Key Laboratory of Molecular Epigenetics of MOE and Institute of Genetics & Cytology, Northeast Normal University, Changchun, China.
Key Laboratory of Molecular Epigenetics of MOE and Institute of Genetics & Cytology, Northeast Normal University, Changchun, China; National Demonstration Center for Experimental Biology Education, Northeast Normal University, Changchun, China.
Plant Physiol Biochem. 2019 Aug;141:60-72. doi: 10.1016/j.plaphy.2019.05.015. Epub 2019 May 20.
The MBW complex, consisting of MYB, basic helix-loop-helix (bHLH) and WD40 proteins, regulates multiple traits in plants, such as anthocyanin and proanthocyanidin biosynthesis and cell fate determination. The complex has been widely identified in dicot plants, whereas few studies are concentrated on monocot plants which are of crucial importance to decipher its functional diversities among angiosperms during evolution. In present study, a WD40 gene from Freesia hybrida, designated as FhTTG1, was cloned and functionally characterized. Real-time PCR analysis indicated that it was expressed synchronously with the accumulation of both proanthocyanidins and anthocyanins in Freesia flowers. Transient protoplast transfection and biomolecular fluorescence complementation (BiFC) assays demonstrated that FhTTG1 could interact with FhbHLH proteins (FhTT8L and FhGL3L) to constitute the MBW complex. Moreover, the transportation of FhTTG1 to nucleus was found to rely on FhbHLH factors. Outstandingly, FhTTG1 could highly activate the anthocyanin or proanthocyanidin biosynthesis related gene promoters when co-transfected with MYB and bHLH partners, implying that FhTTG1 functioned as a member of MBW complex to control the anthocyanin or proanthocyanidin biosynthesis in Freesia hybrida. Further ectopic expression assays in Arabidopsis ttg1-1 showed the defective phenotypes of ttg1-1 were partially restored. Molecular biological assays validated FhTTG1 might interact with the endogenous bHLH factors to up-regulate genes responsible for anthocyanin and proanthocyanidin biosynthesis and trichome formation, indicating that FhTTG1 might perform exchangeable roles with AtTTG1. These results will not only contribute to the characterization of FhTTG1 in Freesia but also shed light on the establishment of flavonoid regulatory system in monocot plants, especially in Freesia hybrida.
MBW 复合物由 MYB、碱性螺旋-环-螺旋(bHLH)和 WD40 蛋白组成,调节植物的多种性状,如花青素和原花青素生物合成和细胞命运决定。该复合物已在双子叶植物中广泛鉴定,而关于单子叶植物的研究较少,这对于解析其在被子植物进化过程中的功能多样性至关重要。本研究从杂种百合中克隆并功能鉴定了一个 WD40 基因,命名为 FhTTG1。实时 PCR 分析表明,它与杂种百合花中原花青素和花青素的积累同步表达。瞬时原生质体转染和生物分子荧光互补(BiFC)试验表明,FhTTG1 可以与 FhbHLH 蛋白(FhTT8L 和 FhGL3L)相互作用构成 MBW 复合物。此外,发现 FhTTG1 向核内的转运依赖于 FhbHLH 因子。值得注意的是,当与 MYB 和 bHLH 伴侣共转染时,FhTTG1 可以高度激活花青素或原花青素生物合成相关基因启动子,表明 FhTTG1 作为 MBW 复合物的成员在控制杂种百合中花青素或原花青素的生物合成中起作用。进一步在拟南芥 ttg1-1 中的异位表达试验表明,ttg1-1 的缺陷表型部分得到恢复。分子生物学试验验证了 FhTTG1 可能与内源性 bHLH 因子相互作用,上调负责花青素和原花青素生物合成和毛状体形成的基因,表明 FhTTG1 可能与 AtTTG1 发挥可互换的作用。这些结果不仅有助于杂种百合中 FhTTG1 的特征描述,也为单子叶植物,特别是杂种百合中黄酮类调控系统的建立提供了启示。
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