Regulation of phenotypic switching and heterogeneity in Photorhabdus luminescens cell populations

发光杆菌是一种与特定土壤线虫紧密共生的昆虫致病细菌。这种细菌有一个复杂的生活史，由定植在线虫肠道的共生阶段和昆虫幼虫体内线虫外的致病阶段组成。发光菌以两种不同的表型存在，分别称为初级和次级变异体，它们在遗传上是同质的，但在各种形态和生理特征上不同。在长时间的生长过程中，次级细胞自发产生，结果是20%-50%的细胞在28天后经历了过渡。这与线虫和细菌重新结合并从昆虫身体中出现的时间点有关。经过表型转换的细胞在与线虫的重新结合和共生中被禁用。发光假单胞菌细胞群体表型异质性的原因鲜为人知。假设次生细胞更好地适应土壤中的生活，因此种群分裂被认为是一种更好的对冲策略，确保种群在任何条件下都能生存。然而，发光杆菌细胞从未从土壤中分离出来，可能是因为它们以代谢不活跃的持久细胞的形式存在。到目前为止，诱导从次级变异体到初级变异体的反向切换或能够唤醒宿存细胞的信号尚不清楚。从之前的资助期开始，次级细胞会分泌一种信号，诱导原代细胞的转换过程。此外，原代细胞的出现抑制了次级细胞的生长，揭示了两种表型变体之间的简单社会生物学关系。查明这些信号的化学性质以及次级细胞的命运是将在本供资期间处理的中心问题。此外，还将对表型开关调节的分子机制进行研究。LysR型调节器HexA已被确定为开关过程的全局调节器。从上一个资助期出现的情况是，HEXA是一种多功能的调节因子，可以直接或间接地影响转录和转录后水平的表型异质性，可能是通过小的调节RNA。识别这些小核糖核酸以及HexA感受到的信号以激活单个细胞中的转换过程，是将在当前供资期间解决的进一步中心问题。发光菌表型转换的数学模型将有助于理解细菌表型异质性的一般调控机制。

Photorhabdus luminescens is an insect pathogenic bacterium that lives in close symbiosis with specific soil nematodes. The bacteria have a complex life cycle consisting of a symbiotic stage colonizing the intestinal tract of the nematodes, and a pathogenic stage outside of the nematodes within insect larvae. P. luminescens exist in two phenotypically different forms, designated as primary and secondary variants that are genetically homogeneous but differ in various morphological and physiological traits. During prolonged growth, secondary cells arise spontaneously with the result that 20-50% of the cells have undergone the transition after 28 days. This correlates with the time point when nematodes and bacteria have been re-associated and emerge from the insect cadaver. The cells that have undergone phenotypic switching are disabled in re-association and symbiosis with the nematodes. The reason for phenotypic heterogeneity of P. luminescens cell populations is rarely understood. It is assumed that secondary cells are better adapted for a life in the soil, so that splitting of the population is assumed to be a bet-hedging strategy that ensures survival of the population under any condition. However, P. luminescens cells have never been isolated from the soil, probably because they exist as metabolically inactive persister cells. The signals that induce reverse switching from the secondary to the primary variant or that can awake persister cells are unknown so far. It emerged from the previous funding period that secondary cells secrete a signal that induces the switching process in primary cells. Furthermore, primary cells emerged to inhibit growth of secondary cells revealing a simple sociobiological relation between both phenotypic variants. Identification of the chemical nature of these signals as well as the fate of the secondary cells are central questions that will be addressed in the current funding period. Furthermore, the molecular mechanism of phenotypic switching regulation will be investigated. The LysR-type regulator HexA has been identified as global regulator of the switching process. It emerged from the previous funding period that HexA is a versatile regulator that can directly or indirectly influence phenotypic heterogeneity at the transcriptional as well as the post-transcriptional level, probably via small regulatory RNAs. Identification of these small RNAs as well as the signal that is sensed by HexA to activate the switching process in single cells are further central questions that will be addressed in the current funding period. Mathematical modeling of phenotypic switching of P. luminescens will be used to contribute to the understanding of general regulation mechanisms of phenotypic heterogeneity in bacteria.