Unraveling type IV pilus assembly

皮利是在许多不同的细菌中发现的毛发状附属物，使它们能够“粘”在许多不同的表面。在引起人类疾病的细菌中，它们通常促进对宿主细胞的附着和定植。没有哪种皮利比IV型皮利（Tfp）分布得更广了。IV型Tfp存在于数百种细菌中，包括许多导致危及生命的疾病的细菌，如脑膜炎奈瑟菌（导致脑膜炎和败血症）和霍乱弧菌（导致霍乱）。研究菌毛生物学至关重要，因为更好地了解细菌组装这些菌毛的方式有望为新药的设计提供线索。干扰菌毛组装将阻止定植，从而“解除”病原菌。这是一个有希望的替代大量使用抗生素，导致抗生素耐药性的无情传播，这是一个主要的公共卫生问题在世界范围内。现在有一些非常好的表征皮利的实验证据表明，这种方法可以工作。然而，我们对Tfp是如何组装的理解对于这种方法来说太有限了，需要改进，这是这里提出的研究的主要目标。Tfp生物合成依赖于一个非常复杂的机器，多达18个高度保守的蛋白质。我们已经提供了间接的证据表明，TFP组装依赖于这些蛋白质的一个相对较小的子集。我们最近发现，这种机制可以在细菌模型中自组装，这为“乐高”方法铺平了道路，在这种方法中，最小的Tfp组装机制将在大肠杆菌中重建。因此，本研究计划的两个目的是：1.重组于E. coli中构建了一个最小系统，该系统能够通过N.脑膜炎双球菌2.对Tfp组装机制进行深入的功能表征，以更好地了解其作用模式。这有望对Tfp生物学产生重大影响，并成为未来设计能够干扰Tfp组装的药物的关键，预计将具有非常广泛的作用范围。

Pili are hair-like appendages found in many different bacteria that allow them to "stick" to many different surfaces. In bacteria causing disease in humans, they often promote attachment to host cells and colonization. No pili are more widespread than type IV pili (Tfp) that are found in hundreds of bacterial species, including many that cause life-threatening diseases such as Neisseria meningitidis (responsible for meningitis and septicaemia) and Vibrio cholerae (responsible for cholera).Studying pilus biology is crucial because a better understanding of the way in which bacteria assemble these filaments is expected to provide leads for the design of new drugs. Interfering with pilus assembly would prevent colonization and thus "disarm" pathogenic bacteria. This is a promising alternative to the massive use of antibiotics that has resulted in an unrelenting spread of antibiotic resistance, which is a major public health concern worldwide. There is now experimental evidence for some very well characterized pili that this approach can work. However, our understanding of how Tfp are assembled is far too limited for such an approach and needs to be improved, which is the main objective of the research proposed here.Tfp biogenesis relies on a very complex machinery of up to 18 highly conserved proteins. We have provided indirect evidence that Tfp assembly relies on a relatively small subset of these proteins. Our recent finding that this machinery can self-assemble in the bacterial model par excellence Escherichia coli paves the way for a "Lego" approach in which a minimal Tfp assembly machinery would be reconstituted in E. coli through sequential addition of the required meningococcal components.The two aims of this research project are therefore to:1. reconstitute in E. coli a minimal system that is capable of assembling Tfp through modular co-expression of N. meningitidis pil genes,2. perform and in depth functional characterization of the Tfp assembly machinery in order to better understand its mode of action.This is expected to have a major impact on Tfp biology and be key for the future design of drugs capable of interfering with Tfp assembly, with are expected to have an exceptionally wide spectrum of action.