Kumho Life Science Laboratory, Department of Molecular Biotechnology, College of Agriculture and Life Sciences, Chonnam National University, Gwangju 61166, Republic of Korea.
Department of Agricultural Biotechnology, National Institute of Agricultural Science, RDA, Jeonju 54874, Republic of Korea.
Int J Mol Sci. 2023 Nov 29;24(23):16941. doi: 10.3390/ijms242316941.
The glutelins are a family of abundant plant proteins comprised of four glutelin subfamilies (GluA, GluB, GluC, and GluD) encoded by 15 genes. In this study, expression of subsets of rice glutelins were suppressed using CRISPR-Cas9 gene-editing technology to generate three transgenic rice variant lines, , , and . Suppression of the targeted glutelin genes was confirmed by SDS-PAGE, Western blot, and q-RT-PCR. Transgenic rice variants GluA1, GluB2, and GluC1 showed reduced amylose and starch content, increased prolamine content, reduced grain weight, and irregularly shaped protein aggregates/protein bodies in mature seeds. Targeted transcriptional profiling of immature seeds was performed with a focus on genes associated with grain quality, starch content, and grain weight, and the results were analyzed using the Pearson correlation test (requiring correlation coefficient absolute value ≥ 0.7 for significance). Significantly up- or down-regulated genes were associated with gene ontology (GO) and KEGG pathway functional annotations related to RNA processing (spliceosomal RNAs, group II catalytic introns, small nucleolar RNAs, microRNAs), as well as protein translation (transfer RNA, ribosomal RNA and other ribosome and translation factors). These results suggest that rice glutelin genes may interact during seed development with genes that regulate synthesis of starch and seed storage proteins and modulate their expression via post-transcriptional and translational mechanisms.
谷蛋白是一类丰富的植物蛋白，由四个谷蛋白亚家族（GluA、GluB、GluC 和 GluD）组成，由 15 个基因编码。在这项研究中，使用 CRISPR-Cas9 基因编辑技术抑制水稻谷蛋白的亚组表达，生成了三个转基因水稻变体系、、和。通过 SDS-PAGE、Western blot 和 q-RT-PCR 证实了靶向谷蛋白基因的抑制。转基因水稻变体 GluA1、GluB2 和 GluC1 表现出直链淀粉和淀粉含量降低、脯氨酸含量增加、粒重降低以及成熟种子中不规则形状的蛋白聚集体/蛋白体。对未成熟种子进行了靶向转录谱分析，重点关注与谷物质量、淀粉含量和粒重相关的基因，并使用 Pearson 相关系数测试（要求相关系数绝对值≥0.7 才有意义）对结果进行分析。显著上调或下调的基因与基因本体论（GO）和与 RNA 加工（核酶 RNA、II 类催化内含子、小核仁 RNA、microRNA）以及蛋白翻译（转移 RNA、核糖体 RNA 和其他核糖体和翻译因子）相关的功能注释相关。这些结果表明，水稻谷蛋白基因在种子发育过程中可能与调节淀粉和种子贮藏蛋白合成的基因相互作用，并通过转录后和翻译机制调节其表达。
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