A Clostridium difficile-Specific, Gel-Forming Protein Required for Optimal Spore Germination
A Clostridium difficile-Specific, Gel-Forming Protein Required for Optimal Spore Germination
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DOI：
10.1128/mbio.02085-16
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发表时间：
2017-01-01
期刊：
影响因子：
6.4
通讯作者：
Shen, Aimee
中科院分区：
文献类型：
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作者：
Donnelly, M. Lauren;Li, William;Shen, Aimee
艰难梭菌是一种革兰氏阳性芽孢形成的专性厌氧菌，是全世界抗生素相关性腹泻的主要原因。为了使艰难梭菌启动感染，其耐氧孢子形式必须在哺乳动物宿主的肠道中发芽。虽然几乎所有的孢子形成生物都使用跨膜萌发受体来触发萌发，但艰难梭菌使用假蛋白酶CspC来感知胆盐萌发。CspC激活相关的枯草杆菌样蛋白酶CspB，然后CspB蛋白水解激活皮层水解酶SleC。活化的SleC降解保护性孢子皮层，这是发芽进行的必要步骤。由于CspC并入孢子也依赖于CspA，一个相关的假蛋白酶结构域，Csp家族蛋白在萌发中起关键作用。然而，Csps如何与孢子结合仍是未知的。在这项研究中，我们证明了CspC、CspB和CspA发芽调节因子在孢子中的结合依赖于CD0311（更名为GerG），这是一种以前未被表征的假设蛋白。gerG孢子中Csps水平的降低与对胆盐萌发物的反应性降低和单孢子萌发试验中萌发异质性的增加有关。有趣的是，GerG中心区域富含天冬酰胺的重复序列促进体外自发凝胶形成，尽管它们对于GerG介导的发芽控制是必不可少的。由于GerG仅存在于艰难梭菌中，我们的研究结果表明，利用GerG功能可能是开发艰难梭菌特异性抗感染疗法的有希望的途径。芽孢形成细菌艰难梭菌是卫生保健相关感染的主要原因。虽然一部分抗生素可以治疗艰难梭菌感染（CDI），但CDI疾病易感性的主要决定因素是先前的抗生素暴露，因为它减少了多种微生物群赋予的定植抗性。因此，减少肠道微生物群扰动的治疗方法应该更有效地减少cdi及其复发，这是疾病并发症的主要来源。鉴于孢子萌发是艰难梭菌启动感染的必要条件，并且艰难梭菌使用独特的途径启动萌发，抑制不同发芽要素的方法可以选择性地防止艰难梭菌疾病复发。在这里，我们确定GerG是一种艰难梭菌特异性蛋白，控制发芽信号蛋白进入孢子。由于gerG突变孢子表现出萌发缺陷，并且对萌发物反应较弱，因此gerG可能是开发抗CDI治疗药物的有希望的靶点。
Clostridium difficile is a Gram-positive spore-forming obligate anaerobe that is a leading cause of antibiotic-associated diarrhea worldwide. In order for C. difficile to initiate infection, its aerotolerant spore form must germinate in the gut of mammalian hosts. While almost all spore-forming organisms use transmembrane germinant receptors to trigger germination, C. difficile uses the pseudoprotease CspC to sense bile salt germinants. CspC activates the related subtilisin-like protease CspB, which then proteolytically activates the cortex hydrolase SleC. Activated SleC degrades the protective spore cortex layer, a step that is essential for germination to proceed. Since CspC incorporation into spores also depends on CspA, a related pseudoprotease domain, Csp family proteins play a critical role in germination. However, how Csps are incorporated into spores remains unknown. In this study, we demonstrate that incorporation of the CspC, CspB, and CspA germination regulators into spores depends on CD0311 (renamed GerG), a previously uncharacterized hypothetical protein. The reduced levels of Csps in gerG spores correlate with reduced responsiveness to bile salt germinants and increased germination heterogeneity in single-spore germination assays. Interestingly, asparagine-rich repeat sequences in GerG's central region facilitate spontaneous gel formation in vitro even though they are dispensable for GerG-mediated control of germination. Since GerG is found exclusively in C. difficile, our results suggest that exploiting GerG function could represent a promising avenue for developing C. difficile-specific anti-infective therapies.IMPORTANCE The spore-forming bacterium Clostridium difficile is a leading cause of health care-associated infections. While a subset of antibiotics can treat C. difficile infections (CDIs), the primary determinant of CDI disease susceptibility is prior antibiotic exposure, since it reduces the colonization resistance conferred by a diverse microflora. Thus, therapies that minimize perturbations to the gut microbiome should be more effective at reducing CDIs and their recurrence, the main source of disease complications. Given that spore germination is essential for C. difficile to initiate infection and that C. difficile uses a unique pathway to initiate germination, methods that inhibit distinct elements of germination could selectively prevent C. difficile disease recurrence. Here, we identify GerG as a C. difficile-specific protein that controls the incorporation of germinant signaling proteins into spores. Since gerG mutant spores exhibit germination defects and are less responsive to germinant, GerG may represent a promising target for developing therapeutics against CDI.