アブラナ科植物の受粉シグナル分子の解明

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After forward genetic screen was performed, I isolated mutant lines with defect after pollination process. In this year, I identified the causal gene underlying the mutant phenotype using genome deep sequencing. T-DNA insertion plant line of the identified gene showed similar phenotype as the one isolated from genetic screen suggesting that the identified gene was important for fertilization of Arabidopsis. Bioinformatic analysis revealed that this gene is a putative homolog of Saccharomyces cerevisiae Vacuolar Protein-Sorting associated 13 (ScVPS13), so, the candidate gene was named AtVPS13 in this study. Pollen grain development of vps13 was confirmed to be normal since its mature pollen was in tricellular stage. So, I observed in vivo hydration and germination of the mutant pollen on the stigma. I found that vps13 mutation did not affect hydration of the pollen however, the germination was severely compromised (no mutant pollen germinated within 45 min after pollination, but 94% of wild type pollen germinated). In vitro pollen germination assay also showed similar trend suggesting that VPS13 was important for pollen germination. As a result, the number of seed/silique of mutant plant was almost three times lower than the wild type. I also found that mutation of putative paralog AT4G17140 additional to vps13 results in a severe sterile phenotype of double mutant plant. In summary, I successfully identified novel gene important in successful fertilization of Arabidopsis found that this gene functioned after pollen hydration to support pollen germination on the stigma.