A decrease in transcript stability is one of the important mechanisms that control the sugar repression of alpha-amylase gene expression in rice suspension cells. In this study, we investigated the function of the 3' untranslated region (3'UTR) of a rice alpha-amylase gene, alpha Amy3, in relation to sugar-dependent accumulation of mRNA. By examining the transient expression of chimeric genes in rice protoplasts, we were able to demonstrate that the alpha Amy3 3'UTR mediated the sugar-dependent repression of fused heterologous gene expression. The same kinetics of accumulation of alpha Amy3 mRNA and reporter mRNA carrying the alpha Amy3 3'UTR in response to glucose deprivation were observed, suggesting that the alpha Amy3 3'UTR is sufficient, and probably the major determinant for controlling the abundance of these transcripts. Functional analysis of two subdomains of alpha Amy3 3'UTR by insertion into a sugar-inducible chimeric gene confirmed their roles in sugar repressibility. The regulatory sequences in the alpha Amy3 3'UTR may act as potent determinants of mRNA stability in response to sugar availability. This finding has important implications for studying the regulatory mechanism of sugar repression in eukaryotes.