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Journal of Bacteriology, October 2004, p. 6938-6943, Vol. 186, No. 20
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.20.6938-6943.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Genomic Studies with Escherichia coli MelR Protein: Applications of Chromatin Immunoprecipitation and Microarrays

David C. Grainger,1* Timothy W. Overton,1 Nikos Reppas,2 Joseph T. Wade,2 Eiji Tamai,3 Jon L. Hobman,1 Chrystala Constantinidou,1 Kevin Struhl,2 George Church,2 and Stephen J. W. Busby1

School of Biosciences, The University of Birmingham, Edgbaston, Birmingham, United Kingdom,1 Department of Microbiology, Faculty of Medicine, Kagawa University, Kita-gun, Kagawa, Japan,3 Department of Biological Chemistry and Molecular Pharmacology and Department of Genetics, Harvard Medical School, Boston, Massachusetts2

Received 9 June 2004/ Accepted 22 July 2004

Escherichia coli MelR protein is a transcription activator that is essential for melibiose-dependent expression of the melAB genes. We have used chromatin immunoprecipitation to study the binding of MelR and RNA polymerase to the melAB promoter in vivo. Our results show that MelR is associated with promoter DNA, both in the absence and presence of the inducer melibiose. In contrast, RNA polymerase is recruited to the melAB promoter only in the presence of inducer. The MelR DK261 positive control mutant binds to the melAB promoter but cannot recruit RNA polymerase. Further analysis of immunoprecipitated DNA, by using an Affymetrix GeneChip array, showed that the melAB promoter is the major, if not the sole, target in E. coli for MelR. This was confirmed by a transcriptomics experiment to analyze RNA in cells either with or without melR.

* Corresponding author. Mailing address: School of Biosciences, The University of Birmingham, Edgbaston, Birmingham B15 2TT, United Kingdom. Phone: 44 121 414 5435. Fax: 44 121 414 7366. E-mail: d.grainger{at}bham.ac.uk.

Journal of Bacteriology, October 2004, p. 6938-6943, Vol. 186, No. 20
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.20.6938-6943.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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