Expanding the Regulatory Network Governed by the Extracytoplasmic Function Sigma Factor σ^Hin Corynebacterium glutamicum

Abstract

bThe extracytoplasmic function sigma factor H is responsible for the heat and oxidative stress response in Corynebacterium glutamicum. Due to the hierarchical nature of the regulatory network, previous transcriptome analyses have not been able to discriminate between direct and indirect targets of H . Here, we determined the direct genome-wide targets of H using chromatin immunoprecipitation with microarray technology (ChIP-chip) for analysis of a deletion mutant of rshA, encoding an antifactor of H . Seventy-five H -dependent promoters, including 39 new ones, were identified. H -dependent, heat-inducible transcripts for several of the new targets, including ilvD encoding a labile Fe-S cluster enzyme, dihydroxy-acid dehydratase, were detected, and their 5= ends were mapped to the H -dependent promoters identified. Interestingly, functional internal H -dependent promoters were found in operon-like gene clusters involved in the pentose phosphate pathway, riboflavin biosynthesis, and Zn uptake. Accordingly, deletion of rshA resulted in hyperproduction of riboflavin and affected expression of Zn-responsive genes, possibly through intracellular Zn overload, indicating new physiological roles of H . Furthermore, sigA encoding the primary factor was identified as a new target of H . Reporter assays demonstrated that the H -dependent promoter upstream of sigA was highly heat inducible but much weaker than the known A -dependent one. Our ChIP-chip analysis also detected the H -dependent promoters upstream of rshA within the sigH-rshA operon and of sigB encoding a group 2 factor, supporting the previous findings of their H -dependent expression. Taken together, these results reveal an additional layer of the sigma factor regulatory network in C. glutamicum.

SigR, a hub of multilayered regulation of redox and antibiotic stress responses

“…As in S. coelicolor , Cgl σ H directly regulates MSH metabolism in the synthesis ( mshC ) and recycle ( mca ) of S‐conjugated MSH (Busche et al , ; Si et al , ). The Cgl σ H ‐specific promoter consensus sequence, GGAA‐N 18‐20 ‐GTT (Toyoda et al , ), very closely matches with that of σ R . Among five ECF sigma factors in C. glutamicum , σ M recognizes promoters with σ H ‐specific consensus sequence with overlapping target genes such as trxBC and sufR (Nakunst et al , ; Patek and Nesvera, ).…”

“…Cgl σ H controls responses against heat shock and oxidative stresses (Kim et al , ; Ehira et al , ). The σ H regulon, which was determined via transcriptome (Busche et al , ) and ChIP‐chip (Toyoda et al , ) analyses, consists of > 75 transcription units and shares many genes with the S. coelicolor σ R regulon (Table ) and Mtb σ H regulon. Transcription of the sigH gene is regulated at multiple promoters.…”

“…Transcription of the sigH gene is regulated at multiple promoters. Cgl σ H controls genes for other transcriptional regulators, such as σ A , σ B , σ M , HspR, SufR and ClgR (Engels et al , ; Nakunst et al , ; Ehira et al , ; Busche et al , ; Toyoda et al , ), exhibiting its central role as a regulatory hub. As in S. coelicolor , Cgl σ H directly regulates MSH metabolism in the synthesis ( mshC ) and recycle ( mca ) of S‐conjugated MSH (Busche et al , ; Si et al , ).…”

RNase III mediated cleavage of the coding region of mraZ mRNA is required for efficient cell division in Corynebacterium glutamicum

“…Transcriptome analysis was performed as described previously (Toyoda et al, 2015). Comparative analysis of the deletion mutants with the wild type was performed using a microarray (designated as ORF array) containing only ORFspecific oligonucleotide probes that were spotted onto a glass slide (Takara Bio).…”

Enhanced Glucose Consumption and Organic Acid Production by Engineered Corynebacterium glutamicum Based on Analysis of a pfkB1 Deletion Mutant

“…A DNA microarray assay for transcriptome analysis was conducted on the Agilent eArray platform (Agilent Technologies, Palo Alto, CA) as described previously (31,58). RNA was extracted from the cells during aerobic cultivation on A medium with glucose or fructose in the early exponential phase (at 4 h) and in late exponential/stationary phases (at 12 h).…”