Identification analysis of GAD gene family, and the role of BoGAD 5 in GABA enrichment in broccoli sprouts

Glutamate decarboxylase (GAD) catalyzes the conversion of glutamic acid (Glu) to γ-aminobutyric acid (GABA), playing a crucial role in plant growth and development. A genome-wide identification and expression analysis of the GAD gene family in broccoli were performed to understand their functional characteristics. The GAD gene family in broccoli has amino acid lengths ranging from 408 to 622 and isoelectric points from 5.4 to 8.71. BoGADs were classified into three branches based on gene sequence and phylogenetic analysis, with two branches having distributed members. Promoter element analysis revealed numerous stress and hormone response elements. Nine BoGAD genes were expressed in roots, stems, and leaves of broccoli sprouts, primarily in leaves, with BoGAD 5 showing the highest expression, followed by BoGAD 2. CaCl₂ treatment increased GABA content in broccoli sprouts, and BoGADs gene expression increased, among which BoGAD 8 and BoGAD 5 were the most obvious, indicating that BoGADs were related to GABA enrichment in broccoli sprouts. Transient overexpression of BoGAD 5 in broccoli leaves significantly decreased Glu content while increasing GABA and succinic acid semialdehyde (SSA) content. GAD, γ-aminobutyrate transaminase (GABA-T), and succinate-semialdehyde dehydrogenase (SSADH) activities enhanced after BoGAD 5 overexpression. Thus, BoGAD 5 is associated with GABA enrichment in broccoli sprouts, promoting GABA enrichment through increasing activity of GABA shunt enzymes.