Meng Lingzhi, Qi Changyan, Wang Cuihong, Wang Shuai, Zhou Chunlei, Ren Yulong, Cheng Zhijun, Zhang Xin, Guo Xiuping, Zhao Zhichao, Wang Jie, Lin Qibing, Zhu Shanshan, Wang Haiyang, Wang Zhonghua, Lei Cailin, Wan Jianmin
Institute of Crop Sciences, Chinese Academy of Agriculture Sciences/National Key Facility for Crop Gene Resources and Genetic Improvement, Beijing, 100081, China.
State Key Laboratory of Crop Stress Biology for Arid Areas, College of Agronomy, Northwest A&F University, Yangling, 712100, Shaanxi, China.
Rice (N Y). 2021 Apr 21;14(1):37. doi: 10.1186/s12284-021-00480-1.
Anthocyanins cause purple, brown or red colors in various tissues of rice plants, but the specific determinant factors and regulatory systems for anthocyanin biosynthesis in almost all tissues remain largely unknown. In the present study, we mapped and isolated two complementary genes, OsC1 encoding a R2R3-MYB transcriptional factor and OsDFR encoding a dihydroflavonol 4-reductase, which are responsible for the purple coloration of apiculi and stigmas in indica cultivar Xieqingzao by the map-based cloning strategy. We also identified two tissue-specific pigmentation genes, OsPa for apiculi and OsPs for stigmas, by phylogenetic analysis of all anthocyanin biosynthesis-associated bHLH transcriptional factors in maize and rice, CRISPR/Cas9 knockout and transcriptional expression analysis. The OsC1, OsPa and OsPs proteins are all localized in the nucleus while the OsDFR protein is localized in the nucleus and cytoplasm, and the OsC1 and OsDFR genes are preferentially strongly expressed in both purple-colored tissues while the OsPa and OsPs genes are preferentially strongly expressed in apiculi and stigmas, respectively. OsC1 specifically interacts with OsPa or OsPs to activate OsDFR and other anthocyanin biosynthesis genes, resulting in purple-colored apiculi or stigmas. OsC1 itself does not produce color but can produce brown apiculi when functioning together with OsPa. Loss of function of OsDFR alone leads to brown apiculi and straw-white stigmas. Genotyping and phenotyping of a panel of 176 rice accessions revealed diverse genotypic combinations of OsC1, OsDFR, OsPa and OsPs that enable accurate prediction of their apiculus and stigma pigmentation phenotypes, thus validating the general applicability of the OsC1-OsDFR-OsPa and OsC1-OsDFR-OsPs models to natural populations. Our findings disclosed the biological functions of OsC1, OsPa and OsPs, and shed light on the specific regulatory systems of anthocyanin biosynthesis in apiculi and stigmas, a further step in understanding the regulatory network of anthocyanin biosynthesis in rice.
花青素致使水稻植株的不同组织呈现紫色、棕色或红色，但几乎所有组织中花青素生物合成的具体决定因素和调控系统仍 largely 未知。在本研究中，我们通过图位克隆策略定位并分离出两个互补基因，即编码 R2R3-MYB 转录因子的 OsC1 和编码二氢黄酮醇 4-还原酶的 OsDFR，它们负责籼稻品种协青早颖尖和柱头的紫色着色。我们还通过对玉米和水稻中所有与花青素生物合成相关的 bHLH 转录因子进行系统发育分析、CRISPR/Cas9 敲除和转录表达分析，鉴定出两个组织特异性色素沉着基因，颖尖的 OsPa 和柱头的 OsPs。OsC1、OsPa 和 OsPs 蛋白均定位于细胞核，而 OsDFR 蛋白定位于细胞核和细胞质，并且 OsC1 和 OsDFR 基因在两个紫色组织中均优先强烈表达，而 OsPa 和 OsPs 基因分别在颖尖和柱头中优先强烈表达。OsC1 特异性地与 OsPa 或 OsPs 相互作用以激活 OsDFR 和其他花青素生物合成基因，从而导致颖尖或柱头呈现紫色。OsC1 本身不产生颜色，但与 OsPa 共同作用时可产生棕色颖尖。单独的 OsDFR 功能丧失会导致棕色颖尖和稻草白色柱头。对一组 176 份水稻种质的基因分型和表型分析揭示了 OsC1、OsDFR、OsPa 和 OsPs 的多种基因型组合，能够准确预测它们的颖尖和柱头色素沉着表型，从而验证了 OsC1-OsDFR-OsPa 和 OsC1-OsDFR-OsPs 模型对自然群体的普遍适用性。我们的研究结果揭示了 OsC1、OsPa 和 OsPs 的生物学功能，并阐明了颖尖和柱头中花青素生物合成的特定调控系统，这是进一步了解水稻花青素生物合成调控网络的重要一步。
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