AINTEGUMENTA-LIKE 5 promotes root and leaf growth by regulating CYCLIN D6;1 in Rehmannia glutinosa

AINTEGUMENTA-LIKE (AIL) proteins, members of the APETALA2/Ethylene Responsive Factor (AP2/ERF) transcription factor family, play a crucial role in regulating plant development. To explore the functions of AILs in Rehmannia glutinosa development, we focused on RgAIL5, which is characterized by its nuclear localization and transcriptional activation activity. Overexpression of RgAIL5 in R. glutinosa significantly enhanced root fresh weight, tuberous root diameter, leaf length, and leaf width compared to wild type plants. Conversely, silencing of RgAIL5 significantly delayed tuberous root formation and reduced leaf length and width in R. glutinosa. Yeast one-hybrid, dual-luciferase, and electrophoretic mobility shift assays confirmed that RgAIL5 activates the transcription of RgCYCD6;1 by binding to its promoter. Furthermore, RgCYCD6;1 was shown to interact with RgCDKA;1 through yeast two-hybrid, luciferase complementation imaging, and pull-down assays. Overexpression of either RgCYCD6;1 or RgCDKA;1 individually in R. glutinosa plants promoted tuberous root thickening and leaf enlargement, which might be attributed to enhanced cell proliferation and cell expansion. Overall, our study elucidates the critical regulatory role of the RgAIL5-RgCYCD6;1-RgCDKA;1 module in the growth of roots and leaves in R. glutinosa.