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 （sRNAs）在所有生物王国中都扮演着重要的角色。它们通常通过与特定的靶mrna配对来改变其翻译和/或稳定性或影响蛋白质的活性。小rna也是两种人类病原体单核细胞增生李斯特菌和金黄色葡萄球菌的调控网络的一部分，因此最有可能参与毒力机制。最近发现了许多srna。然而在革兰氏阴性菌中，主要是在大肠杆菌中，许多sRNAs已经被赋予了功能作用，而关于革兰氏阳性菌的知识状态是落后的。在这种情况下，在单核增生乳杆菌中存在五个几乎相同拷贝的sRNA LhrC仍然需要功能分类。有趣的是，LhrC sRNAs在血液中和β -内酰胺抗生素存在期间被上调，这表明其参与了毒力机制。在预期的研究项目中，LhrC的功能将主要在蛋白质组水平上进行探索，通过比较野生型和LhrC所有五个拷贝中缺失的突变体，采用全面的定量方法。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过程，不仅包括细胞质蛋白质组学，还包括膜和细胞外蛋白质部分。因此，确定的假定LhrC目标将得到支持和补充，应用基于生物信息学和生化捕捞的进一步战略。在该项目的第二部分，另一个完全未知功能的sRNA将采用类似的实验方法进行研究。金黄色葡萄球菌的这个sRNA被证明是sigmab调控的，表明在感染过程中可能起作用。该研究将有助于了解致病性革兰氏阳性菌中sRNA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.