Cooperative interaction between ZmKNOX13 and ZmNAC70 improves cold stress response in maize

Maize ( Zea mays L.) is a globally important crop that is highly sensitive to cold stress during the early seedling stage, which severely constrains its development and growth. However, the molecular basis of cold adaptation in maize still remains largely unclear. In this study, we identified ZmKNOX13 , a KNOX family transcription factor, as a key regulator for maize cold tolerance through genetic analysis of a cold-sensitive EMS mutant. Functional analyses using overexpression and CRISPR/Cas9 knockout lines demonstrated that ZmKNOX13 plays an important role in the maize cold stress response. We further showed that ZmKNOX13 physically interacts with ZmNAC70, and the zmnac70 mutant also exhibits a cold-sensitive phenotype. DNA affinity purification sequencing (DAP-seq) identified the cold-responsive bZIP transcription factor ZmLIP15 as a direct target of ZmKNOX13. Electrophoretic mobility shift assays (EMSAs) indicated that both ZmKNOX13 and ZmNAC70 directly bind to the ZmLIP15 promoter. Furthermore, dual luciferase reporter assay demonstrated that ZmNAC70 enhanced ZmKNOX13-mediated activation of the ZmLIP15 promoter under cold stress. Collectively, our findings uncover a novel transcriptional regulatory module that modulates maize cold stress response, and provide potential targets for the molecular improvement of cold-tolerant maize varieties