Abstract
Basic leucine zipper (bZIP) genes are extensively involved in various physiological processes, including seed development, light signal regulation, and stress responses. However, the regulatory roles of soybean GmbZIP genes in mediating abiotic stress responses remain poorly understood. To systematically identify abiotic stress-responsive GmbZIP genes in soybean and elucidate their evolutionary relationships, this study conducted a genome-wide identification and evolutionary analysis of the GmbZIP gene family using bioinformatics methods, combined with transcriptome data from waterlogging, salt, and shade stress treatments. Furthermore, comparative genomics was employed to screen for GmbZIP genes potentially associated with salt stress response, followed by protein-protein interaction network analysis and experimental validation via qRT-PCR. The results showed that a total of 92 GmbZIP genes were identified from the stress-related transcriptome datasets. Phylogenetic analysis classified them into 13 distinct subfamilies, each exhibiting significant evolutionary divergence and diversity in protein physicochemical properties, gene structures, and conserved motifs. Collinearity analysis revealed 143, 122, 104, 49, and 30 collinear gene pairs between soybean and Medicago truncatula (M. truncatula), Solanum lycopersicum (S. lycopersicum), Arabidopsis thaliana, Oryza sativa (O. sativa), and Zea mays (Z. mays), respectively. Segmental duplication was identified as the primary driver of the expansion of this gene family, with purifying selection playing a dominant role during its evolution. KEGG enrichment analysis indicated that GmbZIP genes are predominantly enriched in plant hormone signal transduction pathways. Consistent with this finding, cis-regulatory element analysis of promoters showed that 98.9% of the GmbZIP genes contain five types of hormone-responsive elements. Through comparative genomic screening, 12 candidate GmbZIP genes potentially involved in salt stress response were identified. Both protein-protein interaction network prediction and qRT-PCR expression validation support that these genes may be involved in the regulation of salt stress. In summary, this study presents the first systematic identification and evolutionary analysis of the abiotic stress-responsive GmbZIP gene family in soybean, providing an important foundation for further investigation into the functional mechanisms of these genes in plant abiotic stress responses.