Genome-wide identification of the GT transcription factor family in alfalfa and functional characterization of MsGT16 in salt stress tolerance

Alfalfa is a perennial legume forage crop, development, yield and quality are often affected by abiotic stresses. Members of the GT transcription factor family play important roles in plant growth and development, as well as in light response and abiotic stress response. However, no genome-wide studies exist on GT genes in alfalfa under abiotic stress conditions. A total of 22 GT geneswere identified in alfalfa, which were unevenly distributed across its 8 chromosomes. On the basis of sequence similarity and structural features, the genes were classified into five subfamilies. Collinearity analysis revealed two fragment duplication events in the alfalfa genome. Comparative collinearity analysis revealed 14 collinear GT genes between alfalfa and two model species (Arabidopsis and rice), highlighting the expansion and evolutionary divergence of the GT transcription factor family. Expression profiling under abiotic stress conditions demonstrated that GTs respond to salt, alkali, cold and drought stress. Among these genes, MsGT16 was significantly upregulated under multiple stress treatments. In the salt stress assays, compared with WT control plants, MsGT16-overexpressing Arabidopsis plants displayed stronger growth and higher antioxidant enzyme activity. In conclusion, this study presents the first genome-wide identification of 22 GT transcription factor family members in alfalfa and preliminarily suggests their involvement in abiotic stress responses. We further demonstrated that MsGT16 overexpression could improve the salt tolerance of Arabidopsis, providing evidence for its use as a key candidate gene for developing stress-resistant alfalfa cultivars.