Wisconsin Energy Institute, Great Lakes Bioenergy Research Center, University of Wisconsin-Madison, Madison, Wisconsin, United States of America.
Laboratory of Genetics, University of Wisconsin-Madison, Madison, Wisconsin, United States of America.
PLoS Genet. 2022 Jun 29;18(6):e1010270. doi: 10.1371/journal.pgen.1010270. eCollection 2022 Jun.
Bacterial two-component systems (TCSs) often function through the detection of an extracytoplasmic stimulus and the transduction of a signal by a transmembrane sensory histidine kinase. This kinase then initiates a series of reversible phosphorylation modifications to regulate the activity of a cognate, cytoplasmic response regulator as a transcription factor. Several TCSs have been implicated in the regulation of cell cycle dynamics, cell envelope integrity, or cell wall development in Escherichia coli and other well-studied Gram-negative model organisms. However, many α-proteobacteria lack homologs to these regulators, so an understanding of how α-proteobacteria orchestrate extracytoplasmic events is lacking. In this work we identify an essential TCS, CenKR (Cell envelope Kinase and Regulator), in the α-proteobacterium Rhodobacter sphaeroides and show that modulation of its activity results in major morphological changes. Using genetic and biochemical approaches, we dissect the requirements for the phosphotransfer event between CenK and CenR, use this information to manipulate the activity of this TCS in vivo, and identify genes that are directly and indirectly controlled by CenKR in Rb. sphaeroides. Combining ChIP-seq and RNA-seq, we show that the CenKR TCS plays a direct role in maintenance of the cell envelope, regulates the expression of subunits of the Tol-Pal outer membrane division complex, and indirectly modulates the expression of peptidoglycan biosynthetic genes. CenKR represents the first TCS reported to directly control the expression of Tol-Pal machinery genes in Gram-negative bacteria, and we predict that homologs of this TCS serve a similar function in other closely related organisms. We propose that Rb. sphaeroides genes of unknown function that are directly regulated by CenKR play unknown roles in cell envelope biosynthesis, assembly, and/or remodeling in this and other α-proteobacteria.
细菌双组分系统（TCS）通常通过检测细胞外刺激并通过跨膜感应组氨酸激酶传递信号来发挥作用。然后，该激酶启动一系列可逆磷酸化修饰，以调节同源细胞质反应调节剂作为转录因子的活性。在大肠杆菌和其他研究充分的革兰氏阴性模式生物中，有几个 TCS 被牵连到细胞周期动态、细胞包膜完整性或细胞壁发育的调节中。然而，许多α-变形菌缺乏这些调节剂的同源物，因此缺乏对α-变形菌如何协调细胞外事件的理解。在这项工作中，我们在α-变形菌红球菌中鉴定出一种必需的 TCS，CenKR（细胞包膜激酶和调节剂），并表明其活性的调节会导致主要的形态变化。通过遗传和生化方法，我们剖析了 CenK 和 CenR 之间磷酸转移事件的要求，利用这些信息在体内操纵该 TCS 的活性，并鉴定出 CenKR 在 Rb 中直接和间接控制的基因。sphaeroides。将 ChIP-seq 和 RNA-seq 相结合，我们表明 CenKR TCS 直接参与维持细胞包膜，调节 Tol-Pal 外膜分裂复合物亚基的表达，并间接调节肽聚糖生物合成基因的表达。CenKR 是第一个被报道在革兰氏阴性细菌中直接控制 Tol-Pal 机械基因表达的 TCS，我们预测该 TCS 的同源物在其他密切相关的生物体中具有类似的功能。我们提出，Rb。sphaeroides 受 CenKR 直接调节的未知功能基因在该菌和其他α-变形菌中细胞壁生物合成、组装和/或重塑中发挥未知作用。
