Post-translational regulation of alpha PSM production in Staphylococcus aureus by the small RNA Teg41

项目概要/摘要 金黄色葡萄球菌既是人类的寄生虫，也是一种高度危险的细菌病原体。S.金黄色 致病机制是由大量分泌的和细胞壁相关的毒力因子介导的，包括 一些有效的细胞溶解肽称为酚可溶性调节蛋白（PSM）。PSM是两亲性的， 根据其分类大小不同的螺旋肽。α型PSM的大小约为22个氨基酸 近年来一直是研究的热点。他们被指控参与了 社区获得性甲氧西林耐药链球菌的毒力潜力。金黄色葡萄球菌（CA-MRSA）菌株，特别是那些 美国300系列而α PSM在S.金黄色葡萄球菌感染已被广泛研究， 在我们对它们如何在细菌细胞中产生的理解方面仍然存在重大差距。有五 大多数S.金黄色葡萄球菌菌株（PSMα1-4和δ-毒素）。PSMα1-4编码在 相同的多顺反子转录本（αPSM转录本），但研究表明，这四种转录本的相对水平 肽的变化相当大。PSMα4（位于转录物的3'末端）通常是最丰富的， 而PSMα3（四种肽中最有效的）通常丰度最低。αPSM的这种变化 丰度暗示了转录后调节。该项目的长期目标是 了解有助于在S中产生αPSM的分子机制。金黄色。在本提案中，我们 将专门研究小RNA Teg 41对αPSM产生和毒力的贡献。 初步研究表明，Teg 41在转录后水平上对αPSM的产生产生有积极影响。我们 将研究（i）哪些PSMα1-4肽受Teg 41调节，（ii）Teg 41介导的调节是否 通过与αPSM转录物的直接碱基配对促进，以及（iii）在αPSM产生的哪个阶段，以及如何， teg 41发挥了它的影响力。为了研究这三个目标，我们将使用体外，体内，遗传， 生物化学和分子生物学方法。我们还将利用一些尖端技术， 高通量DNA测序（SHAPE-seq，miR-CATCH和Ribo-seq）。这项研究的结果可以 对人类健康有直接影响。破坏Teg 41介导的α PSM产生可以显著降低 S.金黄色葡萄球菌，因此可能代表治疗干预的新靶点。

PROJECT SUMMARY/ABSTRACT Staphylococcus aureus is both a commensal of humans and a highly dangerous bacterial pathogen. S. aureus pathogenesis is mediated by a large repertoire of secreted and cell wall-associated virulence factors, including a number of potent cytolytic peptides called phenol soluble modulins (PSMs). PSMs are amphipathic, alpha helical peptides that vary in size depending on their classification. The α type PSMs are ~22 amino acids in size and have been the focus of intense study in recent years. They have been implicated in contributing to the high virulence potential of community-acquired methicillin resistant S. aureus (CA-MRSA) strains, in particular those of the USA300 lineage. While the role of αPSMs in S. aureus infection has been extensively investigated, significant gaps still exist in our understanding of how they are produced in the bacterial cell. There are five αPSM peptides produced by most S. aureus strains (PSMα1-4 and the δ-toxin). PSMα1-4 are encoded within the same polycistronic transcript (the αPSM transcript), yet studies have shown that the relative levels of the four peptides vary considerably. PSMα4 (located at the 3' end of the transcript) is commonly the most abundant, while PSMα3 (the most potent of the four peptides) is typically the least abundant. This variation in αPSM abundance is suggestive of post-transcriptional regulation. The long-term objective of this project is to understand the molecular mechanism(s) that contribute to αPSM production in S. aureus. In this proposal, we will specifically investigate the contribution of the small RNA Teg41 to αPSM production and virulence. Preliminary studies show that Teg41 positively influences αPSM production at the post-transcription level. We will investigate (i) which of the PSMα1-4 peptides is/are regulated by Teg41, (ii) if Teg41-mediated regulation is facilitated by direct base pairing with the αPSM transcript, and (iii) at what stage in αPSM production, and how, Teg41 exerts its influence. To investigate these three aims, we will use a combination of in vitro, in vivo, genetic, biochemical, and molecular biology approaches. We will also utilize a number of cutting edge techniques based on high throughput DNA sequencing (SHAPE-seq, miR-CATCH, and Ribo-seq). The results from this study could have direct implications for human health. Disrupting Teg41-mediated αPSM-production could dramatically lower the virulence potential of S. aureus and therefore may represent a novel target for therapeutic intervention.