| Summary: | 碩士 === 國立臺灣大學 === 植物病理與微生物學研究所 === 106 === Peanut witches’ broom phytoplasma (PnWB) infected-Catharanthus roseus plants showed leaf yellowing, witches’ broom, and phyllody (herein referred as leafy flower) symptoms. Our previous studies demonstrated that Phyllody Symptoms1 (PHYL1) of PnWB is the effector to trigger the microRNA396 (miR396) down-regulation, and subsequently up-regulate the miR396-targeted SHORT VEGETATIVE PHASE (SVP) for leafy flower formation in PnWB-infected C. roseus and transgenic Arabidopsis expressing green fluorescent protein (GFP)-PHYL1 fusion gene (GFP-PHYL1 plant). Moreover, we observed the anthocyanin accumulation in the leafy flowers and the gene expressions of anthocyanin pathway were up-regulated in 6-week-old GFP-PHYL1 plants. We speculate that PHYL1 have ability to induce anthocyanin accumulation. In addition, we found that the miR156 expression levels was up-regulated, and the gene expression levels of miR156-regulated SQUAMOSA PROMOTER BINDING PROTEIN LIKE (SPL) genes were down-regulated in the GFP-PHYL1 plant. SPL9 is a negative regulator of anthocyanin biosynthesis; therefore, the anthocyanin accumulation in leafy flowers is caused by the miR156-mediated SPL9 down-regulation. Furthermore, GAs activate the SPLs for flowering; however, SVP inhibit the GAs biosynthesis. Therefore, up-regulated SVP expression, which mediate by low miR396, represses gibberellic acids (GAs) biosynthesis in GFP-PHYL1 plants, resulting in inactivation of SPLs and consequence anthocyanin accumulation. In addition, previous study demonstrated that sucrose can induce expressions of anthocyanin biosynthetic genes. In addition, our results showed that sucrose was accumulated in leafy flower and triggered the anthocyanin biosynthesis. Moreover, our previous study indicated that PHYL1 is an unstable protein in vivo that might need the other bacterial protein(s) for stabilizing. The results of immunoprecipitation (IP) from leafy flowers of PnWB-infected C. roseus indicated that PnWB effector 2 (PnE2) directly interacts with PHYL1, whereas the PnWB effector 1 (PnE1) indirectly interacts with PHYL1 through the interaction with PnE2. According to the amino acid sequence alignment and signal peptide prediction, we found that PnE1 is a novel effector, and PnE2 is an immunodominant membrane protein which anchor on membrane of phytoplasma. In this study, we demonstrated that PHYL1-interfered miRNA expressions have indirectly effects on anthocyanin accumulation of leafy flowers and two bacterial proteins interact with PHYL1. We suggest that anthocyanin and the carbohydrate accumulate in leafy flower might benefit for phytoplasma survival.
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