Université Lyon, Université Lyon 1, INSA Lyon, CNRS UMR 5240 Microbiologie Adaptation et Pathogénie, Villeurbanne, France.
School of Biological and Chemical Sciences, Queen Mary University of London, London, United Kingdom.
mBio. 2022 Jun 28;13(3):e0025322. doi: 10.1128/mbio.00253-22. Epub 2022 May 12.
The phytopathogenic proteobacterium Dickeya dadantii secretes an array of plant cell wall-degrading enzymes and other virulence factors via the type 2 secretion system (T2SS). T2SSs are widespread among important plant, animal, and human bacterial pathogens. This multiprotein complex spans the double membrane cell envelope and secretes fully folded proteins through a large outer membrane pore formed by 15 subunits of the secretin GspD. Secretins are also found in the type 3 secretion system and the type 4 pili. Usually, specialized lipoproteins termed pilotins assist the targeting and assembly of secretins into the outer membrane. Here, we show that in , the pilotin acts in concert with the scaffolding protein GspB. Deletion of profoundly impacts secretin assembly, pectinase secretion, and virulence. Structural studies reveal that GspB possesses a conserved periplasmic homology region domain that interacts directly with the N-terminal secretin domain. Site-specific photo-cross-linking unravels molecular details of the GspB-GspD complex . We show that GspB facilitates outer membrane targeting and assembly of the secretin pores and anchors them to the inner membrane while the C-terminal extension of GspB provides a scaffold for the secretin channel in the peptidoglycan cell wall. Phylogenetic analysis shows that in other bacteria, GspB homologs vary in length and domain composition and act in concert with either a cognate ATPase GspA or the pilotin GspS. Gram-negative bacteria have two cell membranes sandwiching a peptidoglycan net that together form a robust protective cell envelope. To translocate effector proteins across this multilayer envelope, bacteria have evolved several specialized secretion systems. In the type 2 secretion system and some other bacterial machineries, secretins form large multimeric pores that allow transport of effector proteins or filaments across the outer membrane. The secretins are essential for nutrient acquisition and pathogenicity and constitute a target for development of new antibacterials. Targeting of secretin subunits into the outer membrane is often facilitated by a special class of lipoproteins called pilotins. Here, we show that in and some other bacteria, the scaffolding protein GspB acts in concert with pilotin, facilitating the assembly of the secretin pore and its anchoring to both the inner membrane and the bacterial cell wall. GspB homologs of varied domain composition are present in many other T2SSs.
植物病原生质体迪基氏菌通过 2 型分泌系统（T2SS）分泌一系列植物细胞壁降解酶和其他毒力因子。T2SS 在重要的植物、动物和人类细菌病原体中广泛存在。这个多蛋白复合物跨越双层细胞膜包膜，并通过由 15 个分泌蛋白 GspD 亚基组成的大外膜孔分泌完全折叠的蛋白质。分泌蛋白也存在于 3 型分泌系统和 4 型菌毛中。通常，专门的脂蛋白被称为信号肽，可协助将分泌蛋白靶向和组装到外膜中。在这里，我们表明在 中，信号肽与支架蛋白 GspB 协同作用。 的缺失严重影响分泌蛋白的组装、果胶酶的分泌和毒力。结构研究表明，GspB 具有保守的周质同源区结构域，该结构域可直接与 N 端分泌蛋白结构域相互作用。定点光交联揭示了 GspB-GspD 复合物的分子细节。我们表明 GspB 有助于外膜靶向和分泌蛋白孔的组装，并将其锚定在内膜上，而 GspB 的 C 端延伸为肽聚糖细胞壁中的分泌蛋白通道提供了支架。系统发育分析表明，在其他细菌中，GspB 同源物的长度和结构域组成不同，与同源 ATP 酶 GspA 或信号肽 GspS 协同作用。革兰氏阴性菌有两层细胞膜夹着一个肽聚糖网，共同形成一个坚固的保护性细胞膜包膜。为了将效应蛋白穿过这个多层包膜转运，细菌已经进化出几种专门的分泌系统。在 2 型分泌系统和其他一些细菌机制中，分泌蛋白形成大的多聚体孔，允许效应蛋白或纤维穿过外膜。分泌蛋白对于营养物质的获取和致病性至关重要，并且是开发新抗菌药物的目标。将分泌蛋白亚基靶向到外膜通常是由一类特殊的脂蛋白，称为信号肽来促进的。在这里，我们表明在 和其他一些细菌中，支架蛋白 GspB 与信号肽协同作用，促进分泌蛋白孔的组装及其锚定在内膜和细菌细胞壁上。许多其他 T2SS 中都存在具有不同结构域组成的 GspB 同源物。
