The reason for kale reddening in late autumn is BoCBF2/BoMYB2 synergistically regulates <i>BoDFR</i> expression

Ornamental kale is a popular decorative plant, particularly during the cold season. Its newborn leaves turn red due to the accumulation of anthocyanins under low temperature, but the response mechanism to low temperature is still unclear. Here, an anthocyanin synthesis gene, BoDFR, was identified as playing a role in the anthocyanin synthesis pathway via a map-based cloning strategy, and silencing of BoDFR resulted in reduced anthocyanin accumulation. Through adopting RNA-seq and cis-acting element methods, we speculated that BoMYB2 was the upstream transcription factor of BoDFR. Overexpression of BoMYB2 increased anthocyanin accumulation in kale and the expression level of BoDFR. We demonstrated that BoMYB2 binds to the promoter of BoDFR and activates its expression, as evidenced by yeast one-hybrid, electrophoretic mobility shift, GUS activity, and luciferase reporter assays. Further investigations revealed that the cold stress response factor BoCBF2 interacts with the transcription factor BoMYB2 to form a transcriptional regulatory module, BoCBF2/BoMYB2, which synergistically regulates the transcript levels of BoDFR under low temperature. Our findings uncovered connections between the BoCBF2-mediated low-temperature signal, BoMYB2-mediated transcription regulation, and BoDFR-mediated anthocyanin synthesis pathway in the reddening of newborn leaves in kale. Understanding the molecular mechanism behind reddening helps reveal the regular patterns of plant color changes in response to low temperature, providing theoretical support for enhancing ornamental leaf colors.