Project description:These data were used to infer the genome-wide localization of sigma70 and core RNAP beta subunit for the study "The transition between transcriptional initiation and elongation in E. coli is highly variable and often rate-limiting" (Reppas et al. 2006). This study analyzes transfrags with respect to the ChIP profile of the sigma70 and the beta subunit of RNA polymerase. Keywords: ChIP-chip; tiled analysis of mRNA expression
Project description:The in vivo trafficking patterns on DNA by the bacterial regulators of transcript elongation Sigma70, Rho, NusA, and NusG and the explanation for high promoter-proximal levels or peaks of RNA polymerase (RNAP) are unknown. Genome-wide ChIP-chip on E. coli revealed distinct association patterns of regulators as RNAP transcribes away from promoters (Rho first, then NusA, and then NusG). However, the interactions of elongating complexes with these regulators, including a weak interaction with Sigma70, did not differ significantly among most transcription units. A modest variation of NusG signal among genes reflected increased NusG interaction as transcription progresses, rather than functional specialization of elongating complexes. Promoter-proximal RNAP peaks were offset from Sigma70 peaks in the direction of transcription and co-occurred with NusA and Rho peaks, suggesting that the RNAP peaks reflected elongating, rather than initiating, complexes. However, inhibition of Rho did not increase RNAP levels within genes downstream of the RNAP peaks, suggesting the peaks are caused by a mechanism other than simple Rho-dependent attenuation. Chromatin immunoprecipitation (ChIP) experiments were performed using antibodies against RNA polymerase (Beta' subunit), Sigma70, NusA, NusG, or Rho. Differentially labeled ChIP DNA and genomic DNA were competitively hybridized to an E. coli K-12 MG1655 tiling array with overlapping probes at ~24bp spacing across the entire genome. The series contains 17 total datasets.
Project description:We integrated RNAP binding regions (RBRs) and mRNA transcript abundance to determine segments of contiguous transcription originating from promoter regions. To measure RBRs at a genome scale, we employed a ChIP-chip method to E. coli K-12 MG1655 grown in the presence or absence of rifampicin under multiple growth conditions using antibody against E. coli RNAP beta subunit.
Project description:The in vivo trafficking patterns on DNA by the bacterial regulators of transcript elongation Sigma70, Rho, NusA, and NusG and the explanation for high promoter-proximal levels or peaks of RNA polymerase (RNAP) are unknown. Genome-wide ChIP-chip on E. coli revealed distinct association patterns of regulators as RNAP transcribes away from promoters (Rho first, then NusA, and then NusG). However, the interactions of elongating complexes with these regulators, including a weak interaction with Sigma70, did not differ significantly among most transcription units. A modest variation of NusG signal among genes reflected increased NusG interaction as transcription progresses, rather than functional specialization of elongating complexes. Promoter-proximal RNAP peaks were offset from Sigma70 peaks in the direction of transcription and co-occurred with NusA and Rho peaks, suggesting that the RNAP peaks reflected elongating, rather than initiating, complexes. However, inhibition of Rho did not increase RNAP levels within genes downstream of the RNAP peaks, suggesting the peaks are caused by a mechanism other than simple Rho-dependent attenuation.
Project description:We integrated RNAP binding regions (RBRs) and mRNA transcript abundance to determine segments of contiguous transcription originating from promoter regions. To measure RBRs at a genome scale, we employed a ChIP-chip method to E. coli K-12 MG1655 grown in the presence or absence of rifampicin under multiple growth conditions using antibody against E. coli RNAP beta subunit. A twelve ChIP-chip study using immunoprecipitated DNA (IP-DNA) from four separate culture conditions with and/or without rifampicin treatment. The high-density oligonucleotide tiling arrays used were consisted of 371,034 oligonucleotide probes spaced 25 bp apart (25-bp overlap between two probes) across the E. coli genome (NimbleGen). Experiments were conducted as biological duplicates or triplicates (different cultures).
Project description:Mapping the occupancy of FNR, HNS, and IHF throughout the genome of Escherchia coli MG1655 K-12 using an affinity purified antibody under anerobic growth conditions. We also mapped the binding of the M-CM-^_ subunit of RNA Polymerase under both aerobic and anaerobic growth conditions. As a control, we also performed ChIP-chip on FNR in a M-bM-^HM-^Ffnr mutant strain of Escherchia coli MG1655 K-12. We also examined FNR immunoprecipitation at various FNR concentrations using IPTG and Ptac::fnr (PK8263). The M-bM-^HM-^Fhns/M-bM-^HM-^FstpA strains were also used. Descirbed in the manuscript Genome-scale Analysis of E. coli FNR Reveals the Complexity of Bacterial Regulon Structure Mapping of occupancy of FNR, NNS, IHF and M-CM-^_ of RNAP in the genome of Escherchia coli MG1655 K-12 under aerobic or anaerobic growth conditions. Immunoprecipitated DNA compared to INPUT for each sample.
Project description:Mapping the occupancy of ArcA throughout the genome of Escherchia coli MG1655 K-12 using an affinity purified antibody under anaerobic and aerobic growth conditions. As a control, we also performed ChIP-chip onArcA in a ∆arcA mutant strain of Escherchia coli MG1655 K-12. Described in the manuscript The response regulator ArcA uses a diverse binding site architechture to globally regulate carbon oxidation in E. coli
Project description:Bacterial transcription factors (TFs) regulate gene expression to adapt to changing environments; when combined, the TF’s regulatory actions comprise transcriptional regulatory networks (TRNs). The chromatin immunoprecipitation (ChIP) assay is the major contemporary method for mapping in vivo protein-DNA interactions in the genome. It enables the genome-wide study of transcription factor binding sites (TFBSs) and gene regulation. Here, we present the genome-wide binding for major TFs in E. coli K-12 MG1655.