Project description:In Mycobacterium tuberculosis the alternative sigma factor SigF controls the expression of a particular subset of genes by altering RNA polymerase specificity. Here, we utilize two genome-wide approaches to identify SigF-binding sites: chromatin immunoprecipitation (ChIP-on-chip) and microarray analysis of SigF-mediated transcripts. Since SigF is not an abundant protein in the logarithmic phase of growth, a pristinamyin IA-inducible system was used to control its expression. We identified 67 high-affinity SigF-binding sites and 16 loci where a SigF promoter directs the expression of a transcript. These loci include sigF itself, genes involved in lipid and intermediary metabolism and virulence, and at least one transcriptional regulator (Rv2884), possibly acting downstream of SigF. In addition, SigF was also found to direct the transcription of the gene for small RNA F6. Many loci were also found where SigF may be involved in antisense transcription, and in two cases (Rv1358 and Rv1870c) the SigF-dependent promoter was located within the predicted coding sequence. Quantitative PCR confirmed the microarray findings and 5'-rapid amplification of cDNA ends was used to map the SigF-specific transcriptional start points. A canonical SigF consensus promoter sequence GGTTT-N((15-17))-GGGTA was found prior to 11 genes. Together, these data help to define the SigF regulon and show that SigF not only governs expression of proteins such as the virulence factor, HbhA, but also impacts novel functions, such as noncoding RNAs and antisense transcripts.
Project description:This SuperSeries is composed of the following subset Series: GSE34919: Genome-wide Definition of the SigF Regulon in Mycobacterium tuberculosis (ChIP-chip) GSE34922: Genome-wide Definition of the SigF Regulon in Mycobacterium tuberculosis (Expression) Refer to individual Series
Project description:The genome wide transcriptome analysis to identify the M.tuberculosis sigma factor Rv3286c (SigF) dependent transcripts as well as Pristimaycin induced transcripts
Project description:The genome wide transcriptome analysis to identify the M.tuberculosis sigma factor Rv3286c (SigF) dependent transcripts as well as Pristimaycin induced transcripts Pristinamycin induced transcripts were identified by comparing the cDNA extracted from H37Rv::pMY769 with and without pristinamycin (3 biological replicates). SigF controlled transcripts were identified by comparing the cDNA extracted from H37Rv::pMYsigF with and without pristinamycin (4 biological replicates).
Project description:The genome wide ChIP-on-chip analysis to identify the DNA binding sites for the M.tuberculosis sigma factor Rv3286c (SigF) SigF binding sites were determined by microarray analysis of anti-sigF immunoprecipitated DNA from H37Rv(ΔrsbW/sigF)::pmySigF compared to H37Rv(ΔrsbW/sigF)::pMY769 (both cultured with pristinamycin for 3 days). 4 biological replicates were performed.
Project description:Mycobacterium smegmatis SigF is a group III sigma factor. Its ortholog in M. tuberculosis is reported to have role in regulation and function of cell wall components. In present study we have created an M. smegmatis ΔsigF mutant by allele exchange method. M. smegmatis sigF mutant shows non pigmented phenotype and is more sensitive to hydrogen peroxide generated oxidative stress. DNA microarray analysis of M. smegmatis wild type and ΔsigF mutant suggests that SigF in this species controls the expression of several energy and central intermediary metabolism genes along with regulation of carotenoid biosynthesis.