Newton Hayley J, Sansom Fiona M, Dao Jenny, McAlister Adrian D, Sloan Joan, Cianciotto Nicholas P, Hartland Elizabeth L
Australian Bacterial Pathogenesis Program, Department of Microbiology, Monash University, Victoria 3800, Australia.
Infect Immun. 2007 Dec;75(12):5575-85. doi: 10.1128/IAI.00443-07. Epub 2007 Sep 24.
The environmental pathogen Legionella pneumophila possesses five proteins with Sel1 repeats (SLRs) from the tetratricopeptide repeat protein family. Three of these proteins, LpnE, EnhC, and LidL, have been implicated in the ability of L. pneumophila to efficiently establish infection and/or manipulate host cell trafficking events. Previously, we showed that LpnE is important for L. pneumophila entry into macrophages and epithelial cells. In further virulence studies here, we show that LpnE is also required for efficient infection of Acanthamoeba castellanii by L. pneumophila and for replication of L. pneumophila in the lungs of A/J mice. In addition, we found that the role of LpnE in host cell invasion is dependent on the eight SLR regions of the protein. A truncated form of LpnE lacking the two C-terminal SLR domains was unable to complement the invasion defect of an lpnE mutant of L. pneumophila 130b in both the A549 and THP-1 cell lines. The lpnE mutant displayed impaired avoidance of LAMP-1 association, suggesting that LpnE influenced trafficking of the L. pneumophila vacuole, similar to the case for EnhC and LidL. We also found that LpnE was present in L. pneumophila culture supernatants and that its export was independent of both the Lsp type II secretion system and the Dot/Icm type IV secretion system. The fact that LpnE was exported suggested that the protein may interact with a eukaryotic protein. Using LpnE as bait, we screened a HeLa cell cDNA library for interacting partners, using the yeast two-hybrid system. Examination of the protein-protein interaction between LpnE and a eukaryotic protein, obscurin-like protein 1, suggested that LpnE can interact with eukaryotic proteins containing immunoglobulin-like folds via the SLR regions. This investigation has further characterized the contribution of LpnE to L. pneumophila virulence and, more specifically, the importance of the SLR regions to LpnE function.
环境病原体嗜肺军团菌拥有五种来自四肽重复蛋白家族且带有Sel1重复序列（SLRs）的蛋白质。其中三种蛋白质，即LpnE、EnhC和LidL，与嗜肺军团菌有效建立感染和/或操纵宿主细胞运输事件的能力有关。此前，我们表明LpnE对嗜肺军团菌进入巨噬细胞和上皮细胞很重要。在本文进一步的毒力研究中，我们表明LpnE对于嗜肺军团菌有效感染卡氏棘阿米巴以及在A/J小鼠肺部的复制也是必需的。此外，我们发现LpnE在宿主细胞侵袭中的作用取决于该蛋白质的八个SLR区域。一种缺少两个C末端SLR结构域的LpnE截短形式无法弥补嗜肺军团菌130b的lpnE突变体在A549和THP-1细胞系中的侵袭缺陷。lpnE突变体在避免与LAMP-1结合方面表现受损，这表明LpnE影响嗜肺军团菌液泡的运输，与EnhC和LidL的情况类似。我们还发现LpnE存在于嗜肺军团菌培养上清液中，并且其分泌独立于Lsp II型分泌系统和Dot/Icm IV型分泌系统。LpnE被分泌这一事实表明该蛋白质可能与真核蛋白质相互作用。我们使用酵母双杂交系统，以LpnE为诱饵，筛选HeLa细胞cDNA文库中的相互作用伙伴。对LpnE与一种真核蛋白质——类 obscurin 蛋白1之间的蛋白质 - 蛋白质相互作用的研究表明，LpnE可以通过SLR区域与含有免疫球蛋白样结构域的真核蛋白质相互作用。这项研究进一步阐明了LpnE对嗜肺军团菌毒力的贡献，更具体地说，是SLR区域对LpnE功能的重要性。
