Exploring Different Mutations at a Single Amino Acid Position of Cucumber green mottle mosaic virus Replicase to Attain Stable Symptom Attenuation
Exploring Different Mutations at a Single Amino Acid Position of Cucumber green mottle mosaic virus Replicase to Attain Stable Symptom Attenuation
复制标题
探索黄瓜绿斑驳花叶病毒复制酶单个氨基酸位置的不同突变以获得稳定的症状减弱
DOI：
10.1094/phyto-03-17-0107-r
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发表时间：
2017-09-01
期刊：
影响因子：
3.2
通讯作者：
Gu, Qinsheng
中科院分区：
文献类型：
--
作者：
Liu, Liming;Peng, Bin;Gu, Qinsheng
黄瓜绿色斑驳花叶病毒（CGMMV）是乙型病毒属（处女病毒科）的一种，对瓜类作物造成严重的经济损失。控制CGMMV的一种可能性是使用交叉保护，这需要稳定的减毒分离物。本研究构建了CGMMV hn分离株的感染性克隆。出乎意料的是，该克隆携带一个非保守突变，涉及单个核苷酸变化，导致复制酶蛋白残基284上的Arg被Cys取代；该突变与延迟症状诱导和RNA积累相关，这在时间过程实验中得到证实。病毒后代的测序显示，野生型症状的恢复和RNA积累的增加与突变向野生型序列的逆转相关，这一现象发生在接种后约7至10天。因此，Arg284似乎是至关重要的，但不是病毒感染的严格必要条件。随后，在编码Arg284的三联体中构建了另外四个突变体并进行了分析。结果表明，不同突变体的症状和发病时间不同，致病性增强和RNA积累均与Arg284的逆转相关。因此，突变的性质强烈地影响了突变体的遗传稳定性。至少有两个突变体在接种后30天内没有发生逆转，这些突变体被定义为获得稳定症状衰减的良好候选者，可用于交叉保护。
Cucumber green mottle mosaic virus (CGMMV) is a member of the genus Tobamovirus (family Virgaviridae) that causes serious economic losses in cucurbit crops. A possibility for CGMMV control is the use of cross-protection, for which stable attenuated isolates are required. In this study, an infectious clone was constructed for the hn isolate of CGMMV. Unexpectedly, this clone carried a nonconserved mutation involving a single nucleotide change resulting in the replacement of Arg by Cys at residue 284 of the replicase protein; this mutation correlated with delayed symptom induction and RNA accumulation, as shown in time-course experiments. Sequencing of the viral progeny showed that restoration of wild-type symptoms and increased RNA accumulation correlated with reversion of the mutation to the wild-type sequence, a phenomenon that occurred at approximately 7 to 10 days postinoculation. Thus, Arg284 seems to be crucial but not strictly necessary for virus infection. Subsequently, four other mutants in the triplet encoding Arg284 were constructed and assayed. Results showed that symptoms and their timing were diverse for the different mutants, with enhanced pathogenicity and RNA accumulation always correlating with reversion to Arg284. Therefore, the nature of the mutation strongly influenced the genetic stability of the mutant. At least two mutants were identified for which reversion did not occur by 30 days postinoculation, and these were defined as good candidates to attain stable symptom attenuation that could be useful in cross-protection.