Role of intragenic binding of cAMP responsive protein (CRP) in regulation of the succinate dehydrogenase genes Rv0249c-Rv0247c in TB complex mycobacteria.
Role of intragenic binding of cAMP responsive protein (CRP) in regulation of the succinate dehydrogenase genes Rv0249c-Rv0247c in TB complex mycobacteria.
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DOI：
10.1093/nar/gkv420
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发表时间：
2015-06-23
影响因子：
14.9
通讯作者：
McDonough KA
中科院分区：
文献类型：
--
作者：
Knapp GS;Lyubetskaya A;Peterson MW;Gomes AL;Ma Z;Galagan JE;McDonough KA
细菌病原体适应宿主内不断变化的环境，信号分子 3', 5'-环单磷酸腺苷 (cAMP) 促进了这一过程。在这项研究中，我们以结核菌 (TB) 复合菌为模型，对牛分枝杆菌 BCG 中 cAMP 响应蛋白 CRP 的体内 DNA 结合和基因调控进行了表征。染色质免疫沉淀随后进行深度测序 (ChIP-seq) 显示 CRP 与约 900 个 DNA 结合区域相关，其中大部分发生在基因内。最高度富集的结合区域位于假定的铜转运蛋白基因 (ctpB) 的上游，并且 crp 缺失的细菌表现出对铜毒性的敏感性增加。对毒力相关的 Rv0249c-Rv0247c 琥珀酸脱氢酶基因上游的四个 CRP 结合位点的详细突变分析表明，CRP 通过两个启动子直接调节 Rv0249c-Rv0247c 的表达，其中一个启动子需要 Rv0250c 的基因内序列才能最大表达。高比例的基因内 CRP 结合位点以及我们证明这些基因内 DNA 序列对生物学相关基因表达有显着贡献，极大地扩展了分枝杆菌基因调控分析必须考虑的基因组空间。这些发现对于一种重要的细菌病原体也具有实际意义，其中影响药物靶标相关基因表达的突变的鉴定被广泛用于快速耐药性筛查。
Bacterial pathogens adapt to changing environments within their hosts, and the signaling molecule adenosine 3′, 5′-cyclic monophosphate (cAMP) facilitates this process. In this study, we characterized in vivo DNA binding and gene regulation by the cAMP-responsive protein CRP in M. bovis BCG as a model for tuberculosis (TB)-complex bacteria. Chromatin immunoprecipitation followed by deep-sequencing (ChIP-seq) showed that CRP associates with ∼900 DNA binding regions, most of which occur within genes. The most highly enriched binding region was upstream of a putative copper transporter gene (ctpB), and crp-deleted bacteria showed increased sensitivity to copper toxicity. Detailed mutational analysis of four CRP binding sites upstream of the virulence-associated Rv0249c-Rv0247c succinate dehydrogenase genes demonstrated that CRP directly regulates Rv0249c-Rv0247c expression from two promoters, one of which requires sequences intragenic to Rv0250c for maximum expression. The high percentage of intragenic CRP binding sites and our demonstration that these intragenic DNA sequences significantly contribute to biologically relevant gene expression greatly expand the genome space that must be considered for gene regulatory analyses in mycobacteria. These findings also have practical implications for an important bacterial pathogen in which identification of mutations that affect expression of drug target-related genes is widely used for rapid drug resistance screening.
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影响因子：
2.8
作者：
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通讯作者：
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