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Cold-Induced Promoter Methylation Attenuates ScMYB7-Mediated Repression of the CBF Pathway: A Proposed Mechanism for Sugarcane Cold Adaptation

Weiwei Liu,Qiuyue Ou,Meichang Feng,Yue Li,Linghan Huang,Xuan Peng,Xiaoqiang Zhong,Guoqiang Huang,Hengbo Wang,Jinlong Guo

Cold stress is a critical environmental factor adversely affecting plant growth and development. As a tropical-origin crop constituting the primary global source of sucrose, sugarcane ( Saccharum spp. hybrid) exhibits particular vulnerability to suboptimal temperature conditions, with chilling injury substantially compromising its yield potential. Despite its agricultural significance, the molecular mechanisms underlying cold acclimation in sugarcane remain poorly characterized. Here, we report a cold-repressed 1R-MYB gene, ScMYB7 , from sugarcane, whose promoter ( pro-ScMYB7 ) contains multiple cis -acting elements, including two cytosine—phosphate diester—guanine (CpG) islands. Bisulfite sequencing PCR (BSP) and qPCR results showed that low-temperature treatment increased the methylation level of the CpG islands in the promoter to reduce the transcription of the ScMYB7 gene. The outcomes of GUS enzyme activity measurement of the promoter also indicated that low-temperature treatment inhibits the promoter's transcriptional activity, and methylation inhibitors could alleviate this inhibition. By generating transgenic Arabidopsis lines overexpressing ScMYB7 , ScMYB7's roles in regulating cold tolerance were investigated. We observed that the transgenic plants reduced cold tolerance, featured by a decreased survival rate after recovery, fluctuated physiological traits, and significantly lower expression levels of the C-repeat binding factor (CBF)-dependent pathway genes ( AtCBFs , AtCOR15 , and AtRD29A ). Yeast one-hybrid assays demonstrated direct binding of ScMYB7 to the AtCBF1 promoter, while repression of sugarcane ScDREB1A occurred indirectly. Furthermore, the dual-luciferase reporter assay indicated that ScMYB7 was able to inhibit the expression of the AtCBF1 or ScDREB1A . Taken together, we propose a model in which ScMYB7 acts as a repressor of cold tolerance via the CBF-dependent pathway. Under low-temperature stress, increased methylation of the pro-ScMYB7 promoter reduces ScMYB7 expression, thereby alleviating its repression of sugarcane DREB/CBF-type transcription factors and enhancing cold adaptation.