Faculty of Agriculture, Ehime University, Matsuyama, Ehime 790-8566, Japan.
Department of Agronomy, Kansas State University, Manhattan, KS 66506, U.S.A.
Phytopathology. 2020 Jan;110(1):146-152. doi: 10.1094/PHYTO-12-18-0478-FI. Epub 2019 Nov 21.
Cucumber green mottle mosaic virus (CGMMV), a member of the genus , is a major threat to economically important cucurbit crops worldwide. An attenuated strain (SH33b) derived from a severe strain (SH) of CGMMV caused a reduction in the viral RNA accumulation and the attenuation of symptoms, and it has been successfully used to protect muskmelon plants against severe strains in Japan. In this study, we compared GFP-induced silencing suppression by the 129K protein and the methyltransferase domain plus intervening region (MTIR) of the 129K protein between the SH and SH33b strains, respectively. As a result, silencing suppression activity (SSA) in the GFP-silenced plants was inhibited efficiently by the MTIR and 129K protein of SH strain, and it coincided with drastically reduced accumulation of GFP-specific small interfering RNAs (siRNAs) but not by that of SH33b strain. Furthermore, analyses of siRNA binding capability (SBC) by the MTIR of 129K protein and 129K protein using electrophoretic mobility shift assay revealed that SBC was found with the MTIR and 129K protein of SH but not with that of SH33b, suggesting that a single amino acid mutation (E to G) in the MTIR is responsible for impaired SSA and SBC of SH33b. These data suggest that a single amino acid substitution in the intervening region of 129K protein of CGMMV resulted in attenuated symptoms by affecting RNA silencing suppression.
黄瓜绿斑驳花叶病毒（CGMMV），属于 属，是全球重要经济作物葫芦科作物的主要威胁。从强毒株（SH）衍生而来的弱毒株（SH33b）导致病毒 RNA 积累减少和症状减弱，已成功用于保护日本的厚皮甜瓜植物免受强毒株的侵害。在本研究中，我们比较了 SH 和 SH33b 株系的 129K 蛋白和甲基转移酶结构域加间隔区（MTIR）对 GFP 诱导的沉默抑制作用。结果表明，SH 株系的 MTIR 和 129K 蛋白可有效抑制 GFP 沉默植物中的沉默抑制活性（SSA），并且与 GFP 特异性小干扰 RNA（siRNA）积累明显减少一致，但 SH33b 株系则没有。此外，通过电泳迁移率变动分析对 129K 蛋白的 MTIR 和 129K 蛋白的 siRNA 结合能力（SBC）进行分析表明，SH 株系的 MTIR 和 129K 蛋白具有 SBC，但 SH33b 株系则没有，表明 MTIR 中的单个氨基酸突变（E 到 G）导致 SH33b 的 SSA 和 SBC 受损。这些数据表明，CGMMV 129K 蛋白间隔区中的单个氨基酸替换通过影响 RNA 沉默抑制导致症状减弱。
