ZmFKF1b antagonizes the bifunctional repressor ZmEREB214 to coordinate drought tolerance and flowering time in maize

The coordination of drought tolerance and flowering time represents a fundamental evolutionary trade-off constraining crop yield potential, yet the underlying molecular mechanisms in maize remain poorly understood.

This study aims to elucidate the upstream transcriptional regulatory mechanisms governing ZmFKF1b expression and identify downstream effectors through which ZmFKF1b coordinately regulates drought stress responses and flowering time in maize.

DAP-seq and RNA-seq were employed to identify downstream target genes. Yeast one-hybrid (Y1H), dual-luciferase reporter, and EMSA validated transcription factor-promoter interactions; Y2H, BiFC, and Co-IP confirmed protein-protein interactions. Transgenic knockout and overexpression lines were generated in maize inbred line B104 via Agrobacterium-mediated transformation.

ZmDOF29 enhances drought tolerance and delays maize flowering by directly repressing ZmFKF1b expression. Under long-day (LD) conditions, ZmDOF29 constitutively delays flowering; under short-day (SD) conditions, this repressive function is conditionally activated by drought to antagonize drought-induced flowering acceleration. ZmFKF1b physically interacts with ZmEREB214, a positive regulator of drought tolerance, and attenuates its transcriptional repressor activity. Specifically, ZmEREB214 directly binds to the promoters of ZmSINAT4 (a negative regulator of drought tolerance) and ZmMADS1 (a positive regulator of flowering), repressing their expression to enhance drought tolerance and delay flowering, respectively. Physiologically, elevated ZmEREB214 or ZmDOF29 activity enhances antioxidant enzyme activities (SOD, POD, CAT), reduces MDA accumulation, and increases proline and chlorophyll content under drought. Conversely, ZmFKF1b accumulation attenuates ZmEREB214-mediated repression, thereby reducing drought tolerance and accelerating flowering.

This study elucidates a hierarchical ZmDOF29-ZmFKF1b-ZmEREB214 regulatory cascade governing the drought tolerance-flowering time trade-off in maize. ZmEREB214 serves as a bifunctional hub that directly represses ZmSINAT4 (drought) and ZmMADS1 (flowering), while ZmFKF1b antagonizes this activity through protein interaction. These findings deepen our mechanistic understanding of how crops coordinate stress acclimation with reproductive development, and provide precise molecular targets for breeding climate-resilient, high-yielding maize varieties.