miR156b-targeted <i>VvSBP8/13</i> functions downstream of the abscisic acid signal to regulate anthocyanins biosynthesis in grapevine fruit under drought

Abstract

Anthocyanins are the primary color components of grapevine berries and wines. In cultivation practices, a moderate water deficit can promote anthocyanin accumulation in red grape skins. Our previous study showed that abscisic acid (ABA) plays a key role in this process. Herein, we identified a microRNA, vv-miR156b, that is generated in grapevine berries in response to drought stress, along with increasing anthocyanin content and biosynthetic structural gene transcripts. In contrast, vv-miR156b short tandem target mimic (STTM) function-loss callus exhibits the opposite phenotype. Results from <i>in vivo</i> and <i>in vitro</i> experiments revealed that the ABA-signaling-regulated transcription factor VvAREB2 binds directly to the ABA-responsive element (ABRE) of the <i>MIR156b</i> promoter and activates miR156b expression. Furthermore, two miR156b downstream targets, <i>VvSBP8</i> and <i>VvSBP13</i>, exhibited reduced grape anthocyanin content in their overexpressors but there was a contrary result in their CRISPR-edited lines, the decrease in anthocyanin content was rescued in miR156b and SBP8/13 double overexpressors. We further demonstrated that both <i>VvSBP8</i> and <i>VvSBP13</i>, encoding transcriptional repressors, displayed sufficient ability to interact with VvMYC1 and VvMYBA1, thereby interfering with MYB-bHLH-WD (MBW) repeat transcriptional complex formation, resulting in the repression of anthocyanin biosynthesis. Our findings demonstrate a direct functional relationship between ABA signaling and the miR156-SBP-MBW complex regulatory module in driving drought-induced anthocyanin accumulation in grape berries.