Modulation of the Vitis vinifera cv. 'Chardonnay' microRNA and mRNA transcriptomes in response to aster yellows phytoplasma-infection

[摘要] ENGLISH ABSTRACT: Aster yellows (AY) phytoplasmas are part of a group of cell wall-less plant pathogenic bacteria responsible for a detrimental disease known as grapevine yellows (GY). The molecular mechanisms of AY phytoplasma pathogenicity on highly susceptible cultivars, such as Chardonnay, are still largely unknown. This has sparked considerable interest to gain knowledge about the basis of host susceptibility to GY in order to develop control strategies that may mitigate the scale of infection or even prevent spread. Leaf total RNA was extracted from both healthy and AY-infected plants to generate small RNA (sRNA) sequencing libraries, as well as mRNA sequencing libraries. These libraries were subjected to Illumina transcriptome sequencing (small RNA-seq and mRNA-seq, respectively), and comparative transcriptome profiling, to explore the involvement of microRNA (miRNA) and gene expression pathways in AY phytoplasma-infected Chardonnay. Multiple known miRNA sequence variants (isomiRs) were identified, and 13 known miRNAs were shown to be differentially expressed. A total of 175 novel miRNA precursor sequences, each derived from a previously uncharacterised genomic location, were identified, of which 23 were differentially expressed. Some of these novel miRNAs shared high sequence similarity with conserved miRNAs from other plant species, as well as known grapevine miRNAs. The relative expression of some of these known and novel miRNAs was confirmed with stem-loop RT-qPCR analysis, thereby validating the trend of miRNA expression in the normalised sRNA-seq read count data. miRNA target prediction, using a complementary-based in silico approach, followed by functional annotation, allowed the identification of potential target genes involved in plant morphology, hormone signalling, nutrient homeostasis, as well as plant stress. mRNA-seq results showed that 175 genes were differentially expressed in the AY phytoplasma-infected leaf material. Functional annotation of differentially expressed genes (DEGs) enabled the identification of mRNAs involved in plastid and cell wall metabolism/architecture, signalling, innate immunity, pathogen defence, secondary metabolism and photosynthesis. RT-qPCR analysis was used to validate the trend of expression of significant DEGs. Taken together, this study presents the first report on the modulation of miRNAs and genes associated with AY phytoplasma-infection in Chardonnay. The knowledge generated during this study may be crucial in understanding disease symptom development in AY phytoplasma-infected grapevines. Importantly, the findings of this study may also aid in developing GY disease control strategies and could provide added insight for future plant pathogenesis-related studies.

[发布机构] Stellenbosch University