Mechanisms of (p)ppGpp action in Staphylococcus aureus

细菌的严格反应以信使分子(P)ppGpp的合成为特征，它干扰许多细胞过程，包括转录、复制和翻译。然而，(P)ppGpp积累导致的表型结果因物种而异，并可由不同的潜在机制调节。根据物种的不同，严格的反应对不同的生物学过程至关重要，包括分化、生物膜形成、抗生素耐受性、次生代谢物的产生或毒力。现在很清楚的是，最初在大肠杆菌中表现出的严格反应与在FirMicuts中的反应之间存在根本的区别。在参与信使合成和降解的主要参与者以及(P)ppGpp的下游影响方面都存在差异。我们阐明了金黄色葡萄球菌的严格反应的主要参与者和机制。在金黄色葡萄球菌中，严格的反应在毒力、吞噬小体逃逸和抗生素耐受性方面起着重要作用。从我们之前的工作中，我们确定了几个在(P)ppGpp合成时立即激活的基因。严格反应过程中基因激活的分子机制尚不清楚，但这种基因激活对于在胁迫条件下的生存很重要，可能是以一种物种特有的方式。我们的目标是揭示(P)ppGpp依赖的基因激活的分子机制和功能后果。此外，我们将开发(P)ppGpp传感器来评估可能的随机(P)ppGpp合成在单细胞水平上的应激耐受中的作用。最近，关于REL、RelQ和RelP的(P)ppGpp合成酶活性的主要结构方面的研究从Firmicuts获得。然而，许多已提出的机制尚未在活体生物体中得到证实。我们将建立方法来深入了解金黄色葡萄球菌中(P)ppGpp合成酶/水解酶活性的变构调节的功能后果。通过这个项目，我们将阐明导致金黄色葡萄球菌特定严格反应表型的潜在分子机制。

The stringent response of bacteria is characterised by the synthesis of the messenger molecule (p)ppGpp, which interferes with many cellular processes, including transcription, replication and translation. However, the phenotypic consequences resulting from (p)ppGpp accumulation vary among species and can be mediated by different underlying mechanisms. Depending on the species, the stringent response is crucial for diverse biological processes, including differentiation, biofilm formation, antibiotic tolerance, production of secondary metabolites or virulence. It is now clear that there are fundamental differences between the stringent response initially characterised in E. coli and the response in Firmicutes. Differences are seen in both the main players involved in synthesis and degradation of the messengers and in the downstream effects of (p)ppGpp. We elucidated major players and mechanisms of the stringent response in Staphylococcus aureus. In S. aureus the stringent response plays important roles in virulence, phagosomal escape and antibiotic tolerance. From our previous work we identified several genes that are immediately activated upon (p)ppGpp synthesis. The molecular mechanisms of gene activation during the stringent response are not well understood, but such gene activation is important for survival under stress conditions probably in a species specific manner. We aim to unravel the molecular mechanisms and functional consequences of (p)ppGpp-dependent gene activation. Furthermore, we will develop (p)ppGpp sensors to assess the role of possibly stochastic (p)ppGpp synthesis in stress tolerance on the single cell level. Recently, major structural insights concerning the (p)ppGpp synthetase activities of Rel, RelQ and RelP from Firmicutes were gained. However, many of the proposed mechanisms are not yet confirmed in vivo in living organisms. We will establish methods to gain insights into the functional consequence of the allosteric regulation of the (p)ppGpp synthetase/hydrolase activities in S. aureus. With this project we will elucidate the underlying molecular mechanisms leading to the specific stringent response phenotype of S. aureus.