Expression of alpha-amylase genes in cultured rice suspension cells is induced by sucrose starvation. To study the mechanism of sugar metabolite regulation on the expression of individual alpha-amylase genes, DNA fragments specific to each of eight rice alpha-amylase genes were synthesized and used as gene-specific probes. Comparison of the relative abundance of mRNA revealed that expression of the eight alpha-amylase genes in rice cells was differentially regulated by sucrose starvation. Accumulation of all the alpha-amylase mRNAs increased in response to sucrose starvation; however, levels of the alphaAmy3 and alphaAmy8 mRNAs were distinctly higher and constituted 90% of total alpha-amylase mRNAs. RNA gel blot and nuclear run-on transcription analyses demonstrated a positive correlation between the increased transcription rates and the elevated steady-state levels of alpha-amylase mRNAs induced by sucrose starvation. The half-lives of alphaAmy3, alphaAmy7, and alphaAmy8 were prolonged by sucrose-starvation; however, the stability of the three mRNAs seems controlled by different mechanisms. The translation inhibitors cycloheximide and anisomycin preferentially blocked the sucrose-suppressed expression of alphaAmy3 but not that of alphaAmy7 and alphaAmy8. These inhibitors also enhanced the sucrose starvation-induced accumulation of alphaAmy3 mRNA but not that of alphaAmy7 or alphaAmy8 mRNAs. Cycloheximide did not significantly alter the transcription rates of alpha-amylase genes, suggesting that labile proteins may selectively stabilize the alphaAmy7 and alphaAmy8 mRNAs but destabilize the alphaAmy3 mRNA.

SummaryCarbon metabolites suppress the expression of a-amylase genes in germinating seeds and in suspension-cultured cells of rice. We have used suspension cell culture as a model system to study the mechanisms of metabolic regulation of a-amylase gene expression in rice. Both transcription rate and mRNA stability increased as cells were starved of sucrose; the transcription rate of a-amylase genes in cells starved of sucrose for 24 h was seven times greater than in cells provided with sucrose. The half-life of a-amylase mRNA was less than 1 h in cells provided with sucrose, but increased to 12 h in cells starved of this sugar. A protein synthesis inhibitor, cycloheximide (CHX), induced massive accumulation of a-amylase mRNA in cells provided with sucrose. The longer half-life of mRNA induced by sucrose starvation and the massive accumulation of mRNA caused by CHX were specific to the a-amylase genes, since actin genes were not similarly affected. Our findings suggest that both transcriptional and post-transcriptional control mechanisms are important in the metabolic regulation of a-amylase gene expression and de n o w synthesized proteins are involved in these mechanisms. The expression of a-amylase and actin genes is regulated in an opposite manner by sugars, which also suggests the operation of a differential regulatory mechanism under different growth conditions.