School of Optometry, University of California, Berkeley, CA 94720, USA.
Microb Pathog. 2011 Nov;51(5):305-12. doi: 10.1016/j.micpath.2011.08.001. Epub 2011 Aug 9.
We previously showed that ADP-ribosylation (ADP-r) activity of ExoS, a type III secreted toxin of Pseudomonas aeruginosa, enables bacterial replication in corneal and respiratory epithelial cells and correlates with bacterial trafficking to plasma membrane blebs (bleb-niche formation). Here, we explored another type III secreted toxin, ExoY, for its impact on intracellular trafficking and survival, and for virulence in vivo using a murine corneal infection model. Chromosomal or plasmid-mediated expression of exoY in invasive P. aeruginosa (strain PAO1) enabled bacteria to form and traffic to epithelial membrane blebs in the absence of other known effectors. In contrast, plasmid expression of any of four adenylate cyclase mutant forms of exoY did not enable bleb-niche formation, and bacteria localized to perinuclear vacuoles as for effector-null mutant controls. None of the plasmid-complemented bacteria used in this study showed ADP-r activity in the absence of ExoS and ExoT. In contrast to ADP-r activity of ExoS, bleb-niche formation induced by ExoY's adenylate cyclase activity was not accompanied by enhanced intracellular replication. In vivo results showed that ExoY-adenylate cyclase activity promoted P. aeruginosa corneal virulence in susceptible mice. Together the data show that adenylate cyclase activity of P. aeruginosa ExoY, similarly to the ADP-r activity of ExoS, can mediate bleb-niche formation in epithelial cells. While this activity did not promote intracellular replication in vitro, ExoY conferred increased virulence in vivo in susceptible mice. Mechanisms for bleb-niche formation and relationships to intracellular replication and virulence in vivo require further investigation for both ExoS and ExoY.
我们之前曾表明，铜绿假单胞菌 III 型分泌毒素 ExoS 的 ADP-核糖基化（ADP-r）活性使细菌能够在角膜和呼吸道上皮细胞中复制，并与细菌向质膜泡（泡巢形成）的运输相关。在此，我们使用鼠角膜感染模型，研究了另一种 III 型分泌毒素 ExoY 对细胞内运输和存活的影响及其在体内的毒力。在侵袭性铜绿假单胞菌（PAO1 株）中，染色体或质粒介导的 exoY 表达使细菌能够在没有其他已知效应物的情况下形成并运输到上皮细胞膜泡。相比之下，质粒表达任何四种腺嘌呤核苷酸环化酶突变形式的 exoY 都不能形成泡巢，并且细菌像效应物缺失突变体对照一样定位于核周空泡。在这项研究中使用的任何质粒互补细菌都没有显示 ExoS 和 ExoT 缺失时的 ADP-r 活性。与 ExoS 的 ADP-r 活性相反，由 ExoY 的腺苷酸环化酶活性诱导的泡巢形成并没有伴随着增强的细胞内复制。体内结果表明，ExoY-腺嘌呤核苷酸环化酶活性促进了易感小鼠中铜绿假单胞菌的角膜毒力。这些数据表明，铜绿假单胞菌 ExoY 的腺苷酸环化酶活性，类似于 ExoS 的 ADP-r 活性，可介导上皮细胞中的泡巢形成。虽然这种活性在体外没有促进细胞内复制，但 ExoY 在易感小鼠中赋予了更高的体内毒力。需要进一步研究 ExoS 和 ExoY 以了解泡巢形成的机制以及与体内细胞内复制和毒力的关系。
