Transcription factor WRKY53 transcriptionally activates sensitive to proton rhizotoxicity 1 to promote C-repeat binding factors 1 and 3 mediated chilling tolerance in apple.

Chilling stress limits apple (Malus domestica) growth and productivity, particularly in spring, when chilling events can severely impair bud break and early shoot development. However, the regulatory mechanisms underlying chilling adaptation in apple remain incompletely understood. Here, we identified the C2H2 zinc-finger transcription factor Sensitive to Proton Rhizotoxicity 1 (MdSTOP1) from apple as a chilling-responsive regulator whose transcript and protein abundance increased under chilling stress. We further demonstrated that MdSTOP1 enhances apple tolerance to prolonged chilling at non-freezing temperatures, accompanied by sustained photosynthetic capacity, improved osmotic adjustment, and increased reactive oxygen species (ROS)-scavenging capacity. Chromatin immunoprecipitation sequencing (ChIP-seq) and subsequent molecular assays identified C-repeat binding factor 1 (MdCBF1) and C-repeat binding factor 3 (MdCBF3) as direct targets transcriptionally activated by MdSTOP1. In addition, apple WRKY transcription factor 53 (MdWRKY53) was identified as an upstream regulator of MdSTOP1, was induced by chilling stress, and directly activated MdSTOP1 transcription, thereby enhancing chilling tolerance in transgenic apple lines. Together, these findings establish an MdWRKY53-MdSTOP1-MdCBF1/3 regulatory module that promotes chilling tolerance in apple.