Pilus assembly in Gram-positive bacteria

性状（由申请人提供）：皮利或菌毛存在于许多革兰氏阳性病原体的表面，包括白喉棒状杆菌、肠球菌和链球菌。这些纤维状蛋白聚合物在细菌致病的许多方面起着关键作用，如粘附宿主组织、侵入、生物膜形成和调节宿主免疫。这些皮利在细菌细胞壁上的组装通过分选酶依赖性机制发生，这首先在C.白喉，白喉的病原体。C.白喉杆菌表达三种不同的异三聚体菌毛结构，根据形成菌毛杆的主要亚基命名为SpaA-、SpaD-和SpaH-型皮利。SPA型的特点是 由spaA-srtA- spaB-spaC基因座编码的菌毛。该原型菌毛由形成菌毛轴的SpaA、位于菌毛顶端的SpaC和位于菌毛基部的SpaB组成。重要的是，SpaA皮利介导棒状杆菌对咽上皮细胞的粘附，咽上皮细胞是棒状杆菌的主要部位。白喉感染我们发现，菌毛亚基的聚合需要一个同源的转肽酶命名为菌毛特异性分选酶（SrtA）通过赖氨酸介导的转肽。所得聚合物通过管家分选酶SrtF连接到细胞壁。近年来，革兰氏阳性菌皮利的生物学研究取得了很大进展，确立了皮利在致病机制中的重要作用，以及其对开发针对链球菌病原体的多价疫苗的适用性。然而，在我们对以下方面的理解中存在重大差距：（a）如何控制皮利的表面表达，（B）菌毛蛋白如何以特定顺序组装，以及（c）是什么决定了菌毛聚合和细胞壁锚定及其在细菌发病机制中的衍生物。在目前的建议中，我们通过使用多学科方法来寻求这些基本问题的答案。具体而言，我们试图描绘的机制，控制表面组装的皮利的特点SrtA-SrtF与其基板和SarA，一种新的膜结合蛋白所需的SrtF介导的细胞壁锚定，并通过发现特定的反式作用因子，调节srtF的表达。其次，我们的目标是进一步阐明菌毛生物发生的基本机制，通过破译的分子决定因素和潜在的相互作用，精心安排的菌毛有序组装，并通过揭示潜在的伴侣和辅助因子参与菌毛生物发生。最后，我们的目标是建立两个范式原则，即棒状杆菌皮利的表面组装显着促进细菌发病机制，以及结合皮利和毒素抗原的疫苗增强保护功效。所产生的结果肯定不仅会提供一个清晰的了解菌毛大会和菌毛介导的致病性革兰氏阳性菌的基本机制，而且知识，改善健康。

DESCRIPTION (provided by applicant): Pili or fimbriae are present on the surface of many gram-positive pathogens, including Corynebacterium diphtheriae, enterococci and streptococci. These fiber-like protein polymers play a pivotal role in many facets of bacterial pathogenesis such as adherence to host tissues, invasion, biofilm formation and modulation of host immunity. The assembly of these pili on the bacterial cell wall occurs by a sortase-dependent mechanism, which was first demonstrated in C. diphtheriae, the causative agent of diphtheria. C. diphtheriae expresses three distinct heterotrimeric pilus structures, named SpaA-, SpaD-, and SpaH-type pili according to the major subunit that forms the pilus shaft. Well characterized is the SpaA-type pilus that is encoded by the spaA-srtA- spaB-spaC locus. This archetype pilus is composed of SpaA forming the shaft, SpaC located at the tip and SpaB at the pilus base. Importantly, the SpaA pili mediate corynebacterial adherence to pharyngeal epithelial cells, the major site of C. diphtheriae infection. We showed that polymerization of the pilus subunits requires a cognate transpeptidase enzyme named pilin-specific sortase (SrtA) via lysine-mediated transpeptidation. The resulting polymer is joined to the cell wall by the housekeeping sortase SrtF. Recently, a great deal of progress has been made on the biology of Gram-positive pili, establishing the prominent roles of pili in pathogenesis as well as their suitability for developing multivalent vaccines against streptococcal pathogens. However, there are major gaps in our understanding of (a) how surface expression of pili is controlled, (b) how pilins are assembled in a specific order, and (c) what governs the decision between pilus polymerization and cell wall anchoring and its ramification in bacterial pathogenesis. In the current proposal, we seek answers to these fundamental questions by using a multidisciplinary approach. Specifically, we seek to delineate the mechanism that controls surface assembly of pili by characterizing SrtA-SrtF interactions with their substrates and SarA, a novel membrane bound protein required for SrtF-mediated cell wall anchoring, and by discovering specific trans-acting factors that regulate srtF expression. Secondly, we aim to further elucidate the fundamental mechanisms of pilus biogenesis by deciphering the molecular determinants and the underlying interactions that orchestrate the ordered assembly of the pilus and by uncovering potential chaperones and cofactors involved in pilus biogenesis. Finally, we aim to establish two paradigmatic principles that surface assembly of corynebacterial pili significantly contributes to bacterial pathogenesis and that a vaccine combining pilus and toxin antigens enhances protective efficacy. The results generated will certainly provide not only a clear understanding of the basic mechanisms of pilus assembly and pilus-mediated pathogenesis in Gram-positive bacteria but also knowledge that improves health.