Carbohydrate Starvation Stimulates Differential Expression of Rice α-Amylase Genes That Is Modulated through Complicated Transcriptional and Posttranscriptional Processes

Abstract

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.

The 3′ untranslated region of a rice α-amylase gene functions as a sugar-dependent mRNA stability determinant

“…Five micrograms of total RNA was loaded in each lane. The same blot was stripped and rehybridized with indicated probes as described previously (8). The probes were Luc, Act1 (22), and rRNA (23) cDNAs and ␣Amy3-specific DNA (8).…”

“…The same blot was stripped and rehybridized with indicated probes as described previously (8). The probes were Luc, Act1 (22), and rRNA (23) cDNAs and ␣Amy3-specific DNA (8). The Luc and ␣Amy3 probes hybridized to a same blot, and the Act and rRNA probes hybridized to another parallelprepared blot.…”

“…In rice germinating embryos and cultured suspension cells, expression of ␣-amylase genes is activated by sugar depletion and repressed by sugar provision (3,5,6). An increase in both the transcription rate and mRNA half-life under sucrose depletion contributes to the increase in the steady-state level of ␣-amylase mRNA (7,8). Studies in transgenic rice confirmed that the ␣-amylase gene promoters control sugar-dependent repression of reporter gene expression (9)(10)(11).…”

“…Previously, we have shown that the mRNA half-life of one rice ␣-amylase gene, ␣Amy3, decreased from 6 h to 1.5 h under repressed conditions (8). By examining the transient expression of chimeric genes in rice protoplasts, the 3Ј UTR of ␣Amy3 mRNA was shown to mediate sugar-dependent repression of fused heterologous gene expression (17).…”

The SnRK1A Protein Kinase Plays a Key Role in Sugar Signaling during Germination and Seedling Growth of Rice

“…aAmy3 and aAmy8 constitute two major rice a-amylases expressed under sugar-depleted conditions and are important for the catabolism of starch as a carbon source when sugar levels become too low to sustain respiration and growth in rice suspension cells and germinating embryos Yu, 1999;Chen et al, 2006). Sugar repression of a-amylase gene expression involves the control of transcription rate and mRNA stability (Chan et al, 1994;Sheu et al, 1994Sheu et al, , 1996Yu, 1998a, 1998b), and common regulatory elements in aAmy3 and aAmy8 sugar response complexes (SRCs) have been identified (Hwang et al, 1998;Lu et al, 1998;Toyofuku et al, 1998;Chen et al, 2006). The 100-bp aAmy3 SRC contains three essential motifs, the GC box, the G box, and the TA box, for high-level promoter activity under sugar-depleted conditions (Lu et al, 1998).…”