Carotenoid Cleavage Dioxygenase 1 Catalyzes Lutein Degradation To Influence Carotenoid Accumulation and Color Development in Foxtail Millet Grains

Foxtail millet is a minor but economically important crop in certain regions of the world. Millet color is often used to judge grain quality, yet the molecular determinants of millet coloration remain unclear. Here, we explored the relationship between SiCCD1 and millet coloration in yellow and white millet varieties. Carotenoid levels declined with grain maturation and were negatively correlated with SiCCD1 expression, which was significantly higher in white millet as compared to yellow millet during the color development stage. Cloning of the SiCCD1 promoter and CDS sequences from these different millet varieties revealed the presence of two additional cis-regulatory elements within the SiCCD1 promoter in white millet varieties, including an enhancer-like GC motif element associated with anoxic specific inducibility and a GCN4-motif element associated with endosperm expression. Dual-luciferase reporter assays confirmed that SiCCD1 promoter fragments containing these additional cis-acting elements derived from white millet varieties were significantly more active than those from yellow millet varieties, consistent with the observed SiCCD1 expression patterns. Further in vitro enzyme detection assays confirmed that SiCCD1 primarily targets and degrades lutein. Together, these data suggest that SiCCD1 promoter variation was a key factor associated with the observed differences in SiCCD1 expression, which in turn led to the difference in millet coloration.