ZmWRKY104–ZmCCaMK interaction enhances brassinosteroid-promoted salt tolerance in maize ( <i>Zea mays</i> L.) via antioxidant defense

Brassinosteroid (BR)-mediated salt tolerance is a crucial mechanism for maize (Zea mays L.) adaptation to saline-alkaline environments. This study aimed to elucidate the molecular mechanism underlying BR-induced salt tolerance in maize, focusing on the regulatory roles of ZmWRKY104 and ZmCCaMK. Key results showed that ZmWRKY104 directly interacts with ZmCCaMK in the nucleus in a non-phosphorylation-dependent manner, forming a novel regulatory module. BR treatment upregulates ZmWRKY104 expression, and overexpression of ZmWRKY104 significantly enhances the activities of antioxidant enzymes (APX and SOD). Co-expression of ZmWRKY104 and ZmCCaMK synergistically promotes the antioxidant defense system in maize. Transgenic maize overexpressing ZmWRKY104 exhibits obvious salt tolerance advantages under 100 mM NaCl stress compared to wild-type plants, including reduced leaf yellowing, increased plant height and root length, as well as decreased electrolyte leakage (EL) and malondialdehyde (MDA) content. Collectively, this study identifies a novel non-phosphorylation-dependent WRKY-CCaMK regulatory module in the BR signaling pathway, which enhances BR-induced maize salt tolerance by synergistically activating antioxidant defense. The findings highlight ZmWRKY104 as a candidate gene and provide a potential molecular mechanism for salt-tolerant maize breeding in saline-alkaline regions of northern China.