title: Transcription Factor Binding Site Mapping Using ChIP-Seq.
abstract: Transcription factors (TFs) play a central role in regulating gene expression in all bacteria. Yet until recently, studies of TF binding were limited to a small number of factors at a few genomic locations. Chromatin immunoprecipitation followed by sequencing (ChIP-Seq) provides the ability to map binding sites globally for TFs, and the scalability of the technology enables the ability to map binding sites for every DNA binding protein in a prokaryotic organism. We have developed a protocol for ChIP-Seq tailored for use with mycobacteria and an analysis pipeline for processing the resulting data. The protocol and pipeline have been used to map over 100 TFs from Mycobacterium tuberculosis, as well as numerous TFs from related mycobacteria and other bacteria. The resulting data provide evidence that the long-accepted spatial relationship between TF binding site, promoter motif, and the corresponding regulated gene may be too simple a paradigm, failing to adequately capture the variety of TF binding sites found in prokaryotes. In this article we describe the protocol and analysis pipeline, the validation of these methods, and the results of applying these methods to M. tuberculosis.
authors.affiliation: Department of Biomedical Engineering, Boston University, Boston, MA 02215.
authors.forename: Suma
authors.initials: S
authors.lastname: Jaini
authors.affiliation: Bioinformatics Program, Boston University, Boston, MA 02215.
authors.forename: Anna
authors.initials: A
authors.lastname: Lyubetskaya
authors.affiliation: Bioinformatics Program, Boston University, Boston, MA 02215.
authors.forename: Antonio
authors.initials: A
authors.lastname: Gomes
authors.affiliation: Department of Biomedical Engineering, Boston University, Boston, MA 02215.
authors.forename: Matthew
authors.initials: M
authors.lastname: Peterson
authors.affiliation: Macrogen Clinical Laboratory, Macrogen Corp, Rockville, MD 20850.
authors.forename: Sang
authors.initials: S
authors.lastname: Tae Park
authors.affiliation: National Emerging Infectious Diseases Laboratory, Boston University, Boston, MA 02118.
authors.forename: Sahadevan
authors.initials: S
authors.lastname: Raman
authors.affiliation: Department of Medicine and Department of Microbiology and Immunology, Stanford Medical School, Stanford, CA 94305.
authors.forename: Gary
authors.initials: G
authors.lastname: Schoolnik
authors.affiliation: Department of Biomedical Engineering, Boston University, Boston, MA 02215.
authors.forename: James
authors.initials: J
authors.lastname: Galagan
mesh_terms: D001665:Binding Sites; D047369:Chromatin Immunoprecipitation / Q000379:methods*; D019295:Computational Biology / Q000379:methods; D004247:DNA / Q000378:metabolism*; D005827:Genetics, Microbial / Q000379:methods; D008967:Molecular Biology / Q000379:methods*; D009169:Mycobacterium tuberculosis / Q000235:genetics*; D011485:Protein Binding; D017422:Sequence Analysis, DNA / Q000379:methods*; D014157:Transcription Factors / Q000378:metabolism*
chemical_list: D014157:Transcription Factors; D004247:DNA