DcbHLH1 interacts with DcMYB1 and DcMYB2 to dynamically regulate petal pigmentation in Dianthus caryophyllus

The carnation (Dianthus caryophyllus L.) is a beloved and widely embraced ornamental flower worldwide, celebrated for vibrant hues, delightful fragrance, and prolonged blooming season. Within the petals of carnation blooms, anthocyanins, natural pigments, are responsible for the diverse range of captivating colors they display. However, the intricate molecular processes orchestrating the synthesis of these anthocyanins in carnations remain unclear. This study unveils DcbHLH1 as a pivotal regulator actively participating in anthocyanin biosynthesis within carnation flowers. Y1H and dual-LUC provided evidences that DcbHLH1 exhibited direct binding to the promoters of DcCHS and DcDFR, consequently leading to the activation of their gene expressions. In the earlier stages (S2 and S3) of blooming, DcMYB1 synergistically enhances the transcriptional activation capacity of DcbHLH1 through their collaborative interaction. As the flower matures into the later stages (S4 and S5), DcMYB1 expression exhibited a rapid decline. Concurrently, the repressor gene, DcMYB2, experienced a swift upsurge in expression, ultimately inhibiting anthocyanin biosynthesis and curtailing the transcriptional activation activity of DcbHLH1 through direct interaction with DcbHLH1. Our results show a narrative of the dynamic interplay between DcbHLH1-DcMYB1 and DcbHLH1-DcMYB2, unveiling their roles in orchestrating the array of hues that adorn carnation petals over various flowering stages. These findings offer novel insights into the regulatory mechanisms influencing anthocyanin biosynthesis in ornamental plants.