Department of Microbiology, Bose Institute, P-1/12 C.I.T. Scheme VIIM, Kolkata 700054, India.
Microbiology (Reading). 2022 May;168(5). doi: 10.1099/mic.0.001192.
A previous study reported that the (Msm) protein MSMEG_2295 is a repressor controlling the expression of several genes, including that for MSMEG_5125, a putative isoprenoid binding protein belonging to the YceI family, and DinB2, a DNA damage repair enzyme. This repressor is encoded by the first gene of the operon that also expresses the gene for DinB2. Targeted inhibition of MSMEG_5125 using CRISPRi technology resulted in a significant loss of Msm's respiratory activity and viability. Since this protein has been predicted to be an isoprenoid binding protein, we suspected a role of menaquinones, which are isoprenoid naphthoquinones, in the observed phenomenon. Accordingly, we tested whether MSMEG_5125's deficiency-induced lethality could be reversed by adding menaquinone. The result was positive, implying cooperation between MSMEG_5125 and menaquinone in bringing about respiration. Inhibition of expression led to the induction of and , two hallmark genes of the MSMEG_2295 regulon. This result suggests that when MSMEG_5125 becomes limiting, a feedback-loop derepresses the MSMEG_2295 regulon genes, including its own. Interestingly, menaquinone functioned as an inducer of , indicating that it is likely to mediate the feedback mechanism. This result also strengthens our hypothesis that the functions of menaquinone and MSMEG_5125 are interrelated. Menaquinone also induced the MSMEG_2295-controlled operon () not induced following the inactivation of . Therefore, the activation mechanism of MSMEG_2295-regulated genes may not be the same for all, although derepression is likely to be a common feature. , menaquinone abolished MSMEG_2295's DNA binding activity by interacting with it, confirming its role as an inducer. Therefore, a menaquinone-MSMEG_5125-regulated gene expression circuit controls Msm respiration and possibly oxidative stress-induced DNA damage repair.
先前的研究报告指出，（Msm）蛋白 MSMEG_2295 是一种阻遏物，可控制包括 MSMEG_5125 在内的多个基因的表达，MSMEG_5125 是一种假定的异戊烯基结合蛋白，属于 YceI 家族，以及 DinB2，一种 DNA 损伤修复酶。该阻遏物由操纵子的第一个基因编码，该基因也表达 DinB2 基因。使用 CRISPRi 技术靶向抑制 MSMEG_5125 导致 Msm 的呼吸活性和活力显著丧失。由于该蛋白已被预测为异戊烯基结合蛋白，因此我们怀疑类异戊二烯萘醌（menaquinones）在观察到的现象中起作用。因此，我们测试了 MSMEG_5125 缺乏诱导的致死性是否可以通过添加menaquinone 来逆转。结果为阳性，表明 MSMEG_5125 和 menaquinone 之间存在协同作用，从而促进呼吸作用。表达抑制导致 和 的诱导，这两个基因是 MSMEG_2295 调控子的标志性基因。这一结果表明，当 MSMEG_5125 变得有限时，反馈环会解除 MSMEG_2295 调控子基因的阻遏，包括其自身。有趣的是，menaquinone 作为 的诱导物发挥作用，表明它可能介导反馈机制。这一结果也进一步证实了我们的假设，即 menaquinone 和 MSMEG_5125 的功能是相互关联的。menaquinone 还诱导了 MSMEG_2295 控制的操纵子 （），而不是在 失活后诱导。因此，尽管去阻遏可能是一个共同特征，但 MSMEG_2295 调控基因的激活机制可能并非对所有基因都相同。此外，menaquinone 通过与 MSMEG_2295 相互作用，使其 DNA 结合活性丧失，证实了其作为诱导物的作用。因此，一个menaquinone-MSMEG_5125 调控的基因表达回路控制 Msm 的呼吸作用，并可能控制氧化应激诱导的 DNA 损伤修复。
