RsWRKY40 coordinates the cold stress response by integrating RsSPS1-mediated sucrose accumulation and the CBF-dependent pathway in radish (Raphanus sativus L.).

Cold stress adversely affects crop growth and development. Radish is an important root vegetable crop, and its taproot formation is susceptible to low temperatures. However, the molecular basis of the cold stress response has not yet been fully dissected in radish. Here, a sucrose phosphate synthase gene (RsSPS1) was identified through a genome-wide association study and transcriptome analysis. RsSPS1 was responsible for sucrose synthesis, and sucrose was shown to be involved in taproot growth, cambium activity, and cold tolerance in radish. RsSPS1 regulated cambium activity and cold stress response by modulating sucrose content. Moreover, RsWRKY40 was identified as the upstream transcription activator of RsSPS1 by binding to its promoter. RsWRKY40 functioned in cambium activity and cold tolerance by modulating RsSPS1-mediated sucrose accumulation. Furthermore, RsWRKY40 promoted the RsCBF1 and RsCBF2 expression levels, resulting in elevated cold resilience. RsWRKY40 also enhanced its own transcription, forming a positive auto-regulatory loop to regulate cold stress response in radish. Together, a transcription module of RsWRKY40 orchestrated cold stress response by integrating sucrose accumulation and the CBF-dependent pathway was uncovered. These findings would provide novel insight into the molecular mechanism underlying cold-responsive sucrose accumulation and cambium activity and facilitate the genetic improvement of cold tolerance in radish breeding programs.