Defects in type III secretion correlate with internalization of Pseudomonas aeruginosa by epithelial cells
Defects in type III secretion correlate with internalization of Pseudomonas aeruginosa by epithelial cells
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DOI：
10.1128/iai.66.4.1413-1420.1998
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发表时间：
1998-04-01
影响因子：
3.1
通讯作者：
Engel, JN
中科院分区：
文献类型：
--
作者：
Hauser, AR;Fleiszig, S;Engel, JN
先前对铜绿假单胞菌临床分离株的表征已证明细胞毒性与上皮细胞内化之间呈负相关。为了进一步研究这种关系，我们测试了PA103，一种细胞毒性铜绿假单胞菌菌株，和33个同基因非细胞毒性转座子突变体的Madin-Darby犬肾细胞内化。大多数突变体没有被内化，表明细胞毒性和上皮细胞的细菌摄取之间的负相关性不是绝对的。六个非细胞毒性突变体，然而，表现出可测量的内化水平，通过标准的氨基糖苷类排除试验，即使野生型菌株PA103的内化是不可检测的。所有六个人都有蛋白质分泌缺陷的证据，涉及两种蛋白质，一种40 kDa蛋白质和一种32 kDa蛋白质。这些蛋白质分别命名为PepB（假单胞菌胞外蛋白B）和PepD，每种蛋白质都具有III型转运蛋白的特征。此外，核苷酸测序研究表明PepB和PepD分别是耶尔森氏菌属的III型分泌蛋白YopB和YopD的同源物。这是将效应分子转运到真核细胞的细胞质区室所必需的。因此，虽然PA103中的许多突变导致细胞毒性的丧失而没有内化的明显增加，但III型分泌蛋白PepB和PepD的转运缺陷与细胞毒性的丧失和内化的获得两者相关。这些结果与需要PepB和PepD易位到宿主细胞中的内化抑制剂的III型分泌一致。
Previous characterization of Pseudomonas aeruginosa clinical isolates has demonstrated an inverse correlation between cytotoxicity and internalization by epithelial cells. To further investigate this relationship, we tested PA103, a cytotoxic P. aeruginosa strain, and 33 isogenic noncytotoxic transposon mutants for internalization by Madin-Darby canine kidney cells. The majority of the mutants were not internalized, demonstrating that an inverse correlation between cytotoxicity and bacterial uptake by epithelial cells is not absolute. Six of the noncytotoxic mutants, however, demonstrated measurable levels of internalization by standard aminoglycoside exclusion assays even though internalization of wild-type strain PA103 was not detectable. All six had evidence of protein secretion defects involving two proteins, a 40-kDa protein and a 32-kDa protein. These proteins, designated PepB (for Pseudomonas exoprotein B) and PepD, respectively, each had characteristics of type III transported proteins. In addition, nucleotide sequencing studies demonstrated that PepB and PepD are homologs of YopB and YopD, respectively, type III secreted proteins of Yersinia spp. necessary for the translocation of effector molecules into the cytoplasmic compartment of eukaryotic cells. Thus, while many mutations in PA103 result in loss of cytotoxicity without an appreciable increase in internalization, defects in transport of type III secretion proteins PepB and PepD correlate with both loss of cytotoxicity and gain of internalization. These results are consistent with type III secretion of an inhibitor of internalization that requires PepB and PepD for translocation into the host cell.