A light-responsive transcriptional network involving HY5 and PIF3 negatively regulates MYB3 expression.

R2R3-MYB transcription factors (TFs) orchestrate the regulation of numerous plant metabolic and developmental pathways by modulating gene expression through promoter binding. Within the phenylpropanoid pathway, specific MYB TFs control the biosynthesis of key secondary metabolites such as flavonols, anthocyanins, and lignins. MYB3, a subgroup 4 R2R3-MYB TF, has been implicated in secondary cell wall biosynthesis, abiotic stress responses, and other physiological processes. However, its upstream regulatory mechanisms remain poorly understood. Here, we demonstrate that MYB3 expression is significantly downregulated under light conditions compared to darkness, suggesting a negative regulatory role of light. In silico promoter analysis of MYB3 identified multiple light-responsive elements (LREs), including binding motifs for the light-regulated TFs HY5 and PIF3. Binding of HY5 and PIF3 to distinct regions of the MYB3 promoter was confirmed through yeast one-hybrid (Y1H) and chromatin immunoprecipitation (ChIP) assays. Furthermore, quantitative expression analyses in transgenic and mutant lines revealed that both HY5 and PIF3 act as transcriptional repressors of MYB3. Collectively, our findings highlight a previously uncharacterized light-mediated transcriptional cascade involving HY5 and PIF3 as upstream regulators of MYB3, contributing to the broader understanding of light-regulated secondary metabolism in plants.