Zhejiang Key Laboratory of Crop Germplasm, Institute of Modern Seed Industry, Zhejiang University, Hangzhou 310058, China.
Shandong (Linyi) Institute of Modern Agriculture, Zhejiang University, Linyi 276000, China.
Int J Mol Sci. 2022 Jun 7;23(12):6357. doi: 10.3390/ijms23126357.
The endosperm is the main nutrient source in cereals for humans, as it is a highly specialized storage organ for starch, lipids, and proteins, and plays an essential role in seed growth and development. Active DNA demethylation regulates plant developmental processes and is ensured by cytosine methylation (5-meC) DNA glycosylase enzymes. To find out the role of in seed development, the null mutant of was generated using the CRISPR/Cas9 system. The null mutant of was stable and heritable, which affects the major agronomic traits, particularly in rice seeds. The null mutant of showed longer and narrower grains, and seeds were deformed containing an underdeveloped and less-starch-producing endosperm with slightly irregularly shaped embryos. In contrast to the transparent grains of the wild type, the grains of the null mutant of were slightly opaque and rounded starch granules, with uneven shapes, sizes, and surfaces. A total of 723 differential expression genes (DEGs) were detected in the null mutant of by RNA-Seq, of which 290 were downregulated and 433 were upregulated. The gene ontology (GO) terms with the top 20 enrichment factors were visualized for cellular components, biological processes, and molecular functions. The key genes that are enriched for these GO terms include starch synthesis genes ( and ) and cellulose synthesis genes (, , , and ). Genes encoding polysaccharides and glutelin were found to be downregulated in the mutant endosperm. The glutelins were further verified by SDS-PAGE, suggesting that glutelin genes could be involved in the null mutant of seed phenotype and could have the key role in the regulation of glutelins. Furthermore, 378 differentially alternative splicing (AS) genes were identified in the null mutant of , suggesting that the gene has an impact on AS events. Our findings indicated that the function on rice endosperm development in the null mutant of could be influenced through regulating gene expression and AS, which could provide the base to properly understand the molecular mechanism related to the gene in the regulation of rice seed development.
胚乳是人类谷物的主要营养来源，因为它是淀粉、脂质和蛋白质的高度特化储存器官，在种子生长和发育中起着至关重要的作用。活性 DNA 去甲基化调节植物发育过程，并由胞嘧啶甲基化 (5-meC) DNA 糖苷酶确保。为了研究 在种子发育中的作用，使用 CRISPR/Cas9 系统生成了 的 null 突变体。 的 null 突变体是稳定且可遗传的，这会影响主要的农艺性状，特别是在水稻种子中。 的 null 突变体表现出更长和更窄的谷物，并且种子变形，含有发育不良且淀粉产量较低的胚乳，胚乳形态不规则。与野生型的透明谷物相比， 的 null 突变体的谷物略带不透明且圆形的淀粉颗粒，形状、大小和表面不均匀。通过 RNA-Seq 在 的 null 突变体中检测到 723 个差异表达基因 (DEG)，其中 290 个下调，433 个上调。可视化了具有前 20 个富集因子的基因本体 (GO) 术语，用于细胞成分、生物过程和分子功能。这些 GO 术语中富集的关键基因包括淀粉合成基因（ 和 ）和纤维素合成基因（ 、 、 、和 ）。在突变体胚乳中发现编码多糖和谷蛋白的基因下调。谷蛋白通过 SDS-PAGE 进一步验证，表明谷蛋白基因可能参与 的种子表型，而 可能在调节谷蛋白方面发挥关键作用。此外，在 的 null 突变体中鉴定出 378 个差异剪接 (AS) 基因，表明 基因对 AS 事件有影响。我们的研究结果表明， 在 的 null 突变体中对水稻胚乳发育的功能可能通过调节基因表达和 AS 受到影响，这为正确理解 基因在调节水稻种子发育中的分子机制提供了基础。
