Nothing wasted: Peptidoglycan recycling in mycobacteria

结核分枝杆菌是一个活生生的悖论。在细菌中，它拥有最复杂和富含碳的细胞壁之一，但它却在人类肺部这种营养贫乏的环境中茁壮成长。考虑到分枝杆菌细胞壁的成分是先天免疫系统的有效触发因素，这一点更令人印象深刻。因此，为了支持宿主体内的稳定生长，这些细菌必须非常高效地利用可用的营养物质，并能够躲避免疫系统的攻击。分枝杆菌细胞壁最里面的成分是一种叫做肽聚糖（PG）的复杂大分子。虽然PG的教科书插图表明它是一种静态的袋状结构，但实际上它是一种经过大量修饰的动态分子，可以防止细胞爆炸，同时赋予细胞形状。在细菌的整个生命周期中，PG必须不断地仔细地被切割和重塑，以支持生长和分裂。研究人员长期以来一直在研究PG，主要是因为它的本质。事实上，第一种临床抗生素青霉素就是专门针对PG的生物合成的。尽管多年来人们对这种生物合成已经有了很多了解，但对于PG合成后发生了什么却知之甚少。对于结核分枝杆菌的PG来说更是如此，结核分枝杆菌是一种全球性病原体，也是传染病中人类的第二大杀手。例如，分枝杆菌PG片段一旦被裂解酶从细胞壁上切割下来，其命运就不得而知了。这是一个重要的问题，因为如上所述，这些PG片段是免疫系统的有效刺激物，需要大量的稀缺资源来制造。有趣的是，这些相同的PG片段能够触发细菌从其生活方式的休眠阶段复苏。潜伏的结核分枝杆菌在产生活动性感染之前，能够在宿主体内潜伏数年而不被发现。这严重复杂化了对疾病和相关暴发的控制，因为即使它们被发现，休眠结核分枝杆菌也具有异常的抗生素耐药性。这些观点清楚地表明，了解结核分枝杆菌细胞壁碎片的命运是必要的。在一些细菌中，已经描述了复杂的PG回收系统，其工作原理是将PG片段带回细胞中进行再利用。在迄今为止观察到的所有病例中，这种循环系统对细菌感染宿主的能力很重要。尽管如此，PG的再循环从未在结核分枝杆菌中进行过研究。研究这一途径是很重要的，因为分枝杆菌PG循环系统的元件很可能是新的抗生素靶点。在这个奖学金期间，我将使用新的和令人兴奋的技术来证明分枝杆菌回收它们的PG。我将采用高分辨率的方法来探索被回收的PG片段的精确结构。然后，我将确定PG回收所需的细胞机制，并描述该途径中涉及的关键蛋白质。最后，我将探讨肺结核复苏与PG回收的关系。这项工作将解决结核分枝杆菌悖论，并展示它如何能够平衡其营养和生长需求，其营养贫乏的环境和需要隐藏自己从我们的免疫系统。

Mycobacterium tuberculosis is a living paradox. Amongst bacteria, it possesses one of the single most complex and carbon-rich cell walls and yet it thrives in the decidedly nutrient poor environment of the human lung. This is made even more impressive considering the fact that components of the mycobacterial cell wall are potent triggers of the innate immune system. Therefore, to support stable growth in their host, these bacteria must be exquisitely efficient with their available nutrients and be able to hide themselves from the immune system. The innermost component of the mycobacterial cell wall is a complex macromolecule called peptidoglycan (PG). While textbook illustrations of PG suggest that it is a static bag-like structure, in reality it is a heavily modified and dynamic molecule that prevents the cell from exploding while simultaneously giving it shape. Throughout the life cycle of the bacterium, PG must constantly and carefully be cut and remodeled to support growth and division. Researchers have long studied PG, chiefly because of its essential nature. Indeed, the first clinical antibiotic, penicillin, specifically targets the biosynthesis of PG. While much of this biosynthesis has been understood for many years, relatively little is known about what happens to PG after it is built. This is even truer for the PG of M. tuberculosis, which is a global pathogen and the second leading killer of people amongst infectious diseases. For example, nothing is known about the fate of mycobacterial PG fragments once lytic enzymes have cut them from the cell wall. This is an important question because as mentioned above, these PG fragments are potent stimulators of the immune system and command a great deal of scarce resources to make. Intriguingly, these same PG fragments are capable of triggering resuscitation of the bacterium from the dormant phase of its lifestyle. Dormant M. tuberculosis is capable of persisting undetected for years inside of its host before producing an active infection. This severely complicates control of the disease and associated outbreaks, because even if they are detected, dormant M. tuberculosis are exceptionally antibiotic resistant. These points make it clear that an understanding of the fate of M. tuberculosis cell wall fragments is needed. In some bacteria complex PG recycling systems have been described which work by bringing PG fragments back into the cell to be reused. In all cases observed to-date this recycling system is important to the ability of the bacterium to infect its host. Despite this, PG recycling has never been investigated in M. tuberculosis. It is important that this pathway be investigated because it is very likely that the elements of a mycobacterial PG recycling system are novel antibiotic targets. During this Fellowship I will use new and exciting techniques to demonstrate that mycobacteria recycle their PG. I will take a high-resolution approach by exploring the precise structure of the PG fragments that are being recycled. I will then determine the cellular machinery that is required for PG recycling and characterise the key proteins involved in the pathway. Finally, I will examine the relationship between tuberculosis resuscitation and PG recycling. This work will address the M. tuberculosis paradox and show how it is able to balance its nutritional and growth requirements with its nutrient poor environment and the need to hide itself from our immune system.