RNAi screen reveals an Abl kinase-dependent host cell pathway involved in Pseudomonas aeruginosa internalization.
RNAi screen reveals an Abl kinase-dependent host cell pathway involved in Pseudomonas aeruginosa internalization.
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DOI：
10.1371/journal.ppat.1000031
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发表时间：
2008-03-21
期刊：
影响因子：
6.7
通讯作者：
Engel, Joanne N.
中科院分区：
文献类型：
--
作者：
Pielage, Julia F.;Powell, Kimberly R.;Kalman, Daniel;Engel, Joanne N.
致病细菌铜绿假单胞菌被非吞噬细胞内化是由肌动蛋白细胞骨架的重排促进的，但被这种细菌篡夺的宿主途径尚不清楚。我们使用rnai介导的基因失活已知调控果蝇S2细胞中肌动蛋白细胞骨架的约80个基因，以鉴定铜绿假单胞菌进入所需的宿主分子。这项工作揭示了Abl酪氨酸激酶、接头蛋白Crk、小gtpase Rac1和Cdc42以及p21活化激酶是导致铜绿假单胞菌内化的宿主信号通路的组成部分。使用多种互补方法，我们验证了该途径在哺乳动物细胞中的作用。值得注意的是，P. aeruginosa的III型分泌毒素ExoS和ExoT通过干扰GTPase功能来靶向这一途径，并且在ExoT的情况下，通过消除P. aeruginosa诱导的abl依赖性Crk磷酸化。总之，这项工作揭示了铜绿假单胞菌利用Abl途径进入宿主细胞，并揭示了铜绿假单胞菌III型分泌系统调节这一内化途径的意想不到的复杂性。我们的研究结果进一步证明了使用RNAi筛选来识别被微生物病原体篡位的宿主信号级联的适用性，这些微生物病原体可能是针对抗生素耐药感染治疗的新疗法的潜在靶点。铜绿假单胞菌感染是医院获得性感染的主要原因之一，其死亡率接近40%，多重耐药感染很常见，而且还在增加。宿主细胞对铜绿假单胞菌的内化似乎在这种机会性细菌的发病机制中起着重要作用，但参与这一过程的宿主细胞因素尚不完全清楚。我们在果蝇S2细胞中使用靶向RNAi筛选来鉴定宿主肌动蛋白细胞骨架的一个调节子子集，这些调节子有助于细菌进入，并证实它们参与了哺乳动物细胞的感染。我们发现铜绿假单胞菌可以通过其III型分泌系统将细菌毒素ExoS和ExoT注入宿主细胞，以一种复杂的方式调节这一内化途径。所鉴定的宿主细胞分子可作为新型药物的靶标，用于治疗对常规抗生素具有耐药性的感染，并可适用于广泛的病原体。
Internalization of the pathogenic bacterium Pseudomonas aeruginosa by non-phagocytic cells is promoted by rearrangements of the actin cytoskeleton, but the host pathways usurped by this bacterium are not clearly understood. We used RNAi-mediated gene inactivation of ∼80 genes known to regulate the actin cytoskeleton in Drosophila S2 cells to identify host molecules essential for entry of P. aeruginosa. This work revealed Abl tyrosine kinase, the adaptor protein Crk, the small GTPases Rac1 and Cdc42, and p21-activated kinase as components of a host signaling pathway that leads to internalization of P. aeruginosa. Using a variety of complementary approaches, we validated the role of this pathway in mammalian cells. Remarkably, ExoS and ExoT, type III secreted toxins of P. aeruginosa, target this pathway by interfering with GTPase function and, in the case of ExoT, by abrogating P. aeruginosa–induced Abl-dependent Crk phosphorylation. Altogether, this work reveals that P. aeruginosa utilizes the Abl pathway for entering host cells and reveals unexpected complexity by which the P. aeruginosa type III secretion system modulates this internalization pathway. Our results furthermore demonstrate the applicability of using RNAi screens to identify host signaling cascades usurped by microbial pathogens that may be potential targets for novel therapies directed against treatment of antibiotic-resistant infections. Mortality from Pseudomonas aeruginosa infections, one of the leading causes of hospital acquired infections, approaches 40%, and multiple drug resistant infections are common and increasing. Internalization of P. aeruginosa by the host cell appears to play a fundamental role in the pathogenesis of this opportunistic bacterium, but the host cell factors involved in this process are incompletely understood. We used a targeted RNAi screen in Drosophila S2 cells to identify a subset of regulators of the host actin cytoskeleton that contribute to bacterial entry and confirmed their involvement in infection of mammalian cells. We found that P. aeruginosa can modulate this internalization pathway in a complex manner by injecting the bacterial toxins ExoS and ExoT into the host cell via its type III secretion system. The identified host cell molecules may serve as targets for novel drugs to treat infections resistant to conventional antibiotics and may be applicable to a wide range of pathogens.
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DOI：
10.1073/pnas.0506461102
发表时间：
2005-09-20
影响因子：
11.1
作者：
Cheng, LW;Viala, JPM;Portnoy, DA
通讯作者：
Portnoy, DA
影响因子：
4.8
作者：
Deng, Q;Sun, JJ;Barbieri, JT
通讯作者：
Barbieri, JT
影响因子：
3.1
作者：
Hauser, AR;Fleiszig, S;Engel, JN
通讯作者：
Engel, JN
DOI：
10.1006/bbrc.2002.6402
发表时间：
2002-02-15
影响因子：
3.1
作者：
Fraylick, JE;Rucks, EA;Olson, JC
通讯作者：
Olson, JC
影响因子：
3.1
作者：
Hauser, AR;Engel, JN
通讯作者：
Engel, JN