Genomic studies with Escherichia coli MelR protein: applications of chromatin immunoprecipitation and microarrays.
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.
Grainger
David C
DC
School of Biosciences, The University of Birmingham, Edgbaston, Birmingham B15 2TT, United Kingdom. d.grainger@bham.ac.uk.
Overton
Timothy W
TW
Reppas
Nikos
N
Wade
Joseph T
JT
Tamai
Eiji
E
Hobman
Jon L
JL
Constantinidou
Chrystala
C
Struhl
Kevin
K
Church
George
G
Busby
Stephen J W
SJ
D002843:Chromatin; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D012321:DNA-Directed RNA Polymerases / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial; D016680:Genome, Bacterial; D008553:Melibiose / Q000378:metabolism*; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D011233:Precipitin Tests / Q000379:methods*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D027981:Symporters / Q000235:genetics / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*
D002843:Chromatin; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C077649:MelR protein, E coli; D027981:Symporters; D015534:Trans-Activators; C021548:melibiose permease; D008553:Melibiose; D012321:DNA-Directed RNA Polymerases