金黄色葡萄球菌内源信号分子一氧化氮的功能及机制研究

一氧化氮 (NO) 作为信号分子首先在真核生物中被发现可由一氧化氮合酶 (NOS) 产生，后来发现某些革兰氏阳性菌如金黄色葡萄球菌可通过细菌来源的NOS (bNOS) 产生低浓度的NO。研究发现，细菌内源NO对自身耐药性、致病性以及压力应答有重要调控作用，但相关的分子机制研究甚少。我们前期在临床来源的VISA菌株中发现，bNOS敲除导致万古霉素抗性明显降低，自溶、生物被膜形成以及侵染巨噬细胞能力均增强；我们通过蛋白质组学鉴定了金黄色葡萄球菌中可被其内源NO亚硝基化修饰的蛋白质靶点，并获得了WalK和MgrA相应靶点的点突变菌株，发现该类突变均导致万古霉素敏感性增加、自溶增强。深入研究金黄色葡萄球菌bNOS的表达调控机制以及内源NO调控致病性和耐药性的分子机制，将从分子层面解析细菌内源NO在细菌信号传递中的重要性，并可能为金黄色葡萄球菌及其他细菌病原体的感染治疗提供新策略。

Nitric oxide (NO) is an important signaling molecule first found to be synthesized by NO synthases (NOS) in eukaryotes and functions as a regulator of physiology and immunity. Furthermore, several gram-positive bacteria including Staphylococcus aureus possess a bacterial NOS-like protein (bNOS), which synthesizes low concentration of NO. Studies suggest that bacterial endogenous NO plays an important role in regulating antibiotic resistance, virulence and stress responses, but the relevant molecular mechanisms remain unclear. Our previous work has indicated that bNOS deletion in a clinical vancomycin-intermediate S. aureus causes reduced vancomycin resistance and increased autolytic activity, biofilm formation, and phagocyte internalization. In addition, a mass spectrometry-based proteomic analysis was applied to identify the S-nitrosocysteine residues modified by endogenous NO in S. aureus strain XN108, and then the related cysteine point mutation of walR(C67S), walR(C33S) and mgrA(C12S) were constructed. All of these mutant strains exhibit increased vancomycin susceptibility and autolytic activity. This project aims to investigate the transcriptional factors controlling the expression of bNOS and the molecular mechanism of how endogenous NO regulating antibiotic resistance and virulence in S. aureus. This study will not only deepen our understanding of the significance of endogenous NO in bacterial signal transduction, but also provide novel strategies for the treatment of infection caused by S. aureus and other bacterial pathogens.