RNA-dependent RNA polymerase 1 of Nicotiana tabacum modulates ToLCGV pathogenesis by influencing a number of defence-related genes in N. benthamiana plants. Key means of plants protecting themselves from the invading viruses is through RNA silencing. RNA-dependent RNA polymerase-1 (RDR1) is one of the crucial proteins of the RNA silencing pathway, which is induced after infection by viruses. RDR1 functions in the generation of small interfering RNAs (siRNAs) against the viral genome, thus it is antiviral in nature. Here, we used the transgenic Nicotiana benthamiana plant expressing N. tabacum NtRDR1 and observed reduced susceptibility towards Tomato leaf curl Gujarat virus (ToLCGV) infection compared to the wild-type N. benthamiana plants. To understand the reason for such reduced susceptibility, we prepared high-definition small RNA (sRNA) cDNA libraries from ToLCGV-infected wild-type N. benthamiana and NtRDR1 expressing N. benthamiana lines and carried out next-generation sequencing (NGS). We found that upon ToLCGV infection the majority of siRNAs generated from the host genome were of the 24 nucleotide (nt) class, while viral siRNAs (vsiRNAs) were of the 21–22-nt class, indicating that transcriptional gene silencing (TGS) is the major pathway for silencing of host genes while viral genes are silenced, predominantly, by post transcriptional gene silencing (PTGS) pathways. We estimated the changes in the expression of various defence-related genes, such as Constitutively Photomorphogenic-9 (COP9) signalosome (CSN) complex subunit-7, Pentatricopeptide repeat containing protein (PPRP), Laccase-3, Glutathione peroxidase-1 (GPX-1), Universal stress protein (USP) A-like protein, Heat shock transcription factor B4 (HSTF-B4), Auxin response factor-18 (ARF18), WRKY-6 and Short chain dehydrogenase reductase-3a. The differential expression of these genes might be linked with the enhanced tolerance of NtRDR1 N. benthamiana transgenic plants to ToLCGV. Our study suggests that reduced expression of subunit-7 of CSN complex and WRKY6, and increased expression of USPA-like protein might be linked with the reduced susceptibility of NtRDR1-transgenic N. benthamiana plants to ToLCGV.

中文翻译：

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烟草RNA依赖性RNA聚合酶1影响番茄卷叶古吉拉特病毒感染后本塞姆氏烟草中防御相关基因的表达


烟草的 RNA 依赖性 RNA 聚合酶 1 通过影响本塞姆氏烟草植物中的许多防御相关基因来调节 ToLCGV 发病机制。植物保护自身免受病毒入侵的关键手段是通过 RNA 沉默。 RNA 依赖性 RNA 聚合酶-1 (RDR1) 是病毒感染后诱导的 RNA 沉默途径的关键蛋白之一。 RDR1 的功能是产生针对病毒基因组的小干扰 RNA (siRNA)，因此它本质上具有抗病毒作用。在这里，我们使用表达烟草 NtRDR1 的转基因本塞姆氏烟草植物，并观察到与野生型本塞姆氏烟草植物相比，对古吉拉特邦番茄卷叶病毒 (ToLCGV) 感染的敏感性降低。为了了解敏感性降低的原因，我们从感染 ToLCGV 的野生型本塞姆氏烟草和表达 NtRDR1 的本氏烟草品系中制备了高清小 RNA (sRNA) cDNA 文库，并进行了下一代测序 (NGS)。我们发现，ToLCGV 感染后，宿主基因组产生的大多数 siRNA 属于 24 核苷酸 (nt) 类，而病毒 siRNA (vsiRNA) 属于 21-22 nt 类，表明转录基因沉默 (TGS)宿主基因沉默的主要途径，而病毒基因主要通过转录后基因沉默（PTGS）途径沉默。 我们估计了各种防御相关基因的表达变化，例如组成型光形态发生-9 (COP9) 信号体 (CSN) 复合体亚基-7、五肽重复含有蛋白 (PPRP)、Laccase-3、谷胱甘肽过氧化物酶-1 (GPX) -1)、通用应激蛋白 (USP) A 样蛋白、热休克转录因子 B4 (HSTF-B4)、生长素反应因子-18 (ARF18)、WRKY-6 和短链脱氢酶还原酶-3a。这些基因的差异表达可能与 NtRDR1 本塞姆氏烟草转基因植物对 ToLCGV 的耐受性增强有关。我们的研究表明，CSN 复合体和 WRKY6 亚基 7 表达的减少以及 USPA 样蛋白表达的增加可能与 NtRDR1 转基因本塞姆氏烟草植物对 ToLCGV 的敏感性降低有关。