Department of Integrated Biological Science, Pusan National Universitygrid.262229.f, Busan, South Korea.
Department of Chemistry, Pusan National Universitygrid.262229.f, Busan, South Korea.
J Bacteriol. 2021 Nov 5;203(23):e0040221. doi: 10.1128/JB.00402-21. Epub 2021 Sep 13.
Mycobacterium smegmatis has two isocitrate lyase (ICL) isozymes (MSMEG_0911 and MSMEG_3706). We demonstrated that ICL1 (MSMEG_0911) is the predominantly expressed ICL in M. smegmatis and plays a major role in growth on acetate or fatty acid as the sole carbon and energy source. Expression of the gene in M. smegmatis was demonstrated to be strongly upregulated during growth on acetate relative to that in M. smegmatis grown on glucose. Expression of was shown to be positively regulated by the RamB activator, and three RamB-binding sites (RamBS1, RamBS2, and RamBS3) were identified in the upstream region of using DNase I footprinting analysis. Succinyl coenzyme A (succinyl-CoA) was shown to increase the affinity of binding of RamB to its binding sites and enable RamB to bind to RamBS2, which is the most important site for RamB-mediated induction of expression. These results suggest that succinyl-CoA serves as a coinducer molecule for RamB. Our study also showed that cAMP receptor protein (Crp1; MSMEG_6189) represses expression in M. smegmatis grown in the presence of glucose. Therefore, the strong induction of expression during growth on acetate as the sole carbon source relative to the weak expression of during growth on glucose is likely to result from combined effects of RamB-mediated induction of in the presence of acetate and Crp-mediated repression of in the presence of glucose. Carbon flux through the glyoxylate shunt has been suggested to affect virulence, persistence, and antibiotic resistance of Mycobacterium tuberculosis. Therefore, it is important to understand the precise mechanism underlying the regulation of the gene encoding the key enzyme of the glyoxylate shunt. Using Mycobacterium smegmatis, this study revealed the regulation mechanism underlying induction of expression in M. smegmatis when the glyoxylate shunt is required. The conservation of the - and -acting regulatory elements related to regulation in both M. smegmatis and M. tuberculosis implies that a similar regulatory mechanism operates for the regulation of expression in M. tuberculosis.
耻垢分枝杆菌有两种异柠檬酸裂解酶（ICL）同工酶（MSMEG_0911 和 MSMEG_3706）。我们证明 ICL1（MSMEG_0911）是耻垢分枝杆菌中主要表达的 ICL，并在以乙酸盐或脂肪酸作为唯一碳源和能源的生长中发挥主要作用。证明在以乙酸盐生长时，基因在耻垢分枝杆菌中的表达相对于在葡萄糖上生长时被强烈上调。表明表达受 RamB 激活剂的正调控，并且使用 DNase I 足迹分析在的上游区域鉴定了三个 RamB 结合位点（RamBS1、RamBS2 和 RamBS3）。琥珀酰辅酶 A（琥珀酰-CoA）被证明增加了 RamB 与其结合位点结合的亲和力，并使 RamB 能够结合到 RamBS2，这是 RamB 介导的诱导表达的最重要位点。这些结果表明琥珀酰-CoA 是 RamB 的共诱导分子。我们的研究还表明，环腺苷酸受体蛋白（Crp1；MSMEG_6189）在葡萄糖存在下抑制耻垢分枝杆菌中 的表达。因此，相对于葡萄糖生长时弱表达，在作为唯一碳源的乙酸盐生长时强烈诱导表达可能是由于 RamB 介导的诱导和葡萄糖存在下 Crp 介导的 抑制的综合作用。糖异生途径中的碳通量已被认为影响结核分枝杆菌的毒力、持久性和抗生素耐药性。因此，了解糖异生途径关键酶编码基因调控的精确机制非常重要。本研究使用耻垢分枝杆菌，揭示了在需要糖异生途径时，该途径中关键酶编码基因在耻垢分枝杆菌中的表达诱导的调控机制。在耻垢分枝杆菌和结核分枝杆菌中，与 调控相关的-和-作用调控元件的保守性表明，对于结核分枝杆菌中 的表达调控，存在类似的调控机制。
