The Key Laboratory of Molecular Microbiology and Technology Ministry of Education, Nankai Universitygrid.216938.7, Tianjin, China.
Department of Microbiology, College of Life Sciences, Nankai Universitygrid.216938.7, Tianjin, China.
J Bacteriol. 2021 Sep 23;203(20):e0014521. doi: 10.1128/JB.00145-21. Epub 2021 Aug 2.
FleQ plays a crucial role in motility and biofilm formation by regulating flagellar and exopolysaccharide biosynthesis in Pseudomonas aeruginosa. It has been reported that the expression of FleQ is transcriptionally downregulated by the virulence factor regulator Vfr. Here, we demonstrated that a LysR-type transcriptional regulator, OsaR, is also capable of binding to the promoter region of and repressing its transcription. Through gel shift and DNase I footprinting assays, the OsaR binding site was identified and characterized as a dual LysR-type transcriptional regulator box (AT-N-AT-N-A-N-T). Mutation of the A-T palindromic base pairs in the promoter not only reduced the binding affinity of OsaR but also derepressed transcription . The OsaR binding site was found to cover the Vfr binding site; knockout of or separately exhibited no effect on the transcriptional level of ; however, expression was repressed by overexpression of or . Furthermore, simultaneously deleting both and resulted in an upregulation of , but it could be complemented by the expression of either of the two repressors. In summary, our work revealed that OsaR and Vfr function as two transcriptional repressors of that bind to the same region of but work separately. Pseudomonas aeruginosa is a widespread human pathogen, which accounts for serious infections in the hospital, especially for lung infection in cystic fibrosis and chronic obstructive pulmonary disease patients. P. aeruginosa infection is closely associated with its motility and biofilm formation, which are both under the regulation of the important transcription factor FleQ. However, the upstream regulatory mechanisms of have not been fully elucidated. Therefore, our research identifying a novel regulator of as well as new regulatory mechanisms controlling its expression will be significant for better understanding the intricate gene regulatory mechanisms related to P. aeruginosa virulence and infection.
FleQ 通过调节铜绿假单胞菌鞭毛和胞外多糖生物合成在运动性和生物膜形成中发挥关键作用。据报道，毒力因子调节因子 Vfr 转录下调 FleQ 的表达。在这里，我们证明了一种 LysR 型转录调节因子 OsaR 也能够结合 并抑制其转录。通过凝胶迁移和 DNase I 足迹实验，确定了 OsaR 结合位点，并将其特征化为双 LysR 型转录调节因子盒（AT-N-AT-N-A-N-T）。 突变 启动子中的 A-T 回文碱基对不仅降低了 OsaR 的结合亲和力，而且还解除了 的转录抑制。发现 OsaR 结合位点覆盖了 Vfr 结合位点； 或 单独缺失对 的转录水平没有影响；然而， 表达被 或 的过表达抑制。此外，同时缺失 和 导致 上调，但可以通过表达两个抑制剂中的任一个来补充。总之，我们的工作表明，OsaR 和 Vfr 作为两个结合 相同区域但单独工作的 转录抑制剂。铜绿假单胞菌是一种广泛存在的人类病原体，它在医院中引起严重感染，特别是在囊性纤维化和慢性阻塞性肺疾病患者的肺部感染中。铜绿假单胞菌感染与其运动性和生物膜形成密切相关，这两者都受到重要转录因子 FleQ 的调节。然而， 的上游调控机制尚未完全阐明。因此，我们研究确定了 作为一种新的调控因子，以及控制其表达的新调控机制，这对于更好地理解与铜绿假单胞菌毒力和感染相关的复杂基因调控机制将具有重要意义。
