Cell envelope integrity in mycobacteria: interplay of lipoglycans, peptidoglycan, and capsular polysaccharides

项目总结 脂肪甘露聚糖(LM)和脂阿拉伯甘露聚糖(LAM)是分枝杆菌脂多糖的重要组成部分。 胸膜结核以确定感染。这些脂多糖是在质膜中合成的， 被贩卖到细胞表面。Lm和LAM在分枝杆菌中广泛保守，因此也必须 在致病分枝杆菌和非致病分枝杆菌的生理学中都起着重要作用。然而， 它们的生物合成是如何与其他细胞膜生物合成相协调的，以及它们是如何 有助于细胞包膜的完整性。长期目标是确定脂多糖在我体内的作用。 在创建和维护细胞包膜的过程中识别共生细菌生理学和识别脆弱性 结构。作为下一步，这项建议的目标是获得对脂肪甘油的作用的基本见解- 污垢分枝杆菌和结核分枝杆菌细胞膜中的罐头。中心假设是 LM/LAM生物合成与肽聚糖和壳层a-葡聚糖的生物合成协同作用，正向结合。 致敬多糖的动态平衡，确保活跃生长过程中细胞表面结构的可塑性和完整性。 其基本原理是，在协调多糖生物合成的调控过程中找到漏洞将 为细胞包膜靶向抗菌剂的开发奠定基础。由出版的和初步的指导 来自申请者实验室的数据，这一假设将通过追求两个目标来检验：1)确定 LM/LAM生物合成在抑制肽聚糖降解中的作用；以及2)决定被囊层的作用 来维持细胞包膜的完整性。在第一个目的中，假设LAM缺陷突变体的细胞形状 将测试由于细胞壁变弱而产生的变形。LAM抑制作用的潜在分子机制 将检查肽多聚糖水解酶。在第二个目标中，假设细胞形态缺陷 LAM缺陷突变体可以通过激活SenX3-RegX3双组分系统并增加- 将对a-葡聚糖的生产进行检查。这项拟议的研究具有创新性，因为它揭示了一种新的 分枝杆菌脂多糖在维持糖稳态中的作用，因为它结合了协同作用 申请人及其合作者在细胞正向遗传学和生物正交代谢标记方面的专业知识 WALL生物合成，在概念和执行上都与现状有实质性的背离。建议数 研究意义重大，因为这里获得的数据将为开发能够 通过针对分枝杆菌细胞膜内稳态的调节机制来杀死结核分枝杆菌。这样的一个 药物还可能使细胞膜对其他抗菌剂更具渗透性。

PROJECT SUMMARY Lipomannan (LM) and lipoarabinomannan (LAM) are mycobacterial lipoglycans that are critical for Mycobacte- rium tuberculosis to establish infection. These lipoglycans are synthesized in the plasma membrane and trafficked to the cell surface. LM and LAM are widely conserved among mycobacteria, and therefore must also play a fundamental role in the physiology of both pathogenic and nonpathogenic mycobacterial species. However, it remains obscure how their biosynthesis is coordinated with other cell envelope biosyntheses and how they contribute to the integrity of the cell envelope. The long-term goal is to determine the role of lipoglycans in my- cobacterial physiology and identify vulnerabilities in the process of creating and maintaining the cell envelope structure. As the next step, the objective of this proposal is to gain fundamental insights into the role of lipogly- cans in cell envelope integrity in Mycobacterium smegmatis and M. tuberculosis. The central hypothesis is that LM/LAM biosynthesis, in coordination with peptidoglycan and capsular a-glucan biosyntheses, positively con- tributes to glycan homeostasis, ensuring the plasticity and integrity of cell surface structure during active growth. The rationale is that finding vulnerabilities in the regulatory process that coordinates the glycan biosyntheses will lay the foundation for cell envelope-targeting antimicrobial development. Guided by published and preliminary data from the applicant’s laboratory, this hypothesis will be tested by pursuing two aims: 1) Identify the role of LM/LAM biosynthesis in suppressing peptidoglycan degradation; and 2) Determine the role of the capsular layer in maintaining cell envelope integrity. In the first aim, the hypothesis that the cell shape of a LAM-deficient mutant deforms due to the weakened cell wall will be tested. The potential molecular mechanism of LAM suppressing peptidoglycan hydrolases will be examined. In the second aim, the hypothesis that the morphological defect of the LAM-deficient mutant can be overcome by activating the SenX3-RegX3 two-component system and increas- ing the production of a-glucan will be examined. The proposed research is innovative because it reveals a new role of mycobacterial lipoglycans in maintaining glycan homeostasis, and because it combines the synergistic expertise of the applicant and his collaborators in forward genetics and bioorthogonal metabolic labeling of cell wall biosynthesis, a substantive departure from the status quo in both concept and execution. The proposed research is significant because the data obtained here will form the foundation for developing drugs that could kill M. tuberculosis by targeting a regulatory mechanism of mycobacterial cell envelope homeostasis. Such a drug may also make the cell envelope more permeable to other antimicrobials.