RcWRKY71 - RcGolS1 module enhances cold stress tolerance by oligosaccharides accumulation in rose ( Rosa chinensis )

Roses are globally important ornamental crops worldwide and are widely used as landscape plants in outdoor greening. However, cold stress limits their growth, development, and cultivation range, especially in northern and high-altitude regions. Nevertheless, the transcriptional networks mediating cold stress responses in roses have yet to be fully understood. In this study, we identified and characterized RcWRKY71, a cold-induced WRKY transcription factor from rose (Rosa chinensis). Our findings systematically establish that RcWRKY71 is a key positive regulator of cold hardiness in roses. Overexpressing of RcWRKY71 enhanced cold resistance, as indicated by improved growth performance of callus under low-temperature conditions. Conversely, silencing this gene reduced cold tolerance, resulting in reduced leaf photosynthetic capacity and lower soluble sugar content in roots. Furthermore, we provide conclusive evidence that RcWRKY71 directly binds to and transcriptionally activates its downstream gene RcGolS1. Virus-induced gene silencing (VIGS) of RcGolS1 led to reduced cold tolerance, while its overexpression increased cold resistance by activating raffinose family oligosaccharide (RFOs) biosynthesis, as evidenced by the accumulation of myo-inositol, galactinol, and raffinose. Our study reveals a novel RcWRKY71-RcGolS1 regulatory module within the cold hardiness signaling pathway and provides key molecular evidence explaining how cryoprotective oligosaccharides accumulate to facilitate overwintering adaptation in perennial woody plants. These findings provide valuable genetic resources and a theoretical foundation for breeding new, cold-resistant rose germplasm through genetic engineering.