Genome-Wide Identification, Molecular Evolution, and Abiotic Stress-Responsive Regulation of Cupin Superfamily Genes in Rice (Oryza sativa L.)

The Cupin superfamily, characterized by a conserved β-barrel structure, plays crucial roles in plant growth, development, and stress responses. However, comprehensive analyses of this gene family in rice remains limited. Here, we performed a genome-wide identification, molecular evolution, and expression analysis of Cupin genes in rice under abiotic stress. Utilizing the telomere-to-telomere (T2T) genome of rice, 54 Cupin genes (OsCupins) were identified and classified into four subfamilies (GLP, PIRIN, TRR14, and ARD) based on phylogenetic relationships with Arabidopsis homologs. These genes were unevenly distributed across ten chromosomes, with tandem and segmental duplications driving their expansion. Structural and synteny analyses revealed conserved motifs and orthologous relationships with sorghum and Arabidopsis. The promoter regions of OsCupins were enriched in stress-responsive cis-elements, including ABRE, MYB, and MYC motifs. qRT-PCR data demonstrated the significant upregulation of multiple OsCupins (e.g., OsGLP15, OsGLP38, and OsGLP43) under NaCl and PEG 6000 treatments. Functional validation in yeast showed that the overexpression of OsGLP15, OsGLP38, or OsGLP43 enhanced salt and drought tolerance in yeast, with OsGLP43 exhibiting the strongest stress resilience. Our findings provide insights into the evolutionary dynamics and stress-responsive regulatory mechanisms of the Cupin superfamily in rice, offering potential targets for enhancing abiotic stress tolerance in this critical crop.