Protein secretion pathways in the phylum Bacteroidetes

拟杆菌属的成员是迄今为止人类肠道微生物组中最丰富的革兰氏阴性细菌。最近，我们使用质谱法来检查外膜（OM）蛋白质组和分泌组的最常见的研究成员之一的拟杆菌属，B。fragilis，在实验室条件下生长。尽管我们没有发现任何新的分泌途径，但我们发现这种生物体使用的分泌策略与变形菌（例如大肠杆菌）非常不同。杆菌B。fragilis缺乏许多在变形杆菌中广泛存在的途径（例如，II、III和IV型途径），但同时产生多个I型分泌系统。B。fragilis与Proteobacteria的显著不同之处还在于它产生大量暴露在细胞表面上的脂蛋白。I型途径的底物和脂蛋白通过OM转运的机制尚不清楚。有趣的是，在分泌组中发现的许多蛋白质与Genbank数据库中的蛋白质缺乏显著的同源性，并且可能具有新的功能。 我们的一些实验涉及使用B的肠毒素（ETBF）菌株。fragilis，其特征在于分泌称为fragilysin（BFT）的毒素。虽然ETBF菌株引起腹泻并与结直肠癌相关，但它们尚未得到很好的表征。为了方便我们的研究，我们最近获得了四个ETBF菌株的全基因组序列。序列显示，这些菌株在基因组水平上表现出相当大的变异。只有少数主要位于BFT致病岛（BFT PAI）和侧翼CTn86接合转座子的基因是保守的，在所有四个菌株和第五个菌株的基因组先前测序。有趣的是，系统发育分析强烈表明BFT PAI在进化过程中多次被非转基因（NTBF）菌株获得。在表型水平上，我们发现ETBF菌株比NTBF菌株NCTC 9343更不适合，并且对它产生的生长抑制蛋白敏感。ETBF菌株还显示出比NTBF菌株更大的形成生物膜的趋势，这可能促进肿瘤形成。虽然ETBF菌株的基因组多样性提高了它们致病性不同的可能性，但我们的实验结果也表明它们具有由非通用遗传元件的不同组合所赋予的共同特性。

Members of the genus Bacteroides are by far the most abundant Gram-negative bacteria in the human gut microbiome. Recently we used mass spectrometry to examine the outer membrane (OM) proteome and secretome of one of the most commonly studied members of the Bacteroides genus, B. fragilis, grown under laboratory conditions. Although we did not identify any novel secretion pathways, we found that this organism uses a very different range of secretion strategies than Proteobacteria such as E. coli. B. fragilis lacks many of the pathways that are widespread among the Protebacteria (e.g., the type II, III and IV pathways) but produces multiple type I secretion systems simultaneously. B. fragilis also differs dramatically from Proteobacteria in that it produces a large number of lipoproteins that are exposed on the cell surface. The substrates of the type I pathways and the mechanism by which lipoproteins are transported across the OM are unknown. Interestingly, many of the proteins found in the secretome lack significant homology to proteins that are in the Genbank database and presumably have novel functions. Some of our experiments have involved the use of enterotoxigenic (ETBF) strains of B. fragilis that are distinguished by the secretion of a toxin called fragilysin (BFT). Although ETBF strains cause diarrhea and have been associated with colorectal cancer, they have not been well characterized. To facilitate our studies, we recently obtained complete genome sequences of four ETBF strains. The sequences show that these strains exhibit considerable variation at the genomic level. Only a small number of genes that are located primarily in the BFT pathogenicity island (BFT PAI) and the flanking CTn86 conjugative transposon are conserved in all four strains and a fifth strain whose genome was previously sequenced. Interestingly, phylogenetic analysis strongly suggests that the BFT PAI was acquired by non-toxigenic (NTBF) strains multiple times during the course of evolution. At the phenotypic level, we found that the ETBF strains were less fit than the NTBF strain NCTC 9343 and were susceptible to a growth-inhibitory protein that it produces. The ETBF strains also showed a greater tendency to form biofilms, which may promote tumor formation, than NTBF strains. Although the genomic diversity of ETBF strains raises the possibility that they vary in their pathogenicity, our experimental results also suggest that they share common properties that are conferred by different combinations of non-universal genetic elements.