BolANT3 Positively Regulates Indolic Glucosinolate Accumulation by Transcriptionally Activating BolCYP83B1 in Cabbage

Indolic glucosinolates are a group of plant secondary metabolites found in <i>Brassica</i> vegetables, and their breakdown products could act as important anti-cancer and defense compounds against biotic stresses. Transcriptional regulation plays a key role in modulating the biosynthesis of indolic glucosinolates in the model plant <i>Arabidopsis</i>, but little is known about the transcriptional regulatory landscape of these glucosinolates in <i>Brassica</i> vegetables. In this study, we selected and functionally validated the important biosynthetic gene <i>BolCYP83B1</i> from the indolic glucosinolate pathway in cabbage. Through a yeast one-hybrid assay, we systemically screened and identified upstream regulators of <i>BolCYP83B1</i> in cabbage with BolANTs as the top candidates for further functional validation. Two homologs of BolANTs, BolANT1 and <i>BolANT3</i>, were confirmed to bind the promoter of <i>BolCYP83B1</i> via both a yeast one-hybrid assay and an LUC assay. The overexpression of <i>BolANT3</i> in cabbage significantly increased the accumulation of indolic glucosinolates, while the virus-induced gene silencing (VIGS) of <i>BolANT3</i> significantly reduced the accumulation of indolic glucosinolates in cabbage. Our work provides valuable insights into the transcriptional regulatory mechanisms of indolic glucosinolates in <i>Brassica</i> vegetables.