MiMYB1 and MibHLH2 play key roles in anthocyanin biosynthesis in Matthiola incana flowers. We established a transient expression system using Turnip mosaic virus vector in M. incana. Garden stock (Matthiola incana (L.) R. Br.) is a popular flowering plant observed from winter to spring in Japan. Here we observed that anthocyanin accumulation in ‘Vintage Lavender’ increased with flower development, whereas flavonol accumulation remained constant throughout flower development. We obtained five transcription factor genes, MiMYB1, MibHLH1, MibHLH2, MiWDR1, and MiWDR2, from M. incana floral cDNA contigs. Yeast two-hybrid analyses revealed that MiMYB1 interacted with MibHLH1, MibHLH2, and MiWDR1, but MiWDR2 did not interact with any transcription factor. Expression levels of MiMYB1 and MibHLH2 increased in petals during floral bud development. Their expression profiles correlated well with the temporal profiles of MiF3ʹH, MiDFR, MiANS, and Mi3GT transcripts and anthocyanin accumulation profile. On the other hand, MibHLH1 was expressed weakly in all organs of ‘Vintage Lavender’. However, high expression levels of MibHLH1 were detected in petals of other cultivars with higher levels of anthocyanin accumulation than ‘Vintage Lavender’. MiWDR1 and MiWDR2 maintained constant expression levels in petals during flower development and vegetative organs. Transient MiMYB1 expression in 1-month-old M. incana seedlings using a Turnip mosaic virus vector activated transcription of the endogenous anthocyanin biosynthetic genes MiF3ʹH, MiDFR, and MiANS and induced ectopic anthocyanin accumulation in leaves. Therefore, MiMYB1 possibly interacts with MibHLH2 and MiWDR1, and this trimeric protein complex activates the transcription of anthocyanin biosynthetic genes in M. incana flowers. Moreover, MibHLH1 acts as an enhancer of anthocyanin biosynthesis with the MiMYB1–MibHLH2–MiWDR1 complex. This study revealed the molecular mechanism involved in the regulation of anthocyanin accumulation levels in M. incana flowers.

中文翻译：

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与紫罗兰花色相关的花青素调控和结构基因


MiMYB1 和 MibHLH2 在 Matthiola incana 花的花青素生物合成中发挥关键作用。我们利用芜菁花叶病毒载体在 M. incana 中建立了瞬时表达系统。园林砧木（Matthiola incana (L.) R. Br.）是日本冬季至春季流行的开花植物。在这里，我们观察到“复古薰衣草”中花青素的积累随着花朵的发育而增加，而黄酮醇的积累在整个花朵的发育过程中保持恒定。我们从 M. incana 花 cDNA 重叠群中获得了五个转录因子基因：MiMYB1、MibHLH1、MibHLH2、MiWDR1 和 MiWDR2。酵母双杂交分析表明，MiMYB1 与 MibHLH1、MibHLH2 和 MiWDR1 相互作用，但 MiWDR2 不与任何转录因子相互作用。在花芽发育过程中，花瓣中 MiMYB1 和 MibHLH2 的表达水平增加。它们的表达谱与 MiF3ʹH、MiDFR、MiANS 和 Mi3GT 转录本的时间谱以及花青素积累谱密切相关。另一方面，MibHLH1在‘Vintage Lavender’的所有器官中均弱表达。然而，在花青素积累水平高于“复古薰衣草”的其他品种的花瓣中检测到了 MibHLH1 的高表达水平。 MiWDR1 和 MiWDR2 在花朵发育和营养器官期间在花瓣中保持恒定的表达水平。使用芜菁花叶病毒载体在 1 个月大的 M. incana 幼苗中瞬时表达 MiMYB1，激活内源花青素生物合成基因 MiF3ʹH、MiDFR 和 MiANS 的转录，并诱导叶片中异位花青素积累。因此，MiMYB1可能与MibHLH2和MiWDR1相互作用，并且这种三聚体蛋白复合物激活M. incana花中花青素生物合成基因的转录。 此外，MibHLH1 与 MiMYB1-MibHLH2-MiWDR1 复合物一起充当花青素生物合成的增强剂。这项研究揭示了M. incana 花中花青素积累水平调节的分子机制。