrophoretic Mobility Shift Assay; D020869:Gene Expression Profiling; D016678:Genome; D023281:Genomics / Q000379:methods*; D008722:Methods; D051379:Mice; D018345:Mice, Knockout; D050796:STAT3 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*
chemical_list: D050796:STAT3 Transcription Factor; D014157:Transcription Factors