Characterization of molecular mechanisms of resistance to antimicrobial fatty acids in the staphylococci

背景 大约25%的人类长期受到金黄色葡萄球菌的侵袭。人类的首选定居部位是前鼻孔、腋窝、会阴、手、胸部和四肢，而牛的则是乳头皮肤。因此，它在皮肤上附着的能力是在人类和动物中传播的重要媒介。然而，为了在皮肤上生存，金黄色葡萄球菌必须抵抗皮肤的天然防御屏障，包括具有抗菌活性的长链不饱和游离脂肪酸(UFFAs)。油酸、棕榈油酸、亚油酸和花生四烯酸等uFFA对金黄色葡萄球菌的生长有抑制作用。事实上，这些杀菌脂肪酸是抵御微生物定居的第一道防线的一部分。沙培酸是从皮肤皮脂腺分泌的甘油三酯中释放出来的，在特应性皮炎患者中，其分泌存在缺陷，导致金黄色葡萄球菌携带量增加，对金黄色葡萄球菌感染的易感性增加。鼻腔分泌物还含有亚油酸、花生四烯酸和棕榈油酸，而葡萄球菌脓肿含有大量的亚油酸。金黄色葡萄球菌是皮肤和鼻子的有效定殖者，尽管暴露在抗微生物脂肪酸中 假设 我们假设金黄色葡萄球菌已经或能够进化出对杀菌脂肪酸的抗性机制。我们进一步假设，抗性机制的识别将有助于深入了解uFFA的生物学靶点。 进展 我的实验室已经确定了一种迄今未知的对uFFAs的抗性机制，涉及RNA降解酶体的膜RNaseY的单核苷酸多态性。 目标 阐明RNaseY SNPs与脂肪酸耐药性之间的联系是这项研究计划的首要目标。 目的1：明确RNaseY基因多态性菌株的脂肪酸抗性机制。利用几条平行的研究路线，这一目标将表征RNaseY依赖的脂肪酸抗性机制。 目的2：RNaseY变异体的功能特性。这一目标将表征RNaseY已知功能中与SNP相关的功能增益变化。 目标3：确定SNPs与提高对脂肪酸的抵抗力之间的联系。RNaseY中的氨基酸变化将详细描述它们增强对脂肪酸的抵抗力的能力。 目的4：鉴定和鉴定耐额外脂肪酸结构的金黄色葡萄球菌变异株。这将提供关于脂肪酸结构对金黄色葡萄球菌细胞作用的特定机制的有价值的信息。 意义 这一以发现为基础的研究计划将解决有关金黄色葡萄球菌对杀菌脂肪酸的生理反应以及金黄色葡萄球菌对这些化合物产生抗药性的能力等重要而独特的问题。

Background Approximately 25% of humans are persistently colonized by S. aureus. Preferred sites of colonization of humans are the anterior nares, axillae, perineum, hands, chest and limbs, while in cattle it is the teat skin. Accordingly, its ability to persist on skin is an important mediator of transmission in both humans and animals. Yet, to persist on skin, S. aureus must resist innate defense barriers of the skin including long-chain unsaturated free fatty acids (uFFAs) which possess antimicrobial activity. uFFAs such as sapienic acid, palmitoleic acid, linoleic acid and arachidonic acid inhibit the growth of S. aureus. Indeed, these microbicidal fatty acids are part of the first line of defense against colonization by microbes. Sapienic acid is released from triglycerides secreted by the sebaceous glands in the skin and its secretion is defective in individuals with atopic dermatitis, resulting in increased carriage of S. aureus and increased susceptibility to infection by S. aureus. Nasal secretions also contain linoleic, arachidonic and palmitoleic acids, while staphylococcal abscesses contain abundant quantities of linoleic acid. S. aureus is an effective colonizer of the skin and nose, despite exposure to antimicrobial fatty acids Hypotheses We hypothesize that S. aureus has evolved, or can evolve, mechanisms of resistance to microbicidal fatty acids. We further hypothesize that identification of resistance mechanisms will shed significant insight into the biological targets of uFFAs. Progress My laboratory has identified a heretofore unknown resistance mechanism against uFFAs, involving single nucleotide polymorphisms in RNaseY, a membrane of the RNA degradasome. Aims Elucidating the link between RNaseY SNPs and resistance to fatty acids are the overarching aims of this research program. Aim 1: Define the fatty acid resistance mechanism in strains containing RNaseY polymorphisms. Using several parallel lines of investigation, this aim will characterize the RNaseY-dependent mechanism of resistance to fatty acids. Aim 2: Functional characterization of RNaseY variants. This aim will characterize SNP-associated, gain-of-function changes in known functions of RNaseY. Aim 3: Define the link between SNPs and elevated resistance to fatty acids. Amino acid changes in RNaseY will be characterized in detail for their ability to confer increased resistance to fatty acids. Aim 4: Identification and characterization of S. aureus variants resistant to additional fatty acid structures. This will yield valuable information on fatty acid-structure specific mechanisms of action of fatty acids on the S. aureus cell. Significance This discovery-based research program will address important and unique questions concerning physiological responses of S. aureus to microbicidal fatty acids, and the ability of S. aureus to evolve resistance to these compounds.