Staphylococcus aureus host cell invasion and virulence in sepsis is facilitated by the multiple repeats within FnBPA.
Staphylococcus aureus host cell invasion and virulence in sepsis is facilitated by the multiple repeats within FnBPA.
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DOI：
10.1371/journal.ppat.1000964
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发表时间：
2010-06-24
期刊：
影响因子：
6.7
通讯作者：
Massey RC
中科院分区：
文献类型：
--
作者：
Edwards AM;Potts JR;Josefsson E;Massey RC
金黄色葡萄球菌进入血流可导致转移性脓肿形成和感染性心内膜炎。对这两种条件的发展至关重要的是S。金黄色葡萄球菌与内皮细胞。体内和体外研究表明，葡萄球菌侵袭素FnBPA通过一个涉及纤连蛋白（Fn）桥接至α5β1整联蛋白的过程触发内皮细胞的细菌侵袭。FnBPA的Fn结合区通常含有11个对Fn具有不同亲和力的非相同重复序列（FnBR），其促进多个Fn分子的结合并可促进整联蛋白聚集。因此，我们假设多个重复序列是触发S.金黄色。为了测试这一点，我们构建了含有各种FnBR组合的fnbA变体。体外试验表明，内皮细胞的侵袭可以促进由一个单一的高亲和力，但不是低亲和力的FnBR。使用尼生素诱导系统控制FnBPA的表面表达的研究表明，编码较少FnBR的变体需要更高水平的表面表达来介导侵袭。高表达水平的FnBPA携带单个低亲和力FnBR结合Fn，但不侵入，表明FnBPA对Fn的亲和力对于触发内化是至关重要的。此外，多个FnBR增加了内化的速度，FnBPA的表达水平更高，而不改变摄取机制。使用小鼠脓毒症模型证明了这些发现与发病机制的相关性，该模型表明毒力需要多种FnBR。总之，FnBPA内的多个FnBR促进有效的Fn粘附，触发快速细菌摄取，并且是致病所需的。 金黄色葡萄球菌是菌血症和败血症的常见原因。S.金黄色葡萄球菌可导致心脏瓣膜的定殖和/或扩散到周围组织中以及继发性（转移性）感染的建立。内皮细胞的摄取是由纤连蛋白结合蛋白A（FnBPA）通过纤连蛋白（Fn）桥与称为整合素的宿主细胞受体相互作用触发的。FnBPA含有11个介导与Fn结合的非相同重复序列（FnBR）。先前的工作表明，FnBR区域的部分缺失不会显著影响与Fn的结合或体外宿主细胞的侵袭。因此，不清楚为什么FnBPA包含如此多的重复序列。这项工作表明，重复的数量影响触发入侵所需的表面暴露的FnBPA的量，以及影响吸收的速度。因此，FnBPA内多个重复序列的存在即使在低表面表达水平下也促进粘附和侵袭。使用败血症小鼠模型的实验表明，这些差异对毒力有显著影响;与具有完整重复序列的FnBPA相比，表达FnBPA变体的金黄色葡萄球菌的无FnBR或具有很少FnBR的FnBPA变体的毒性显著更低。
Entry of Staphylococcus aureus into the bloodstream can lead to metastatic abscess formation and infective endocarditis. Crucial to the development of both these conditions is the interaction of S. aureus with endothelial cells. In vivo and in vitro studies have shown that the staphylococcal invasin FnBPA triggers bacterial invasion of endothelial cells via a process that involves fibronectin (Fn) bridging to α5β1 integrins. The Fn-binding region of FnBPA usually contains 11 non-identical repeats (FnBRs) with differing affinities for Fn, which facilitate the binding of multiple Fn molecules and may promote integrin clustering. We thus hypothesized that multiple repeats are necessary to trigger the invasion of endothelial cells by S. aureus. To test this we constructed variants of fnbA containing various combinations of FnBRs. In vitro assays revealed that endothelial cell invasion can be facilitated by a single high-affinity, but not low-affinity FnBR. Studies using a nisin-inducible system that controlled surface expression of FnBPA revealed that variants encoding fewer FnBRs required higher levels of surface expression to mediate invasion. High expression levels of FnBPA bearing a single low affinity FnBR bound Fn but did not invade, suggesting that FnBPA affinity for Fn is crucial for triggering internalization. In addition, multiple FnBRs increased the speed of internalization, as did higher expression levels of FnBPA, without altering the uptake mechanism. The relevance of these findings to pathogenesis was demonstrated using a murine sepsis model, which showed that multiple FnBRs were required for virulence. In conclusion, multiple FnBRs within FnBPA facilitate efficient Fn adhesion, trigger rapid bacterial uptake and are required for pathogenesis. Staphylococcus aureus is a frequent cause of bacteremia and sepsis. Adhesion to and invasion of endothelial cells lining blood vessels by S. aureus can lead to colonization of the heart valves and/or dissemination into surrounding tissues and the establishment of secondary (metastatic) infections. Uptake by endothelial cells is triggered by the interaction of fibronectin-binding protein A (FnBPA) with host cell receptors called integrins via a fibronectin (Fn) bridge. FnBPA contains 11 non-identical repeats (FnBRs) that mediate binding to Fn. Previous work has shown that partial deletions in the FnBR region do not significantly affect binding to Fn or invasion of host cells in vitro. It was therefore unclear why FnBPA contains so many repeats. This work demonstrates that the number of repeats affects the amount of surface-exposed FnBPA required to trigger invasion, as well as affecting the speed of uptake. As such, the presence of multiple repeats within FnBPA facilitates adhesion and invasion even at low surface expression levels. Experiments using a murine model of sepsis demonstrate that these differences have a significant impact on virulence; S. aureus expressing FnBPA variants with no or few FnBRs are significantly less virulent compared to FnBPA with the full complement of repeats.
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