Laboratory of Bacterial Pathogenesis, Department of Microbiology and Immunology, Shanghai Jiao Tong Universitygrid.16821.3c School of Medicine, Shanghai, China.
Department of Microbiology, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, China.
mBio. 2021 Oct 26;12(5):e0209921. doi: 10.1128/mBio.02099-21. Epub 2021 Sep 21.
The two-component system PhoP/PhoQ is essential for Salmonella enterica serovar Typhimurium virulence. Here, we report that PhoP is methylated extensively. Two consecutive glutamate (E) and aspartate (D)/E residues, i.e., E8/D9 and E107/E108, and arginine (R) 112 can be methylated. Individual mutation of these above-mentioned residues caused impaired phosphorylation and dimerization or DNA-binding ability of PhoP to a different extent and led to attenuated bacterial virulence. With the help of specific antibodies recognizing methylated E8 and monomethylated R112, we found that the methylation levels of E8 or R112 decreased dramatically when bacteria encountered low magnesium, acidic pH, or phagocytosis by macrophages, under which PhoP can be activated. Furthermore, CheR, a bacterial chemotaxis methyltransferase, was identified to methylate R112. Overexpression of decreased PhoP activity but increased PhoP stability. Together, the current study reveals that methylation plays an important role in regulating PhoP activities in response to environmental cues and, consequently, modulates Salmonella virulence. Posttranslational modifications (PTMs) play an important role in regulating enzyme activities, protein-protein interactions, or DNA-protein recognition and, consequently, modulate many biological functions. We demonstrated that PhoP, the response regulator of PhoP/PhoQ two-component system, could be methylated on several evolutionally conserved amino acid residues. These amino acid residues were crucial for PhoP phosphorylation or dimerization, DNA-binding ability of PhoP, and Salmonella virulence. Interestingly, methylation negatively regulated the activity of PhoP. A bacterial chemotaxis methyltransferase CheR was involved in PhoP methylation. Methylation of PhoP could stabilize it in an inactive conformation. Our work provides a more informative depiction of PhoP PTM and markedly improves our understanding of the coordinate regulation of bacterial chemotaxis and virulence.
双组分系统 PhoP/PhoQ 对鼠伤寒沙门氏菌的毒力至关重要。在这里，我们报告 PhoP 广泛甲基化。两个连续的谷氨酸 (E) 和天冬氨酸 (D)/E 残基，即 E8/D9 和 E107/E108，以及精氨酸 (R)112 可以被甲基化。这些残基的单个突变导致磷酸化和二聚化或 PhoP 与 DNA 结合能力不同程度受损，导致细菌毒力减弱。借助特异性抗体识别甲基化的 E8 和单甲基化的 R112，我们发现当细菌遇到低镁、酸性 pH 值或巨噬细胞吞噬时，E8 或 R112 的甲基化水平会显著降低，此时 PhoP 可以被激活。此外，细菌趋化性甲基转移酶 CheR 被鉴定为甲基化 R112。PhoP 活性的增加，但 PhoP 稳定性增加。总之，本研究揭示了甲基化在调节 PhoP 活性以响应环境信号方面的重要作用，从而调节沙门氏菌的毒力。
双组分系统 PhoP/PhoQ 对鼠伤寒沙门氏菌的毒力至关重要。在这里，我们报告 PhoP 广泛甲基化。两个连续的谷氨酸 (E) 和天冬氨酸 (D)/E 残基，即 E8/D9 和 E107/E108，以及精氨酸 (R)112 可以被甲基化。这些残基的单个突变导致磷酸化和二聚化或 PhoP 与 DNA 结合能力不同程度受损，导致细菌毒力减弱。借助特异性抗体识别甲基化的 E8 和单甲基化的 R112，我们发现当细菌遇到低镁、酸性 pH 值或巨噬细胞吞噬时，E8 或 R112 的甲基化水平会显著降低，此时 PhoP 可以被激活。此外，细菌趋化性甲基转移酶 CheR 被鉴定为甲基化 R112。PhoP 活性的增加，但 PhoP 稳定性增加。总之，本研究揭示了甲基化在调节 PhoP 活性以响应环境信号方面的重要作用，从而调节沙门氏菌的毒力。
