Integrated control of Caulobacter cell physiology by visible light and stress

描述(申请人提供)：细胞的环境对其生理、发育和进化有深远的影响。新月形杆菌是一个研究细胞周期控制和发育的细菌模型，它为研究环境感知和适应的分子基础提供了一个很好的实验系统。这项提议的目标是定义光和压力信号如何被细菌整合以调节细胞包膜和细胞黏附的分子和细胞机制。LOV-组氨酸激酶(LOV-HKS)是一类新发现的蓝光感受器，在一系列原核生物中都是保守的。虽然LOV-HKS的调控作用还不是很清楚，但这些信号蛋白最近已经被证明控制流产布鲁氏菌的毒力和Caulbacter对光的反应中的细胞黏附。在这些发现之前，卡氏杆菌和布鲁氏菌都不是已知的，也不被推定对可见光有反应。事实上，编码LOV-HKS的大多数物种都是没有预测的光生物学的化学营养体。我们已经在Caulbacter中发现了一个调控网络，其中LOV-HK、LOVK和受体蛋白LOVR与CT形成一个调控反馈环，CT是一种包膜应激西格玛因子，对细胞在渗透和氧化应激下的生存至关重要。这项提议中详细描述的实验将检验这一假设，即LovK/LovR系统是一个新的信号网络的一部分，在该网络中，经典的双组分信号和C依赖的转录控制交叉，以调节细胞被膜的组成，以响应环境中的多种物理和化学信号。我们将使用多种方法来定义化学环境和光环境之间的相互作用如何影响细胞应力适应、细胞膜成分和细胞黏附。具体地说，我们将回答以下问题：(I)LOVK/LOVR双组分系统如何调节细胞被膜成分和细胞对光的黏附，(Ii)LOVK/LOVR光敏网络和CT应激反应网络之间的调控相互作用，以及(Iii)CT、CU和PHYR这三个似乎受CT调控的西格玛因子的转录靶点是什么？在时间允许的情况下，我们使用遗传和生化筛选来确定LovK/LovR黏附途径中的其他调节因子。我们的实验将加深我们对LOV-HKS光调控的理解，LOV-HKS是影响细菌发病机制的一个重要的原核生物学新领域。更广泛地说，这些研究将提供有关细菌在波动环境中感知和整合多种环境刺激的机制的重要数据，这对细菌细胞的生存至关重要。

DESCRIPTION (provided by applicant): The environment of a cell has a profound influence on its physiology, development, and evolution. Caulobacter crescentus, a bacterial model for the study of cell cycle control and development, provides an excellent experimental system to investigate the molecular basis of environmental perception and adaptation. The goal of this proposal is to define the molecular and cellular mechanisms of how light and stress signals are integrated by a bacterium to regulate the cell envelope and cell adhesion. LOV-histidine kinases (LOV-HKs), a newly-discovered class of blue-light photosensors, are conserved across a range of prokaryotes. Although the regulatory roles of LOV- HKs are not well understood, these signaling proteins have recently been shown to control virulence in Brucella abortus and cell adhesion in Caulobacter in response to light. Prior to these discoveries, neither Caulobacter nor Brucella were known or presumed to respond to visible light. Indeed, the majority of species encoding LOV-HKs are chemotrophs with no predicted photobiology. We have uncovered a regulatory network in Caulobacter in which the LOV-HK, LovK, and the receiver protein, LovR, form a form a regulatory feedback loop with CT, an envelope stress sigma factor that is critical for cell survival under osmotic and oxidative stress. The experiments detailed in this proposal will test the hypothesis that LovK/LovR system is part of a novel signaling network in which classical two- component signaling and C-dependent control of transcription intersect to regulate the composition of the cell envelope in response to multiple physical and chemical cues in the environment. We will use a combination of methods to define how interaction between the chemical and light environments affect cellular stress adaptation, cell envelope composition, and cell adhesion. Specifically, we will answer the following questions: (i) How does the LovK/LovR two-component system regulate cell envelope composition and cell adhesion in response to light, (ii) What are the regulatory interactions between the LovK/LovR photosensory network and the CT stress-response network, and (iii) What are the transcriptional targets of CT, CU, and PhyR, three sigma factors that appear to be regulated by CT? Time permitting, we use genetic and biochemical screens to identify additional regulators in the LovK/LovR adhesion pathway. Our experiments will advance our understanding of photoregulation by LOV-HKs, an important new area of prokaryotic biology that impacts bacterial pathogenesis. More generally, these studies will provide important data on mechanisms bacteria use to sense and integrate multiple environmental stimuli in a fluctuating environment, which is critical for bacterial cell survival.