Michel Gérard P F, Durand Eric, Filloux Alain
Laboratoire d'Ingénierie des Systèmes Macromoléculaires, Institut de Biologie Structurale et Microbiologie, Centre National de la Recherche Scientifique, 31Chemin J. Aiguier, 13402 Marseille, France.
J Bacteriol. 2007 May;189(10):3776-83. doi: 10.1128/JB.00205-07. Epub 2007 Mar 9.
The opportunistic human pathogen bacterium Pseudomonas aeruginosa secretes various exoproteins in its surrounding environment. Protein secretion involves different secretory systems, including the type II secretion system, or T2SS, that is one of the most efficient secretory pathways of P. aeruginosa. There are two T2SS in this bacterium, the quorum-sensing-regulated Xcp system and the Hxc system, which is only present under phosphate-limiting conditions. Like T2SS of other bacteria, the Xcp T2SS is species specific, and this specificity mainly involves two proteins, XcpP (GspC family) and the secretin XcpQ (GspD family), which are the gatekeepers of the system. Interestingly, an orphan secretin, XqhA, was previously reported as being able to functionally replace the XcpQ secretin. In this study, we identified another gene, which we named xphA (xcpP homologue A), which is located next to xqhA. We showed that deletion of the xphA gene in an xcpP mutant caused the disappearance of the residual secretion observed in this mutant strain, indicating that the protein XphA plays a role in the secretion process. Our results also revealed that complementation of an xcpP/xcpQ mutant can be obtained with the gene couple xphA/xqhA. The XphA and XqhA proteins (the P(A)Q(A) subunit) could thus form, together with XcpR-Z, a functional hybrid T2SS. A two-dimensional polyacrylamide gel electrophoresis analysis showed that except for the aminopeptidase PaAP, for which secretion is not restored by the P(A)Q(A) subunit in the xcpP/xcpQ deletion mutant, each major Xcp-dependent exoprotein is secreted by the new hybrid machinery. Our work supports the idea that components of the GspC/GspD families, such as XphA/XqhA or XcpP/XcpQ, are assembled as a specific tandem within the T2SS. Each of these pairs may thus confer a different level of secretion specificity, as is the case with respect to PaAP. Finally, using a chromosomal xphA-lacZ fusion, we showed that the xphA-xqhA genes are transcribed from an early stage of bacterial growth. We thus suggest that the P(A)Q(A) subunit might be involved in the secretion process at a different growth stage than XcpP/XcpQ.
机会性人类病原菌铜绿假单胞菌在其周围环境中分泌多种胞外蛋白。蛋白质分泌涉及不同的分泌系统，包括II型分泌系统（T2SS），它是铜绿假单胞菌最有效的分泌途径之一。该细菌中有两个T2SS，即群体感应调节的Xcp系统和仅在磷酸盐限制条件下存在的Hxc系统。与其他细菌的T2SS一样，Xcp T2SS具有种属特异性，这种特异性主要涉及两种蛋白质，XcpP（GspC家族）和分泌素XcpQ（GspD家族），它们是该系统的守门人。有趣的是，先前报道孤儿分泌素XqhA能够在功能上替代XcpQ分泌素。在本研究中，我们鉴定了另一个基因，命名为xphA（xcpP同源物A），它位于xqhA旁边。我们发现，在xcpP突变体中缺失xphA基因会导致该突变菌株中观察到的残余分泌消失，这表明蛋白质XphA在分泌过程中起作用。我们的结果还表明，基因对xphA/xqhA可以互补xcpP/xcpQ突变体。因此，XphA和XqhA蛋白（P(A)Q(A)亚基）可以与XcpR-Z一起形成功能性杂交T2SS。二维聚丙烯酰胺凝胶电泳分析表明，除了氨基肽酶PaAP（在xcpP/xcpQ缺失突变体中，P(A)Q(A)亚基不能恢复其分泌）外，每个主要的Xcp依赖性胞外蛋白都由新的杂交机制分泌。我们的工作支持这样一种观点，即GspC/GspD家族的组分，如XphA/XqhA或XcpP/XcpQ，在T2SS中作为特定的串联体组装。因此，这些对中的每一对可能赋予不同水平的分泌特异性，就像PaAP的情况一样。最后，使用染色体xphA-lacZ融合，我们表明xphA-xqhA基因从细菌生长的早期阶段开始转录。因此，我们认为P(A)Q(A)亚基可能在与XcpP/XcpQ不同的生长阶段参与分泌过程。
