Wheat yellow dwarf virus disease is infected by barley yellow dwarf virus (BYDV), which causes leaf yellowing and dwarfing symptoms in wheat, thereby posing a serious threat to China's food production. The infection of plant viruses can produce large numbers of vsiRNAs, which can target host transcripts and cause symptom development. However, few studies have been conducted to explore the role played by vsiRNAs in the interaction between BYDV-GAV and host wheat plants. In this study, small RNA sequencing was conducted to profile vsiRNAs in BYDV-GAV-infected wheat plants. The putative targets of vsiRNAs were predicted by the bioinformatics software psRNATarget. RT-qPCR and VIGS were employed to identify the function of selected target transcripts. To confirm the interaction between vsiRNA and the target, 5′ RACE was performed to analyze the specific cleavage sites. From the sequencing data, we obtained a total of 11,384 detected vsiRNAs. The length distribution of these vsiRNAs was mostly 21 and 22 nt, and an A/U bias was observed at the 5′ terminus. We also observed that the production region of vsiRNAs had no strand polarity. The vsiRNAs were predicted to target 23,719 wheat transcripts. GO and KEGG enrichment analysis demonstrated that these targets were mostly involved in cell components, catalytic activity and plant-pathogen interactions. The results of RT-qPCR analysis showed that most chloroplast-related genes were downregulated in BYDV-GAV-infected wheat plants. Silencing of a chlorophyll synthase gene caused leaf yellowing that was similar to the symptoms exhibited by BYDV-GAV-inoculated wheat plants. A vsiRNA from an overlapping region of BYDV-GAV MP and CP was observed to target chlorophyll synthase for gene silencing. Next, 5′ RACE validated that vsiRNA8856 could cleave the chlorophyll synthase transcript in a sequence-specific manner. This report is the first to demonstrate that BYDV-GAV-derived vsiRNAs can target wheat transcripts for symptom development, and the results of this study help to elucidate the molecular mechanisms underlying leaf yellowing after viral infection.

中文翻译：

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大麦黄矮病毒-GAV衍生的vsiRNAs通过靶向叶绿素合酶参与小麦叶片黄化症状的产生

小麦黄矮病毒病是由大麦黄矮病毒(BYDV)感染，引起小麦叶片变黄、出现矮化症状，严重威胁我国粮食生产。植物病毒的感染可以产生大量的vsiRNAs，它们可以靶向宿主转录本并导致症状发展。然而，很少有研究探讨 vsiRNA 在 BYDV-GAV 与宿主小麦植物之间的相互作用中所起的作用。在这项研究中，进行了小 RNA 测序以分析 BYDV-GAV 感染的小麦植物中的 vsiRNA。vsiRNA 的推定靶标由生物信息学软件 psRNATarget 预测。RT-qPCR 和 VIGS 用于鉴定所选目标转录本的功能。为了确认 vsiRNA 和目标之间的相互作用，进行 5' RACE 以分析特定的切割位点。从测序数据中，我们总共获得了 11,384 个检测到的 vsiRNA。这些 vsiRNA 的长度分布主要为 21 和 22 nt，并且在 5' 末端观察到 A/U 偏差。我们还观察到 vsiRNA 的生产区域没有链极性。预计 vsiRNA 可靶向 23,719 个小麦转录本。GO 和 KEGG 富集分析表明，这些靶标主要参与细胞成分、催化活性和植物病原体相互作用。RT-qPCR 分析结果表明，大多数叶绿体相关基因在 BYDV-GAV 感染的小麦植株中下调。叶绿素合酶基因的沉默导致叶片变黄，这与接种 BYDV-GAV 的小麦植物表现出的症状相似。观察到来自 BYDV-GAV MP 和 CP 重叠区域的 vsiRNA 靶向叶绿素合酶进行基因沉默。接下来，5' RACE 验证了 vsiRNA8856 可以以序列特异性方式切割叶绿素合酶转录本。本报告首次证明 BYDV-GAV 衍生的 vsiRNAs 可以针对小麦转录本进行症状发展，这项研究的结果有助于阐明病毒感染后叶片变黄的分子机制。