Genomic analysis of TIFY genes in two Medicago and insight into MsTF2-mediated abiotic stress tolerance.

key messageBioinformatics analysis revealed 39 TIFY genes in two Medicago species, with MsTF2 significantly improving abiotic stress tolerance in both prokaryotic and eukaryotic cells. Transcription factor (TF) serves as crucial regulatory proteins in eukaryotes, facilitating RNA polymerase's ability to initiate transcription at particular promoter regions. TIFY proteins are crucial for plant growth and development, as well as signal transduction and stress responses. The TIFY gene family plays crucial roles in stress responses in higher plants, yet comprehensive studies on this protein family in Medicago species remain unexplored. The study identified 39 TIFY genes, comprising 19 in Medicago sativa and 20 in Medicago truncatula, which were unevenly distributed across their respective eight chromosomes. Synteny analysis indicated that tandem and segmental duplications were the primary drivers of TIFY family expansion in these two species. Promoter cis-element analysis revealed an enrichment of stress-responsive motifs, suggesting their functional involvement in abiotic stress adaptation. Transcriptomic and RT-qPCR analyses demonstrated that most TIFY members exhibited strong induction under drought, cold, and high-salinity conditions. Notably, MsTF2, a nuclear-localized protein, displayed the most pronounced stress-responsive expression among all examined TIFY genes. Heterologous expression of MsTF2 in both prokaryotic and eukaryotic systems conferred significant enhancement of tolerance to drought, extreme temperature, and high-salinity stresses. These findings provide a molecular framework for understanding MsTF2-mediated stress resistance and highlight potential genetic targets for improving abiotic stress resilience in Medicago through molecular breeding strategies.