Unknown,Transcriptomics,Genomics,Proteomics

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ChIP-Seq with anti-FLAG antibodies to determine binding of LexA-3xFLAG to DNA in Streptomyces venezuelae treated with Mitomycin C (MMC) overnight

ABSTRACT: LexA is a transcriptional repressor for genes requiring expression primarily during SOS response in response to stress such as that caused by DNA alkylating agent Mitomycin C (MMC). LexA undergoes self-cleavage under stress conditions thereby lifting the repression on genes whose expression is required for stress response. In our experiment LexA binding sites in the genome were determined in untreated and MMC treated (overnight, 14h) cultures using a LexA-3xFLAG strain and anti-FLAG monoclonal antibodies. Wild type Streptomyces venezuelae (unFLAGged LexA) cultures, with and without MMC treatment were used as negative controls.

INSTRUMENT(S): Illumina HiSeq 2000

ORGANISM(S): Streptomyces venezuelae

SUBMITTER: Kathry Stratton

PROVIDER: E-MTAB-11603 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:The SOS response is a conserved pathway that is activated under certain stress conditions and is regulated by the repressor LexA and the activator RecA. The food-borne pathogen Listeria monocytogenes contains RecA and LexA homologs, but their roles in Listeria have not been established. In this study, we identified the SOS regulon in L. monocytogenes by comparing the transcription profiles of the wild-type strain and the ÎrecA mutant strain after exposure to the DNA damaging agent mitomycinC (MMC). The SOS response is an inducible pathway involved in DNA repair, restart of stalled replication forks, and in induction of genetic variation in stressed and stationary phase cells. It is regulated by LexA and RecA. LexA is an autoregulatory repressor which binds to a consensus sequence in the promoter region of the SOS response genes, thereby repressing transcription. A consensus LexA binding motif for L. monocytogenes has not been identified thus far. Generally, the SOS response is induced under circumstances in which single stranded DNA accumulates in the cell. This results in activation of RecA, which in turn stimulates cleavage of LexA, and ultimately in the induction of the SOS response. Keywords: stress response, loop design, SOS response, mitomycin c, listeria monocytogenes, RecA, LexA Triple loop design of 3 independent experiments (series A, B, and C) using Wt strain and recA mutant (activator of the SOS response). Sampling was done before (t=0) and 1 hour after (t=60) exposure to mitomycin C (MMC) for both the wild-type strain and the recA mutant strain. One series therefore contains 4 samples and the complete experiments consists of 12 samples. Each of the 3 series was designed in a loop (wt, t=0 =>wt, t=60 => recA, t=60 => recA, t=0 => wt, t=0). These 3 loops were connected using series B as the central loop. Series B was connected to series A as follows: wt, t=0 (B) => recA, t=0 (A) and wt, t=60 (B) => recA, t=60 (A). Series B was connected to series C as follows: recA, t=0 (B) => wt, t=0 (C) and recA, t=60 (B) => wt, t=60 (C).

Dataset Information

ChIP-Seq with anti-FLAG antibodies to determine binding of LexA-3xFLAG to DNA in Streptomyces venezuelae treated with Mitomycin C (MMC) overnight

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