A calmodulin-like protein CML41 mediates differential cold tolerance between 'Dongzao' and its autotetraploid 'Riguang' in an ICE-CBF-dependent manner.

Ziziphus jujuba Mill. 'Dongzao' ('Dongzao') is the most widely planted fresh jujube cultivar in China. Ziziphus jujuba Mill. 'Riguang' ('Riguang'), obtained through colchicine-induced mutagenesis, exhibits superior fruit size and quality compared to 'Dongzao', but its significantly reduced cold tolerance limits large-scale cultivation. To elucidate the molecular mechanisms underlying the differential cold tolerance between 'Dongzao' and 'Riguang', we examined the expression levels of ZjCML41, ZjCBF1, ZjCBF3, and ZjICE2, which were significantly upregulated in 'Dongzao' relative to 'Riguang'. Overexpression of ZjCML41 in Arabidopsis and jujube increased cold tolerance, whereas transient silencing of ZjCML41 in jujube decreased cold tolerance. Additionally, ZjbZIP25 could interact with the promoter region of ZjCML41 and positively regulate its expression, thereby enhancing cold tolerance. Moreover, ZjCML41 interacted with ZjIQD15, which in turn positively influenced cold tolerance by modulating the ICE-CBF pathway and reactive oxygen species scavenging activity. The proposed regulatory module comprising ZjbZIP25, ZjCML41, and ZjIQD15 enhances cold tolerance in 'Dongzao' by activating the ICE-CBF pathway, while downregulation of these genes reduces cold tolerance in 'Riguang'. These findings provide valuable theoretical insights and potential practical strategies for improving and regulating cold tolerance in 'Riguang'.