Triacylglycerol metabolism and Mtb virulence

描述（由申请人提供）：结核分枝杆菌具有在人类宿主内以静止形式存在多年的能力，只有在宿主免疫系统衰退后才会重新激活并产生传染病。静止形式的细菌被认为是在宿主适应性免疫诱导的生长停滞状态。因此，了解在复制和非复制状态之间转换的代谢适应机制将有助于开发针对持续状态下独特代谢途径的抗结核药物。本项目的目的是剖析三酰基甘油（TAG）代谢在结核分枝杆菌持续和再生中的作用。该建议基于结核分枝杆菌实验衍生的代谢模型，该模型的特征是在持续状态建立期间，从生物合成前体合成的关键中心代谢途径到TAG形成的协调碳通量。结核分枝杆菌中的TAG积累被认为是其长期持续和疾病激活期间再生的关键碳和能量燃料。为了验证这一假设，研究人员采用了一种新的方法来设计TAG的代谢途径，从而可以操纵TAG的代谢动力学来评估其在结核分枝杆菌休眠和再生过程中的功能。首先将采用一种工程策略，结合靶向删除一个主要的TAG合酶基因和有条件地高表达一个高活性TAG脂肪酶基因，来产生一个TAG缺陷结核分枝杆菌菌株。然后，在体外休眠模型和兔结核模型中评估TAG的代谢动态以及TAG对结核分枝杆菌存活和再生长的作用。如果成功，该项目的研究结果有望提供新的研究方向，重点关注TAG代谢，由此可以开发出新的药物来解决持久性杆菌并防止疾病重新激活。此外，缺乏tag的结核分枝杆菌可能是结核病疫苗的潜在候选者。

DESCRIPTION (provided by applicant): Mycobacterium tuberculosis has the ability to persist for many years in a quiescent form within a human host, only to reactivate and produce communicable disease after the host immune system falters. The quiescent form of the bacteria is believed to be in a state of growth arrest induced by host adaptive immunity. Thus, understanding the mechanisms of metabolic adaptation during the shift between replicative and nonreplicative state will facilitate the development of anti-tuberculosis drugs that target unique metabolic pathways in the persistent state. The goal of this project is to dissect the roles of triacylglycerol (TAG) metabolism in M. tuberculosis persistence and regrowth. The proposal is based on an experiment-derived metabolic model of M. tuberculosis, which is marked with a coordinated carbon flux from key central metabolic pathways for biosynthetic precursor synthesis towards the formation of TAG during establishment of the persistent state. TAG accumulation in M. tuberculosis is proposed to serve as critical carbon and energy fuel for both its long-term persistence and regrowth during activation of the disease. To test the hypothesis, a novel approach is utilized to engineer the TAG metabolic pathways so that the dynamics of TAG metabolism can be manipulated to evaluate its functions during M. tuberculosis dormancy and regrowth. A TAG-deficient M. tuberculosis strain will first be generated by employing an engineering strategy that combines targeted deletion of a major TAG synthase gene with the conditional high expression of a highly active TAG lipase gene. Then, the dynamics of TAG metabolism and roles of TAG on M. tuberculosis survival and re-growth will be assessed in in vitro dormancy models and in a rabbit tuberculosis model. If successful, findings from this project are expected to provide new lines of research focusing on TAG metabolism from which new drugs could be developed to tackle the persistent bacilli and to prevent the disease from reactivation. Moreover, the TAG-deficient M. tuberculosis could be a potential candidate for a tuberculosis vaccine. PUBLIC HEALTH RELEVANCE: The goal of the program is to study the roles of triacylglycerol (TAG) metabolism in Mycobacterium tuberculosis survival during dormancy and bacterial re-growth during disease reactivation. The experimental procedures combine molecular engineering and biochemical/physiological assays using models of M. tuberculosis dormancy in vitro and in vivo. The program is expected to gain new insights on the metabolic adaptation mechanisms of the tubercle bacilli during transition between different growth states and to offer new strategies to tackle the persistent infection and disease reactivation.