Flavanones and flavones are excellent source of bioactive compounds but the molecular basis of their highly efficient production remains elusive. Chalcone isomerase (CHI) family proteins play essential roles in flavonoid biosynthesis but little are known about the transcription factors controlling their gene expression. Here, we identified a type IV CHI (designated as CitCHIL1) from citrus which enhances the accumulation of citrus flavanones and flavones (CFLs). CitCHIL1 participates in a CFL biosynthetic metabolon and assists the cyclization of naringenin chalcone to (2S)‐naringenin, which leads to the efficient influx of substrates to chalcone synthase (CHS) and improves the catalytic efficiency of CHS. Overexpressing CitCHIL1 in Citrus and Arabidopsis significantly increased flavonoid content and RNA interference‐induced silencing of CitCHIL1 in citrus led to a 43% reduction in CFL content. Three AP2/ERF transcription factors were identified as positive regulators of the CitCHIL1 expression. Of these, two dehydration‐responsive element binding (DREB) proteins, CitERF32 and CitERF33, activated the transcription by directly binding to the CGCCGC motif in the promoter, while CitRAV1 (RAV: related to ABI3/VP1) formed a transcription complex with CitERF33 that strongly enhanced the activation efficiency and flavonoid accumulation. These results not only illustrate the specific function that CitCHIL1 executes in CFL biosynthesis but also reveal a new DREB‐RAV transcriptional complex regulating flavonoid production.

中文翻译：

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三个 AP2/ERF 家族成员通过调节柑橘中 IV 型查尔酮异构酶调节类黄酮合成

黄烷酮和黄酮是生物活性化合物的极好来源，但其高效生产的分子基础仍然难以捉摸。查尔酮异构酶 (CHI) 家族蛋白在类黄酮生物合成中起着重要作用，但对控制其基因表达的转录因子知之甚少。在这里，我们从柑橘中鉴定出一种 IV 型 CHI（指定为 CitCHIL1），它可以增强柑橘黄烷酮和黄酮 (CFL) 的积累。CitCHIL1 参与 CFL 生物合成代谢并协助柚皮素查耳酮环化为 ( 2S )-柚皮素，从而导致底物有效流入查耳酮合酶 (CHS) 并提高 CHS 的催化效率。在柑橘中过度表达CitCHIL1和拟南芥显着增加类黄酮含量，RNA 干扰诱导的柑橘中 CitCHIL1 沉默导致 CFL 含量降低 43%。三个 AP2/ERF 转录因子被鉴定为CitCHIL1表达的正调节因子。其中，两种脱水反应元件结合 (DREB) 蛋白 CitERF32 和 CitERF33 通过直接结合启动子中的 CGCCGC 基序激活转录，而 CitRAV1（RAV：与 ABI3/VP1 相关）与 CitERF33 形成转录复合物，大大提高了活化效率和类黄酮的积累。这些结果不仅说明了 CitCHIL1 在 CFL 生物合成中执行的特定功能，而且揭示了一种新的 DREB-RAV 转录复合物调节类黄酮的产生。