Accumulation of γ-aminobutyric acid in giant-embryo rice grain in relation to glutamate decarboxylase activity and its gene expression during water soaking

Glutamate decarboxylase (GAD) is the rate-limiting enzyme for gamma-aminobutyric acid (GABA.) synthesis. However, the relationship of GABA accumulation, GAD activity, and its gene expression in rice grain during water soaking is unknown yet. We assessed the role of GAD activity and its gene expression to GABA accumulation after the rice grain was soaked in water for 0, 1, 2, 4, 8, 16, and 32 hr. The results showed a continuous accumulation of GABA in steeped embryo, embryoless brown rice, and complete brown rice, respectively. Among them, embryo of brown rice had the maximal accumulation. The activity of GAD was gradually increasing during water soaking, and the enzyme activity was significantly higher in the embryo than in either the embryoless brown rice or whole brown rice. The results further indicated that the activity of GAD was markedly stimulated by 0.5 mmol/L of Ca2+, whereas it was almost completely inhibited by 1 mmol/L of Hg2+. Accordingly, the content of GABA in embryo paralleled the activity of GAD treated with Ca2+ or Hg2+. To measure the expression of three GAD homologous genes in steeped rice grain, a RT-PCR method was developed using the housekeeping gene Actin as the control. Gene expression studies demonstrated differential expression patterns of the three GAD transcripts: OsGAD1, 2 and 3 in brown rice. These results suggested that the accumulation of GABA in rice grain could be attributed to the increase of OsGAD transcripts.