Evolutionary dynamics and transcriptional diversification of GAD gene family in plants: a pan-species perspective in silico analysis

Glutamate decarboxylase (GAD) catalyzes the biosynthesis of γ-aminobutyric acid (GABA), a key metabolite and signaling molecule in plant growth and stress responses. However, the evolutionary history and regulatory complexity of GAD genes across plant lineages remain poorly understood. We identified 182 GAD genes from 31 plant species, including basal Embryophyta, monocots, and eudicots. Phylogenetic analyses grouped these genes into three major subfamilies (A, B, C), with subfamily A confined to early land plants, suggesting an ancient evolutionary origin. Tandem duplication was the dominant mechanism for expansion in eudicots, whereas annotated splice isoforms suggest potential contributions to transcriptomic diversity in half of the species analyzed. Structural characterization revealed high motif conservation but also specific domain innovations, such as heavy-metal-associated domains in flax. Promoter and intron analyses showed a high abundance of light-, ABA-, MeJA-, and anaerobic-responsive elements. Notably, intron length correlated positively with gene expression across tissues, suggesting a key role for intron-mediated enhancement. RNA-seq data from Dendrobium catenatum, Tagetes erecta, Arabidopsis, rice, and cotton revealed dynamic tissue-specific expression patterns, with certain ancestral GAD genes displaying low, restricted expression, and newer paralogs showing broader profiles. The GAD gene family exhibits lineage- and tissue-specific expression patterns rather than strict subfamily-level functional divergence. Intronic length and regulatory motif content strongly correlate with transcriptional activity. This work offers novel insights into the evolution, expression regulation, and functional diversification of GAD genes across plants, with practical implications for crop improvement and stress resistance.