Regulation of potassium uptake in Caulobacter crescentus [ChIP-seq]. Regulation of potassium uptake in Caulobacter crescentus [ChIP-seq]

Potassium (K+) is an essential physiological element determining membrane potential, intracellular pH, osmotic/turgor pressure, and protein synthesis in cells. Here we describe the regulation of potassium uptake systems in the oligotrophic alpha-proteobacterium Caulobacter crescentus known as a model for asymmetric cell division. We show that C. crescentus can grow in concentrations from the micromolar to the millimolar range by mainly using two K+ transporters to maintain potassium homeostasis, the low affinity Kup and the high affinity Kdp uptake systems. When K+ is not limiting, we found that the kup gene is essential while kdp inactivation does not impact the growth. In contrast, kdp becomes critical but not essential and kup dispensable for growth in K+-limited environments. However, in the absence of kdp, mutations in kup were selected to improve growth in K+-depleted conditions, likely by increasing the affinity of Kup for K+. In addition, mutations in the KdpDE two-component system, which regulates kdpABCDE expression, suggest that the inner membrane sensor regulatory component KdpD mainly works as a phosphatase to limit the growth when cells reach late exponential phase. Our data therefore suggest that KdpE is phosphorylated by another non-cognate histidine kinase. On top of this, we determined the KdpE-dependent and independent K+ transcriptome. Together, our work illustrates how an oligotrophic bacterium responds to fluctuation in K+ availability. Overall design: Examination of KdpE whole genome binding/occupancy (ChIP-Seq) in WT, ∆kdpD and ∆kdpE of Caulobacter crescentus grown in synthetic minimal media M2G-K with 0.025 mM K+ or 0.5 mM K+

钾离子（K+）是维持细胞膜电位、细胞内pH值、渗透/膨压以及细胞内蛋白质合成的必需生理元素。本研究聚焦于作为不对称细胞分裂模型生物的寡营养α-变形菌——新月柄杆菌（Caulobacter crescentus），阐述其钾摄取系统的调控机制。研究发现，新月柄杆菌可通过两种主要的K+转运系统维持钾稳态，在微摩尔至毫摩尔浓度范围内生长：低亲和性Kup转运体与高亲和性Kdp摄取系统。当K+供应不匮乏时，kup基因是必需的，而kdp失活不会对菌体生长造成影响；与之相反，在K+受限环境中，kdp成为关键（但非必需）基因，kup则可被省略。然而，当kdp缺失时，会筛选出kup突变体以提升K+耗尽条件下的生长能力，其机制可能是增强了Kup对K+的亲和性。此外，调控kdpABCDE表达的KdpDE双组分系统的相关突变研究显示，内膜传感器调控蛋白KdpD主要作为磷酸酶发挥作用，在菌体进入指数生长后期限制其生长。据此，本研究数据表明KdpE可通过其他非同源组氨酸激酶实现磷酸化。在此基础上，我们测定了依赖KdpE与不依赖KdpE的K+转录组。综上，本研究阐明了寡营养细菌如何响应K+可用性的波动。整体实验设计：在添加0.025 mM K+或0.5 mM K+的合成最小培养基M2G-K中培养新月柄杆菌野生型（WT）、∆kdpD与∆kdpE菌株，检测KdpE的全基因组结合/占据情况（染色质免疫沉淀测序，ChIP-Seq）。