Exploring the physiological role and mode of action of the small RNA LhrC of Listeria monocytogenes and a recently discovered sigmaB-regulated sRNA of Staphylococcus aureus to check for their possible involvement in pathogenicity

小RNA(SRNA)在所有生命王国中都扮演着重要的角色。它们通常通过与特定的靶mRNAs配对来改变它们的翻译和/或稳定性，或者影响蛋白质的活性。小RNA也是单核细胞增多性李斯特菌和金黄色葡萄球菌这两种人类病原体调控网络的一部分，因此很可能参与毒力机制。最近发现了大量的sRNA。然而，在革兰氏阴性菌中，首先是在大肠杆菌中，许多sRNA被分配了一个功能角色，但关于革兰氏阳性菌的知识状况却落后了。在这种情况下，单核细胞增多性李斯特氏菌中存在于五个几乎相同拷贝中的sRNA LHRC仍然需要进行功能分类。有趣的是，LHRC sRNAs在血液中和在β-内酰胺类抗生素存在时上调，表明参与了毒力机制。在预期的研究项目中，LHRC的功能将主要通过比较野生型和突变型在LHRC的所有五个拷贝中被删除的综合定量方法在蛋白质组水平上进行。更准确地说，蛋白质组学分析将涉及基于14N/15N同位素标记蛋白质的代谢标记程序，不仅包括细胞质蛋白质组，还包括膜和胞外蛋白质组分。因此，将采用进一步的基于生物信息学和生化捕鱼的战略来支持和补充已确定的低风险和可持续发展目标。在该项目的第二部分中，将使用类似的实验方法来研究另一种完全未知功能的sRNA。金黄色葡萄球菌的这个sRNA被证明是受SigmaB调控的，这表明在感染过程中可能起到了作用。这项研究将有助于理解sRNA在致病性革兰氏阳性菌中的作用，并可能为新的抗菌策略提供基础。

Small RNAs (sRNAs) play an important role in all kingdoms of life. They often act by pairing with specific target mRNAs to change their translation and/or stability or act to influence the activity of proteins. Small RNAs are also part of regulatory networks of the two human pathogens Listeria monocytogenes and Staphylococcus aureus and therefore most likely involved in virulence mechanisms. Numerous sRNAs have been discovered recently. Whereas in Gram-negative bacteria, foremost in Escherichia coli, a functional role has been assigned to many sRNAs, the state of knowledge concerning Gram-positive bacteria is lagging behind. In this context, the sRNA LhrC which is present in five almost identical copies in L. monocytogenes still requires functional classification. Interestingly, LhrC sRNAs have been shown to be up-regulated in blood and during the presence of beta-lactam antibiotics pointing to an involvement in virulence mechanisms. In the intended research project LhrC function will be explored primarily on proteome level by comparison of wild type and a mutant deleted in all five copies of LhrC in a comprehensive quantitative approach. More precisely the proteomics analysis will involve a metabolic labeling procedure based on 14N/15N isotopically labeled proteins, and will comprise not only the cytosolic proteome but also the membrane and extracellular protein fraction. Thereby identified putative LhrC targets will be backed up and complemented applying further strategies based on bioinformatics and biochemical fishing. In a second part of the project another sRNA of completely unknown function will be investigated employing a comparable experimental approach. This sRNA of S. aureus was shown to be sigmaB-regulated indicating a possible role during infection. The proposed research will contribute to the understanding of sRNA action in pathogenic Gram-positive bacteria and may provide a basis for new antimicrobial strategies.