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July 1st, 2025
Version: 2
Indian Institute of Science
plant biology
biorxiv

MIR319C activation by CUP-SHAPED COTYLEDON2 delays cell proliferation-to-differentiation transition in Arabidopsis leaf primordia

Alanga, N. S.Open in Google Scholar•Gupta, A.Open in Google Scholar•Mane, V.Open in Google Scholar•Roy, S.Open in Google Scholar•Jayanarayan Perumadaraya, A.Open in Google Scholar•V, A.Open in Google Scholar•Hamant, O.Open in Google Scholar•Nath, U.Open in Google Scholar

The microRNA miR319 regulates leaf size in diverse plant species by reducing the level of the target transcripts that encode JAW-TCPs, the transcription factors (TF) that restrict leaf size by committing the proliferating pavement cells to differentiation. MIR319C, one of the three miR319-producing genes in Arabidopsis, is expressed throughout the incipient leaf primordia, and its expression domain gets restricted to the base at later stages, partly due to its transcriptional repression by JAW-TCPs. However, the factors that activate and maintain MIR319C expression in leaf primordia are yet unknown. Here, we identify the CUP-SHAPED COTYLEDON2 (CUC2) TF as a direct activator of MIR319C transcription. Using a yeast one-hybrid (Y1H) screen, we identified several NAC domain TFs as potential regulators of MIR319C. Subsequent ex vivo binding and transactivation assays suggested that CUC2 binds to a distal promoter region of the MIR319C locus and activates its transcription. Mutants with compromised CUC2 and MIR319C activities resulted in smaller leaves with fewer cells. Detailed morphometric analysis of higher order CUC2 and MIR319 loss-of-function mutants highlighted the crucial role of the CUC2-MIR319 module in maintaining the duration of cell proliferation in leaf primordia. Additionally, the phenotype of mutants with altered CUC2 and MIR319/JAW-TCP activities demonstrated that CUC2 enhances leaf size through the MIR319C-JAW-TCP pathway. Overall, our findings uncovered a novel role for CUC2 in sustaining cell division by activating MIR319C transcription in the leaf primordia.

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