我们前期发现金黄色葡萄球菌SprC（致病岛小RNA C）能结合efb mRNA下调Efb表达，但两者结合的核苷酸及影响此结合和结合后mRNA稳定性的因素未明。Efb是金黄色葡萄球菌重要毒力因子，能抑制宿主天然免疫应答。据研究：细菌sRNA主要通过结合mRNA核糖体结合位点（RBS）影响靶蛋白翻译，RNA伴侣Hfq能促进两者结合，结合后可能触发RNase III活性影响mRNA稳定性。故推测SprC下调Efb表达可能是通过结合efb mRNA RBS实现的；Hfq和RNase III可能促进两者结合及降低mRNA稳定性。本项目拟以分子生物学、生物信息学及生物化学等方法阐明SprC下调Efb表达的分子机制，探讨Hfq在SprC与efb mRNA结合中的作用及RNase III对efb mRNA 稳定性的影响。研究结果有助于深入了解金黄色葡萄球菌致病机理，为改进该菌感染的预防和治疗策略提供线索。

In the previous experiments, we found SprC (small pathogenicity island RNA C) down-regulated the Efb expression by binding to efb mRNA in Staphylococcus aureus (S. aureus). However, the nucleotides interacting between SprC and efb mRNA and the factors affecting the combination of the both RNA and the stability of mRNA after the binding are not to be elucidated. Efb is the significant virulence factor of S. aureus. It can inhibit the host innate immune responses. The previous studies have shown most of bacterial sRNAs regulate the proteins expression in translation level by the binding to the ribosome-binding site (RBS) of the target mRNA. The RNA chaperone Hfq can promote the duplex formation between sRNA and target mRNA. This binding may trigger the hydrolysis activity of RNase III affecting the stability of target mRNA. Therefore, we speculate the mechanism of SprC down-regulating the expression of Efb can be the antisense binding to the RBS of efb mRNA. Hfq and RNase III can produce a marked effect in promoting the binding of SprC to efb mRNA and reducing the stability of efb mRNA, respectively. In this investigation we will elucidate the molecular mechanism of SprC down-regulating Efb expression, and explore the roles of Hfq in promoting the SprC-efb mRNA duplex formation and RNase III in affecting the stability of efb mRNA by the methods of molecular biology, bioinformatics and biochemistry. This study will help us deeply understand the pathogenesis of S. aureus, and give a clue for improving the prevention and therapy strategies of S. aureus infections.

金黄色葡萄球菌是困扰全球公共卫生及人类健康的重要病原菌，其引起的各种临床感染与该菌表达的多种毒力因子密切相关，而这些毒力因子的表达调控在细菌致病机制中发挥着核心作用。当前，细菌的耐药性发展迅速，且不断增强，使得现有的抗菌药物难以满足临床抗感染需求。而针对细菌毒力因子表达的重要调节因子设计的药物可以减弱细菌致病力，减少抗菌药物新耐药机制形成的压力，因而成为研发新型抗感染药物的新方向。sRNA是细菌调节毒力基因表达的一类关键分子，但对涉及sRNA 调控金黄色葡萄球菌毒力因子表达调控机制的了解仍然处于起步阶段。本项目研究内容主要为，在金黄色葡萄球菌非编码小RNA C（SprC）能下调免疫逃避蛋白胞外纤维蛋白原结合蛋白Efb表达基础上研究SprC调控Efb表达的分子机制。分子生物学（如同源重组、RACE技术、northern blot、qRT-PCR、western blot,等）、生物信息学等方法的实验结果显示，SprC通过其 5′-非编码区（5′-UTR）的58个核苷酸与efb mRNA 5′-UTR的78个核苷酸反义结合形成二聚体，在翻译水平负调控Efb蛋白表达。SprC-efb mRNA 复合物不能触发双链特异性核酸内切酶III（RNase III）的水解活性降低efb mRNA 稳定性。RNA伴侣Hfq在SprC调控Efb表达调控过程中也不发挥作用。本研究结果阐明了SprC在Efb表达调控中所起的作用及其分子生物学作用机制，为丰富该菌毒力调节的网络理论、深入了解金黄色葡萄球菌致病机理及有效的干预金黄色葡萄球菌毒力基因表达调控环节改进金黄色葡萄球菌感染的预防和治疗策略提供线索。