Genome-Wide Identification and Functional Validation of the Cold-Tolerance WRKY Transcription Factor in Sweetpotato

Sweetpotato (Ipomoea batatas (L.) Lam.) is a vital global food crop whose production and geographical distribution are severely constrained by low-temperature stress. While WRKY transcription factors are known to regulate plant stress responses, their specific roles in sweetpotato cold tolerance remain largely unexplored. In this study, we conducted a genome-wide identification of the WRKY family in the hexaploid sweetpotato genome, identifying 476 IbWRKYs classified into three major groups and seven subgroups. Synteny analyses revealed that whole-genome duplication (WGD)/segmental duplication (SD) was the primary mode of gene duplication for this conserved family. To identify key cold-responsive regulators, we evaluated the cold tolerance phenotypes of 232 diverse accessions and performed comparative transcriptome profiling on representative cold-tolerant and cold sensitive cultivars. Integrated transcriptomic and co-expression network analyses highlighted IbWRKY460 as a leading cold-responsive candidate. Quantitative RT-PCR confirmed its cold-induced expression, the GFP fusion protein localized to the nucleus, and heterologous expression in yeast provided preliminary evidence of protective activity during low-temperature exposure. Together, these findings prioritize IbWRKY460 for in-planta functional validation and provide a genomic framework for investigating cold-responsive WRKY genes in sweetpotato.