Genome-wide identification, evolutionary and expression analyses of the DUF538 gene family in maize.

The DUF538 (Domain of Unknown Function 538) gene family plays critical roles in plant growth, development, and stress responses, yet its phylogenetic distribution, sequence evolution, and expression characteristics remain poorly understood. Here, we identified 196 DUF538 genes across 13 plants, all of which were specific to land plants. Phylogenetic analysis revealed four distinct subgroups within this family, with subgroups A and B likely originating from ancestral whole-genome duplication (WGD) events in angiosperms. The ancestral subgroups (C/D) exhibited distinct sequence features, biophysical properties, and selective pressures compared to subgroups A/B, whereas A/B underwent gene structure simplification, sequence divergence, and relaxation of selective constraints. In maize, co-expression networks analysis linked DUF538 genes to root development (e.g., flavonoid biosynthesis and kinase signaling) and leaf stress responses (e.g., hydrolase activity and defense signaling). Promoter and expression analyses revealed subgroup-specific enrichment of cis-elements, which correlated with differential expression patterns during development and under abiotic stress. Notably, under CaCl₂ stress, paralogs within subgroups were activated sequentially, sustaining a prolonged response while minimizing functional redundancy through temporal differentiation. Collectively, these findings demonstrate that DUF538 genes have achieved functional diversification through sequence divergence, relaxed selection, and spatiotemporal expression regulation, thereby providing potential targets for enhancing crop stress resilience.