ZmNAC16 increases drought tolerance in maize seedlings via ABA-mediated stomatal closure

Stomatal movement is a crucial response of plants to drought stress. Several NAC transcription factors have been implicated in drought tolerance in maize seedlings, whereas the molecular mechanism by which they directly regulate ABA signaling components within the context of the four-cell stomatal complex remains unknown. In this study, we characterized ZmNAC16 in regulating drought resistance and stomatal closure in maize. Through phenotypic analysis of transgenic overexpression and gene-edited mutant lines, we assessed ZmNAC16 function in stomatal dynamics and drought tolerance. Overexpression of ZmNAC16 in transgenic maize resulted in increased leaf temperature, reduced stomatal conductance, decreased water loss, and enhanced drought tolerance, whereas gene-edited mutants exhibited opposite phenotypes. ZmNAC16 localized to the nucleus and acted as a transcriptional activator. Disruption of ZmNAC16 significantly downregulated key ABA signaling components, including ZmCIPK3 and ZmPP2C81, whose promoters were directly bound by ZmNAC16. Consistently, ZmNAC16 knockout mutants showed impaired ABA-induced stomatal closure, while overexpression lines displayed heightened stomatal sensitivity to ABA. Collectively, our findings demonstrate that ZmNAC16 integrates ABA signaling to regulate stomatal closure and drought tolerance in maize, positioning it as a promising target for enhancing crop resilience.