Involvement of a plasmid-encoded type IV secretion system in the plant tissue watersoaking phenotype of Burkholderia cenocepacia
Involvement of a plasmid-encoded type IV secretion system in the plant tissue watersoaking phenotype of Burkholderia cenocepacia
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DOI：
10.1128/jb.186.18.6015-6024.2004
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发表时间：
2004-09-01
影响因子：
3.2
通讯作者：
Gonzalez, CF
中科院分区：
文献类型：
--
作者：
Engledow, AS;Medrano, EG;Gonzalez, CF
洋葱伯克霍尔德氏菌复合体K56-2株是洋葱伯克霍尔德氏菌复合体的代表菌株，是洋葱伯克霍尔德氏菌复合体的一部分，属于临床上有问题的ET12谱系。该菌株在洋葱组织上产生植物组织浸水(PTW)，这是植物病害相关的特征。利用质粒子诱变技术，获得了PTW表型的突变体。来自ptw阴性突变体的中断基因(PtwD4)的翻译序列与VirD4样蛋白同源。对转移基因同源物近端区域的分析发现，位于92kb驻留质粒上的基因簇与IV型分泌系统同源。通过定点突变，确定ptwD4、PTWC、ptwB4和ptwB10在PTW活性表达中的作用。在ptwD4、ptwB4或ptwB10中断的K56-2衍生物中不表达PTW表型，但在PTWC中断的衍生物中观察到PTW表型。PTW阴性的K56-2衍生物在反式中的互补导致PTW表型完全恢复。此外，对培养上清液的分析表明，推测的Ptw效应蛋白(S)是一种分泌的、热稳定的蛋白(S)，它能引起植物原生质体的质壁分离。在K56-2中发现了与VIRB-VirD系统完全同源的第二个染色体编码的IV型分泌系统。VIRB-VirD系统中关键分泌基因的定点突变不影响PTW表型的表达。我们的发现表明，在K56-2菌株中，pTW IV型分泌系统负责分泌一种植物细胞毒蛋白(S)。
Burkholderia cenocepacia strain K56-2, a representative of the Burkholderia cepacia complex, is part of the epidemic and clinically problematic ET12 lineage. The strain produced plant tissue watersoaking (ptw) on onion tissue, which is a plant disease-associated trait. Using plasposon mutagenesis, mutants in the ptw phenotype were generated. The translated sequence of a disrupted gene (ptwD4) from a ptw-negative mutant showed homology to VirD4-like proteins. Analysis of the region proximal to the transfer gene homolog identified a gene cluster located on the 92-kb resident plasmid that showed homology to type IV secretion systems. The role of ptwD4, ptwC, ptwB4, and ptwB10 in the expression of ptw activity was determined by conducting site-directed mutagenesis. The ptw phenotype was not expressed by K56-2 derivatives with a disruption in ptwD4, ptwB4, or ptwB10 but was observed in a derivative with a disruption in ptwC. Complementation of ptw-negative K56-2 derivatives in trans resulted in complete restoration of the ptw phenotype. In addition, analysis of culture supernatants revealed that the putative ptw effector(s) was a secreted, heat-stable protein(s) that caused plasmolysis of plant protoplasts. A second chromosomally encoded type IV secretion system with complete homology to the VirB-VirD system was identified in K56-2. Site-directed mutagenesis of key secretory genes in the VirB-VirD system did not affect expression of the ptw phenotype. Our findings indicate that in strain K56-2, the plasmid-encoded Ptw type IV secretion system is responsible for the secretion of a plant cytotoxic protein(s).