MdMYB93 activates MdHCT6 expression via transcriptional regulation to enhance chlorogenic acid biosynthesis in apples

Abstract

Chlorogenic acid (CGA), a potent antioxidant with antimicrobial, antiviral, and metabolic regulatory properties, plays multifunctional roles in apple fruit by enhancing postharvest quality, extending shelf life through oxidative stress reduction, and inhibiting enzymatic browning to preserve color, flavor, and nutritional integrity. Despite the established role of hydroxycinnamoyl-CoA transferase (HCT) as a rate-limiting enzyme in CGA biosynthesis, the specific HCT gene responsible for this process and its regulatory mechanisms remain elusive. To address this knowledge gap, we systematically investigated CGA accumulation dynamics during apple storage and functionally characterized MdHCT6, a candidate gene within the HCT family. We found that the chlorogenic acid content in apple fruit increased significantly during postharvest storage compared with the initial storage. Transcriptome analysis showed that the expression level of MdHCT6 was significantly higher than that of other HCT homologues, which was consistent with the reverse transcription quantitative PCR (RT-qPCR) results. In vitro enzymatic assays demonstrated that MdHCT6 catalyzes the synthesis of chlorogenic acid using shikimic acid and quinic acid as precursors, while genetic evidence confirmed its role as a key positive regulator of chlorogenic acid accumulation in apples. Furthermore, we identified the transcription factor MdMYB93 as a direct upstream activator of MdHCT6, establishing a regulatory cascade that governs CGA production. This work not only deciphers the molecular hierarchy of CGA biosynthesis in apples but also provides actionable targets for genetic improvement of antioxidant capacity and postharvest resilience in apple germplasm.