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Abstract：
Viruses employ a series of diverse translational strategies to expand their coding capacity, which produces viral proteins with common domains and entangles virus-host interactions. P3N-PIPO, which is a transcriptional slippage product from the P3 cistron, is a potyviral protein dedicated to intercellular movement. Here, we show that P3N-PIPO from watermelon mosaic virus (WMV) triggers cell death when transiently expressed in Cucumis melo accession PI 414723 carrying the Wmr resistance gene. Surprisingly, expression of the P3N domain, shared by both P3N-PIPO and P3, can alone induce cell death, whereas expression of P3 fails to activate cell death in PI 414723. Confocal microscopy analysis revealed that P3N-PIPO targets plasmodesmata (PD) and P3N associates with PD, while P3 localizes in endoplasmic reticulum in melon cells. We also found that mutations in residues L35, L38, P41, and I43 of the P3N domain individually disrupt the cell death induced by P3N-PIPO, but do not affect the PD localization of P3N-PIPO. Furthermore, WMV mutants with L35A or I43A can systemically infect PI 414723 plants. These key residues guide us to discover some WMV isolates potentially breaking the Wmr resistance. Through searching the NCBI database, we discovered some WMV isolates with variations in these key sites, and one naturally occurring I43V variation enables WMV to systemically infect PI 414723 plants. Taken together, these results demonstrate that P3N-PIPO, but not P3, is the avirulence determinant recognized by Wmr, although the shared N terminal P3N domain can alone trigger cell death.IMPORTANCEThis work reveals a novel viral avirulence (Avr) gene recognized by a resistance (R) gene. This novel viral Avr gene is special because it is a transcriptional slippage product from another virus gene, which means that their encoding proteins share the common N-terminal domain but have distinct C-terminal domains. Amazingly, we found that it is the common N-terminal domain that determines the Avr-R recognition, but only one of the viral proteins can be recognized by the R protein to induce cell death. Next, we found that these two viral proteins target different subcellular compartments. In addition, we discovered some virus isolates with variations in the common N-terminal domain and one naturally occurring variation that enables the virus to overcome the resistance. These results show how viral proteins with common domains interact with a host resistance protein and provide new evidence for the arms race between plants and viruses.
摘要：
病毒采用一系列不同的翻译策略来扩展其编码能力，产生具有共同结构域的病毒蛋白，并纠缠病毒与宿主的相互作用。P3N-PIPO,这是P3顺反子的转录滑移产物，是一种专门用于细胞间运动的potyvirus蛋白。这里,我们表明，西瓜花叶病毒（WMV）的P3N-PIPO在带有Wmr抗性基因的黄瓜甜瓜登录名PI414723中瞬时表达时，会触发细胞死亡。令人惊讶的是,P3N结构域的表达，P3N-PIPO和P3共享，可以单独诱导细胞死亡，而P3的表达未能激活PI414723中的细胞死亡。共聚焦显微镜分析显示，P3N-PIPO靶向胞浆（PD）和P3N与PD相关，而P3定位于甜瓜细胞的内质网。我们还发现，P3N结构域的残基L35、L38、P41和I43的突变分别破坏了P3N-PIPO诱导的细胞死亡，但不影响P3N-PIPO的PD定位。此外，具有L35A或I43A的WMV突变体可以系统地感染PI414723植物。这些关键残基指导我们发现一些可能破坏Wmr抗性的WMV分离株。通过搜索NCBI数据库，我们发现了一些WMV分离株在这些关键位点有变异，和一个天然存在的I43V变异使WMV能够系统地感染PI414723植物。一起来看,这些结果表明，P3N-PIPO，但不是P3，是Wmr认可的无毒力决定因素，尽管共享的N末端P3N域可以单独触发细胞死亡。重要性这项工作揭示了一种由抗性（R）基因识别的新型病毒无毒力（Avr）基因。这种新型病毒Avr基因是特殊的，因为它是另一种病毒基因的转录滑移产物，这意味着它们的编码蛋白质共享共同的N末端结构域，但具有不同的C末端结构域。令人惊讶的是,我们发现确定Avr-R识别的是常见的N端域，但是只有一种病毒蛋白可以被R蛋白识别以诱导细胞死亡。接下来,我们发现这两种病毒蛋白靶向不同的亚细胞区室。此外,我们发现了一些病毒分离株，其常见的N端结构域存在变异，并且有一个天然存在的变异使病毒能够克服耐药性。这些结果显示了具有共同结构域的病毒蛋白如何与宿主抗性蛋白相互作用，并为植物和病毒之间的军备竞赛提供了新的证据。
