Table_1_Integrated transcriptome and microRNA sequencing analyses reveal gene responses in poplar leaves infected by the novel pathogen bean common mosaic virus (BCMV).xls

Recently, a novel poplar mosaic disease caused by bean common mosaic virus (BCMV) was investigated in Populus alba var. pyramidalis in China. Symptom characteristics, physiological performance of the host, histopathology, genome sequences and vectors, and gene regulation at the transcriptional and posttranscriptional levels were analyzed and RT−qPCR (quantitative reverse transcription PCR) validation of expression was performed in our experiments. In this work, the mechanisms by which the BCMV pathogen impacts physiological performance and the molecular mechanisms of the poplar response to viral infection were reported. The results showed that BCMV infection decreased the chlorophyll content, inhibited the net photosynthesis rate (Pn) and stomatal conductance (Gs), and significantly changed chlorophyll fluorescence parameters in diseased leaves. Transcriptome analysis revealed that the expression of the majority of DEGs (differentially expressed genes) involved in the flavonoid biosynthesis pathway was promoted, but the expression of all or almost all DEGs associated with photosynthesis-antenna proteins and the photosynthesis pathway was inhibited in poplar leaves, suggesting that BCMV infection increased the accumulation of flavonoids but decreased photosynthesis in hosts. Gene set enrichment analysis (GSEA) illustrated that viral infection promoted the expression of genes involved in the defense response or plant-pathogen interaction. MicroRNA-seq analysis illustrated that 10 miRNA families were upregulated while 6 families were downregulated in diseased poplar leaves; moreover, miR156, the largest family with the most miRNA members and target genes, was only differentially upregulated in long-period disease (LD) poplar leaves. Integrated transcriptome and miRNA-seq analyses revealed 29 and 145 candidate miRNA−target gene pairs; however, only 17 and 76 pairs, accounting for 2.2% and 3.2% of all DEGs, were authentically negatively regulated in short-period disease (SD) and LD leaves, respectively. Interestingly, 4 miR156/SPL (squamosa promoter-binding-like protein) miRNA−target gene pairs were identified in LD leaves: the miR156 molecules were upregulated, but SPL genes were downregulated. In conclusion, BCMV infection significantly changed transcriptional and posttranscriptional gene expression in poplar leaves, inhibited photosynthesis, increased the accumulation of flavonoids, induced systematic mosaic symptoms, and decreased physiological performance in diseased poplar leaves. This study elucidated the fine-tuned regulation of poplar gene expression by BCMV; moreover, the results also suggested that miR156/SPL modules played important roles in the virus response and development of viral systematic symptoms in plant virus disease.

近日，我国针对毛白杨（Populus alba var. pyramidalis）中由菜豆普通花叶病毒（Bean Common Mosaic Virus, BCMV）引发的新型杨树花叶病开展了相关研究。本实验对该病害的症状特征、宿主生理表现、组织病理学、基因组序列与传毒媒介，以及转录和转录后水平的基因调控情况进行了分析，并通过定量反转录PCR（quantitative reverse transcription PCR, RT-qPCR）完成了基因表达验证。本研究报道了BCMV病原影响宿主生理表现的相关机制，以及杨树抵御病毒侵染的分子调控机制。研究结果显示，BCMV侵染会降低病叶的叶绿素含量，抑制净光合速率（net photosynthesis rate, Pn）与气孔导度（stomatal conductance, Gs），并显著改变叶绿素荧光参数。转录组分析结果表明，杨树叶片中参与类黄酮生物合成通路的绝大多数差异表达基因（differentially expressed genes, DEGs）的表达被上调，而与光合天线蛋白及光合作用通路相关的全部或几乎全部DEGs的表达均被下调，这提示BCMV侵染会增加宿主类黄酮的积累，同时降低宿主的光合能力。基因集富集分析（Gene Set Enrichment Analysis, GSEA）显示，病毒侵染会促进与防御反应或植物-病原互作相关的基因表达。小RNA测序（microRNA-seq）分析结果表明，病叶中有10个miRNA家族表达上调，6个家族表达下调；此外，拥有最多miRNA成员与靶基因的最大家族miR156，仅在长期病害（long-period disease, LD）染病杨树叶片中呈现差异上调表达。整合转录组与microRNA-seq分析共筛选得到29对和145对候选miRNA-靶基因组合；然而在短期病害（short-period disease, SD）与长期病害（LD）染病叶片中，仅分别有17对和76对组合（占全部DEGs的2.2%与3.2%）呈现真实的负调控关系。有趣的是，在LD染病叶片中鉴定得到4对miR156/SPL（squamosa promoter-binding-like protein）-靶基因组合：miR156分子表达上调，而SPL基因表达下调。综上，BCMV侵染会显著改变杨树叶片的转录及转录后基因表达模式，抑制光合过程、增加类黄酮积累、诱导系统性花叶症状，并降低病株的生理表现。本研究阐明了BCMV对杨树基因表达的精细调控机制；此外，研究结果还提示miR156/SPL模块在植物病毒病的病毒响应及系统性病害症状发育过程中发挥了重要作用。