ZmNAC20 positively regulates drought tolerance in maize seedlings via a chromatin-associated network.

The potential yield and establishment of maize are severely restricted by early developmental drought. Although NAC transcription factors are known to control abiotic stress responses, the role of NAC transcription factor 20 (ZmNAC20) in maize drought adaptation remains unclear. We characterized the function of ZmNAC20 in maize seedlings using CRISPR/Cas9 mutagenesis combined with molecular, physiological, and multi-omics approaches. Loss-of-function zmnac20 mutants displayed increased sensitivity to drought stress, indicating that ZmNAC20 positively regulates drought tolerance in maize seedlings. Further analyses revealed that NAC transcription factor 110 (ZmNAC110) functions upstream of ZmNAC20, activating its transcription and physically interacting with the ZmNAC20 protein. CUT&Tag profiling indicated that ZmNAC20 is associated with alterations in H3K27me3 distribution across active genomic regions. By integrating CUT&Tag-seq, AI-based protein-protein interaction predictions, and RNA-seq datasets, we identified candidate downstream components associated with ZmNAC20. Among these, YIGE1 was the most strongly supported candidate. ZmNAC20 activated YIGE1 promoter activity and interacted with the YIGE1 protein. Together, these results identify ZmNAC20 as a positive regulator of drought tolerance in maize seedlings and identify a candidate ZmNAC110-ZmNAC20-YIGE1 regulatory module linked to chromatin-associated drought responses. Because the genetic requirement for YIGE1 was not tested, YIGE1 is presented here as a strong candidate downstream component rather than a confirmed one.