Bretz James R, Mock Norton M, Charity James C, Zeyad Syed, Baker C Jacyn, Hutcheson Steven W
Department of Cell Biology and Molecular Genetics, University of Maryland, College Park, MD 20742, USA.
Mol Microbiol. 2003 Jul;49(2):389-400. doi: 10.1046/j.1365-2958.2003.03616.x.
Pseudomonas syringae strains translocate effector proteins into host cells via the hrp-encoded type III protein secretion system (TTSS) to facilitate pathogenesis in susceptible plants. However, the mechanisms by which pathogenesis is favoured by these effectors are not well understood. Individual strains express multiple effectors with apparently distinct activities that are co-ordinately regulated by the alternative sigma factor HrpL. Genes for several effectors were identified in the P. syringae pv. tomato DC3000 genome using a promoter trap assay to identify HrpL-dependent promoters. In addition to orthologues of avrPphE and hrpW, an unusual allele of avrPphD was detected that carried an IS52 insertion. Using this avrPphD::IS52 allele as a probe, a wild-type allele of avrPphD, hopPtoD1, and a chimeric homologue were identified in the DC3000 genome. This chimeric homologue, identified as HopPtoD2 in the annotated DC3000 genome, consisted of an amino terminal secretion domain similar to that of AvrPphD fused to a potential protein tyrosine phosphatase domain. Culture filtrates of strains expressing HopPtoD2 were able to dephosphorylate pNPP and two phosphotyrosine peptides. HopPtoD2 was shown to be translocated into Arabidopsis thaliana cells via the hrp-encoded TTSS. A DeltahopPtoD2 mutant of DC3000 exhibited strongly reduced virulence in Arabidopsis thaliana. Ectopic expression of hopPtoD2 in P. syringae Psy61 that lacks a native hopPtoD2 orthologue delayed the development of several defence-associated responses including programmed cell death, active oxygen production and transcription of the pathogenesis-related gene PR1. The results indicate that HopPtoD2 is a translocated effector with protein tyrosine phosphatase activity that modulates plant defence responses.
丁香假单胞菌菌株通过hrp编码的III型蛋白分泌系统（TTSS）将效应蛋白转运到宿主细胞中，以促进在易感植物中的致病作用。然而，这些效应蛋白促进致病作用的机制尚未完全了解。单个菌株表达多种具有明显不同活性的效应蛋白，这些效应蛋白由替代sigma因子HrpL协同调节。使用启动子陷阱试验来鉴定HrpL依赖性启动子，在丁香假单胞菌番茄致病变种DC3000基因组中鉴定了几种效应蛋白的基因。除了avrPphE和hrpW的直系同源物外，还检测到一个不寻常的avrPphD等位基因，该等位基因携带一个IS52插入片段。使用这个avrPphD::IS52等位基因作为探针，在DC3000基因组中鉴定出avrPphD的野生型等位基因hopPtoD1和一个嵌合同源物。在注释的DC3000基因组中，这个嵌合同源物被鉴定为HopPtoD2，它由一个类似于AvrPphD的氨基末端分泌结构域与一个潜在的蛋白酪氨酸磷酸酶结构域融合而成。表达HopPtoD2的菌株的培养滤液能够使对硝基苯磷酸酯（pNPP）和两种磷酸酪氨酸肽去磷酸化。结果表明，HopPtoD2能够通过hrp编码的TTSS转运到拟南芥细胞中。DC3000的DeltahopPtoD2突变体在拟南芥中的毒力显著降低。在缺乏天然hopPtoD2直系同源物的丁香假单胞菌Psy61中异位表达hopPtoD2，延迟了几种防御相关反应的发展，包括程序性细胞死亡、活性氧产生和病程相关基因PR1的转录。结果表明，HopPtoD2是一种具有蛋白酪氨酸磷酸酶活性的转运效应蛋白，可调节植物防御反应。
