Heterogeneous cell wall homeostasis as a survival strategy in stationary phase: balancing cell growth, antibiotic production, and envelope stress response in Bacillus subtilis

细胞包膜是一种重要而又高度动态的结构，它保护细菌细胞免受环境的侵害。在整个生长周期中保持其完整性——即使面对抗生素的威胁——对生存是绝对必要的。虽然近年来对平衡生长过程中包膜稳态的理解取得了重大进展，但对平稳期的调控和生理调节知之甚少。当面临不利的生长条件时，革兰氏阳性模式生物枯草芽孢杆菌开始一个复杂的分化程序，导致不同亚群的形成，例如变得有能力，形成生物膜或分化为休眠的内生孢子。这种多样化归因于下注对冲策略，其目的是通过为多种不同的命运“做好准备”，使群体的生存最大化。但到目前为止，对其他固定期生存策略的生理相关性和相关性知之甚少，包括以细胞壁生物合成为目标的抗菌肽（amp）的产生、由此产生的细胞包膜应激反应（CESR）以及剩余营养细胞中的细胞生长。这些将是本建议的重点，它建立在以下基础之上：(i)单个细胞的生长速度是广泛异质性的，导致营养细胞、分化细胞和裂解细胞的群体广泛混合。（ii） AMP的生产也是一种异质性和瞬态策略。（iii）由于amp靶向细胞壁生物合成，因此其抑制作用与活跃生长的细胞相耦合，因此也应该是异质的。这在（iv）观察到的损伤感知Lia系统的双峰诱导中表达出来，该系统响应细胞壁生物合成的脂质II周期的扰动。基于此，我们提出，静止期的Lia反应代表了一个“生理三重and门”，需要AMP的产生、细胞生长和不足的内在保护。为了解决这一假设，并最终解释AMP产生、固定相生长和CESR诱导的异质性之间的相关性，我们追求四个主要目标。(i)我们的目标是提取细胞生长、AMP产生和CESR之间的相关性和表观遗传的定量单细胞图。（ii）我们希望建立一个定量和动态的脂质ii周期数学模型，以模拟AMP产生与CESR诱导之间的相关性。（iii）结合上述两个目标，我们的目标是确定负责决定细胞命运决定的随机/调节过程，包括产孢、AMP的产生和CESR的诱导。（iv）最终，我们希望揭示细胞包膜稳态和lia介导的CESR中表型异质性与固定期生存的生理相关性。

The cell envelope is an essential yet highly dynamic structure that protects the bacterial cell from its environment. Maintaining its integrity throughout the growth cycle - even in the face of antibiotic threat - is absolutely essential for survival. While significant progress has been made in recent years on understanding envelope homeostasis during balanced growth, little is known on the regulatory and physiological adjustments in stationary phase. When faced with unfavorable growth conditions, the Gram-positive model organism Bacillus subtilis embarks on a complex differentiation program, which results in the formation of different subpopulations that e.g. become competent, form biofilms or differentiate into dormant endospores. This diversification is attributed to bet-hedging strategies that aim at maximizing the survival of the population by 'being prepared' for a number of different fates. But so far little is known about the physiological relevance and correlation of other stationary phase survival strategies, including the production of antimicrobial peptides (AMPs) that target cell wall biosynthesis, the resulting cell envelope stress responses (CESR), and the cell growth in the remaining vegetative cells. These will be the focus of this proposal, which rests on the following cornerstones: (i) The growth rate of individual cells is broadly heterogeneous resulting in widely mixed populations of vegetative, differentiating and lysing cells. (ii) AMP production is also a heterogeneous and transient strategy. (iii) Since AMPs target cell wall biosynthesis, their inhibitory action is coupled to actively growing cells and should therefore also be heterogeneous. This is expressed in (iv) the bimodal induction observed for the damage-sensing Lia system, which responds to perturbations of the Lipid II cycle of cell wall biosynthesis. Based on this, we propose that the Lia response in stationary phase represents a 'physiological triple AND-gate' that requires AMP production, cell growth, and insufficient intrinsic protection. To address this hypothesis and ultimately explain the correlation between the heterogeneity in AMP production, stationary phase growth and induction of CESR, we pursue four major goals. (i) We aim at extracting a quantitative, single-cell based picture of the correlation and epigenetic inheritance between cell growth, AMP production, and CESR. (ii) We want to develop a quantitative and dynamic mathematical model of the Lipid II cycle, in order to simulate the correlation between AMP production and the resulting induction of the CESR. (iii) In combining the above two goals, we aim at Identifying the stochastic/regulatory processes responsible for determining the cell fate decision regarding sporulation, AMP production and induction of the CESR. (iv) Ultimately, we would like to unravel the physiological relevance of phenotypic heterogeneity in cell envelope homeostasis and Lia-mediated CESR for stationary phase survival.