Ho et al DNA binding proteins

Ho et al DNA binding proteins - Linking DNA-binding...

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Unformatted text preview: Linking DNA-binding proteins to their recognition sequences by using protein microarrays Su-Wen Ho* † , Ghil Jona †‡§ , Christina T. L. Chen*, Mark Johnston* ¶ , and Michael Snyder ‡¶ *Washington University School of Medicine, 4444 Forest Park Boulevard, St. Louis, MO 63108; and ‡ Department of Molecular, Cellular, and Developmental Biology, Yale University, 266 Whitney Avenue, New Haven, CT 06520-8103 Edited by Stanley Fields, University of Washington, Seattle, WA, and approved May 4, 2006 (received for review October 20, 2005) Analyses of whole-genome sequences and experimental data sets have revealed a large number of DNA sequence motifs that are conserved in many species and may be functional. However, methods of sufficient scale to explore the roles of these elements are lacking. We describe the use of protein arrays to identify proteins that bind to DNA sequences of interest. A microarray of 282 known and potential yeast transcription factors was produced and probed with oligonucleotides of evolutionarily conserved sequences that are potentially functional. Transcription factors that bound to specific DNA sequences were identified. One previ- ously uncharacterized DNA-binding protein, Yjl103, was character- ized in detail. We defined the binding site for this protein and identified a number of its target genes, many of which are involved in stress response and oxidative phosphorylation. Protein microar- rays offer a high-throughput method for determining DNA–protein interactions. proteomics u transcription factor u yeast A fundamental problem in biology is to identify cis-regulatory DNA sequence elements and the proteins that bind to them. Such information is necessary for uncovering gene regulatory networks that control cellular and developmental processes. Genome-wide approaches have revealed many DNA sequence elements that may regulate gene expression: comparison of genome sequences of related organisms has identified thousands of evolutionarily conserved DNA sequence motifs (1–3); com- parison of the sequences adjacent to coregulated sets of genes of an organism often reveals shared sequence motifs (4–7). Veri- fying functionality of these sequences and identifying the pro- teins that bind to them remains a significant challenge. Several methods have recently been developed to map globally the DNA-binding sites of transcription factors. The SELEX method enables in vitro selection of the optimal binding site of a transcription factor (8), although applying it genome-wide may be difficult. In the chromatin immunoprecipitation ‘‘(ChIP)- chip’’ method, chromatin bound by a transcription factor of interest is immunoprecipitated, and the associated DNA is identified by using it to probe a genomic DNA microarray, thereby identifying the targets of the transcription factor (9, 10)....
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Ho et al DNA binding proteins - Linking DNA-binding...

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