The ATP-dependent chromatin remodeling enzyme Fun30 represses transcription by sliding promoter proximal nucleosomes
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ABSTRACT:
ORGANISM(S): Saccharomyces cerevisiae
SUBMITTER: Wei-Hua Wu
PROVIDER: E-GEOD-48571 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:The evolutionarily conserved ATP-dependent chromatin remodeling enzyme Fun30 has recently been shown to play important roles in heterochromatin silencing and DNA repair. However, how Fun30 remodels nucleosomes is not clear. Here we report a nucleosome sliding activity of Fun30 and its role in transcriptional repression. We observed that Fun30 repressed the expression of genes involved in amino acid and carbohydrate metabolism, the stress response, and meiosis. In addition, Fun30 was localized at the 5′ and 3′ ends of genes and within the open reading frames of its targets. Consistent with its role in gene repression, we observed that Fun30-target genes lacked histone modifications often associated with gene activation and showed an increased level of ubiquitinated histone H2B. Furthermore, genome-wide nucleosome mapping analysis revealed that the length of the nucleosome-free region at the 5′ end of a subset of genes was changed in Fun30-depleted cells. In addition, the positions of the -1, +2 and +3 nucleosomes at the 5′ end of target genes were significantly shifted, while position of the +1 nucleosome remained largely unchanged in the fun30Δ mutant. Finally, we demonstrated that affinity purified single-component Fun30 exhibited nucleosome sliding activity in an ATP-dependent manner. These results define a role for Fun30 in regulation of transcription and indicate that Fun30 remodels chromatin at the 5′ end of genes by sliding promoter proximal nucleosomes. Data sets for each strain were generated from three biological replicates and experimental dye swap duplicates of each replicate. The six wildtype raw data files linked below were used to generate normalized, log2 (mutant/WT) ratios for all 3 Samples.
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Project description:The two-component system CBO0366/CBO0365 was recently demonstrated to have a role in cold tolerance of Group I Clostridium botulinum ATCC 3502. The mechanisms under its control, ultimately resulting in increased sensitivity to low temperature, are unknown. A transcriptomic analysis with DNA microarrays was performed to identify the differences in global gene expression patterns of the wild-type ATCC 3502 and a derivative mutant with insertionally inactivated cbo0365 at 37 M-BM-0C and 15 M-BM-0C. Altogether 150 or 141 chromosomal CDSs were found to be differently expressed in the cbo0365 mutant at 37 M-BM-0C or 15 M-BM-0C, respectively, and thus considered to be under direct or indirect transcriptional control of the response regulator CBO0365. Of the differentially-expressed CDSs, expression of 141 CDSs was similarly affected at both temperatures investigated, suggesting that the putative CBO0365 regulon was practically not affected by temperature. The regulon involved genes related to acetone-butanol-ethanol (ABE) fermentation, motility, to arsenic resistance, and phosphate uptake and transport. Deteriorated growth at 17 M-BM-0C was observed for mutants with disrupted ABE fermentation pathway components (crt, bdh and ctfA), arsenic detoxifying machinery components (arsC and arsR), or phosphate uptake mechanism components (phoT), suggesting roles for these mechanisms in cold tolerance of Group I C. botulinum. Electrophoretic mobility shift assays showed recombinant CBO0365 to bind to the promoter regions of crt, arsR, and phoT, as well as to the promoter region of its own operon, suggesting direct DNA-binding transcriptional activation or repression as means for CBO0365 in regulating these operons. The results provide insight to the mechanisms Group I C. botulinum utilize in coping with cold. C. botulinum ATCC 3502 cbo0365 mutant vs. wild type; 3 replicates of each strain; growth at 37C in TPGY broth batch culture and subjected to cold shock to 15C; sampling at mid-log growth phase before cold shock, and 1 h after temperature downshift to 15C (= 2 time points). Dye-swapped hybridization.
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