ChIP-chip: a genomic approach for identifying transcription factor binding sites.

Publisher Summary Transcription factors are key regulatory proteins that can influence the expression of hundreds of genes in response to a particular environmental condition or internal cue. The collective set of genes regulated by a transcription factor therefore defines the state of a cell and can determine cell fate. Among the 6200 predicted proteins in the yeast Saccharomyces cerevisiae, there are about 300 transcription factors. Approximately 85% of the yeast transcription factors have been characterized to some extent and some of these are known to play a critical role in cell cycle initiation, pheromone response, mating type switching, pseudohyphal growth, and nutrient and stress response. This chapter develops an approach in yeast that will comprehensively identify genomic sequences directly bound by transcription factors. This approach has been used to successfully identify targets of the yeast transcription factors SBF and MBF. Briefly, protein-DNA complexes are fixed in vivo with formaldehyde and lysed, and the lysate is sonicated to shear DNA. The transcription factor of interest is purified by immunoprecipitation; the associated DNA is extracted and then amplified and labeled for hybridization to a yeast intergenic array. Two important reagents for this method are specific immunoprecipitating antibodies and a yeast intergenic microarray.

[1]  K. Luo,et al.  SIR2 and SIR4 interactions differ in core and extended telomeric heterochromatin in yeast. , 1997, Genes & development.

[2]  P. Brown,et al.  Exploring the metabolic and genetic control of gene expression on a genomic scale. , 1997, Science.

[3]  P. Brown,et al.  Identification of the Copper Regulon in Saccharomyces cerevisiae by DNA Microarrays* , 2000, The Journal of Biological Chemistry.

[4]  D Botstein,et al.  Desferrioxamine-mediated Iron Uptake in Saccharomyces cerevisiae , 2000, The Journal of Biological Chemistry.

[5]  Renato Paro,et al.  Mapping polycomb-repressed domains in the bithorax complex using in vivo formaldehyde cross-linked chromatin , 1993, Cell.

[6]  Michael E. Cusick,et al.  The Yeast Proteome Database (YPD) and Caenorhabditis elegans Proteome Database (WormPD): comprehensive resources for the organization and comparison of model organism protein information , 2000, Nucleic Acids Res..

[7]  A. Futcher,et al.  Use of polymerase chain reaction epitope tagging for protein tagging in Saccharomyces cerevisiae , 1995, Yeast.

[8]  A. Goffeau,et al.  Genome microarray analysis of transcriptional activation in multidrug resistance yeast mutants , 2000, FEBS letters.

[9]  D. Botstein,et al.  The transcriptional program of sporulation in budding yeast. , 1998, Science.

[10]  C. Allis,et al.  In vivo cross-linking and immunoprecipitation for studying dynamic Protein:DNA associations in a chromatin environment. , 1999, Methods.

[11]  J. Axelrod,et al.  An improved method for photofootprinting yeast genes in vivo using Taq polymerase. , 1989, Nucleic acids research.

[12]  R. Paro,et al.  Analysis of chromatin structure by in vivo formaldehyde cross-linking. , 1997, Methods.

[13]  Alexander Varshavsky,et al.  Mapping proteinDNA interactions in vivo with formaldehyde: Evidence that histone H4 is retained on a highly transcribed gene , 1988, Cell.

[14]  Jun S. Liu,et al.  Gibbs motif sampling: Detection of bacterial outer membrane protein repeats , 1995, Protein science : a publication of the Protein Society.

[15]  Jun S. Liu,et al.  Detecting subtle sequence signals: a Gibbs sampling strategy for multiple alignment. , 1993, Science.

[16]  D. Botstein,et al.  Genomic binding sites of the yeast cell-cycle transcription factors SBF and MBF , 2001, Nature.

[17]  M. Grunstein,et al.  Mapping DNA interaction sites of chromosomal proteins using immunoprecipitation and polymerase chain reaction. , 1999, Methods in enzymology.