猪链球菌（SS）是一种革兰氏阳性致病菌，可引致猪及人脑膜炎等多种疾病，目前对其脑膜炎形成机制还缺乏了解。近年来研究发现细菌小RNA是一种新型毒力调控因子，但尚未有其调控SS毒力机制的报道。申请者前期发现SS小RNA rss04在猪脑脊液中上调表达，促进猪和小鼠脑膜炎形成，该基因缺失后SS毒力降低。本项目通过探讨rss04在SS与小鼠脑微血管内皮细胞和免疫活性细胞互作中发挥的生物学功能，并分析其对SS荚膜形成能力和在猪脑脊液中增殖能力的影响，揭示rss04促进脑膜炎形成的途径；采用MS2识别序列纯化技术，结合转录组、蛋白组及生物信息学等，鉴定rss04靶位与RNA结合蛋白；分析rss04对靶位稳定性及翻译水平的影响，确定与靶位结合区域和RNA结合蛋白的功能，阐明rss04调控靶位机制。该研究有助于理解SS引致脑膜炎机制，也为革兰氏阳性菌RNA结合蛋白功能及小RNA调控靶位机制提供新的参考。

Streptococcus suis (SS) is considered to be a major Gram-positive pathogen worldwide in the swine industry and is also an important zoonotic agent for human. Meningitis is one of the most common features of the infection, but little is known about the mechanisms of SS meningitis. Recent studies have revealed that small RNAs (sRNAs) has emerged as key regulators of virulence in several bacteria. However, at present, there is no report about the mechanism how sRNAs regulate SS virulence. In previous study, we showed that SS sRNA rss04 was up-regulated in pig cerebrospinal fluid and rss04 deletion mutant attenuates SS virulence in a zebrafish infection model. Furthermore, rss04 contributes to SS meningitis in mouse and pig infction models. In this project, to find out the way by which rss04 contributes to meningitis, we will explore the functions of rss04 during SS interaction with mouse brain microvascular endothelial cell, microglia and neutrophil, and evaluate the effects of rss04 on SS capsule production and replication in pig cerebrospinal fluid. MS2 tagging purification, in combination with transcriptome and proteome analysis and bioinformatics prediction, will be performed to identify the rss04 targets involved in meningitis process and RNA binding proteins. To reveal the regulation mechanisms of rss04 on targets, the effects of rss04 on targets mRNA stability and translation will be evaluated, and the binding regions between rss04 and targets and roles of RNA binding proteins in regulation will be determined. These findings not only contribute to understanding the SS meningitis mechanism, but also increase the knowledges of the RNA binding proteins involved in sRNAs regulation and the mechanism of sRNAs regulation in Gram-positive bacteria.