In animal pathogens, assembly of the type III secretion system injectisome requires the presence of so-called pilotins, small lipoproteins that assist the formation of the secretin ring in the outer membrane. Using a combination of functional assays, interaction studies, proteomics, and live-cell microscopy, we determined the contribution of the pilotin to the assembly, function, and substrate selectivity of the T3SS and identified potential new downstream roles of pilotin proteins. In absence of its pilotin SctG, Yersinia enterocolitica forms few, largely polar injectisome sorting platforms and needles. Accordingly, most export apparatus subcomplexes are mobile in these strains, suggesting the absence of fully assembled injectisomes. Remarkably, while absence of the pilotin all but prevents export of early T3SS substrates, such as the needle subunits, it has little effect on secretion of late T3SS substrates, including the virulence effectors. We found that although pilotins interact with other injectisome components such as the secretin in the outer membrane, they mostly localize in transient mobile clusters in the bacterial membrane. Together, these findings provide a new view on the role of pilotins in the assembly and function of type III secretion injectisomes.

中文翻译：

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Pilotins 是参与细菌 III 型分泌系统的组装和底物特异性的移动 T3SS 组件

在动物病原体中，III 型分泌系统注射体的组装需要所谓的 Pilotins 的存在，这是一种小脂蛋白，有助于在外膜中形成促胰液素环。通过结合功能测定、相互作用研究、蛋白质组学和活细胞显微镜，我们确定了 Pilotin 对 T3SS 的组装、功能和底物选择性的贡献，并确定了 Pilotin 蛋白潜在的新下游作用。在缺乏 Pilotin SctG 的情况下，小肠结肠炎耶尔森氏菌形成很少的、主要是极性的注射体分选平台和针。因此，大多数输出​​装置子复合体在这些菌株中是可移动的，表明不存在完全组装的注射体。值得注意的是，虽然 Pilotin 的缺失几乎阻止了早期 T3SS 底物（例如针亚基）的输出，但它对晚期 T3SS 底物（包括毒力效应子）的分泌几乎没有影响。我们发现，尽管 Pilotins 与其他注射体成分（例如外膜中的促胰液素）相互作用，但它们大多定位于细菌膜中的瞬时移动簇中。总之，这些发现为 Pilotins 在 III 型分泌注射体的组装和功能中的作用提供了新的观点。