Control of α-Amylase mRNA Accumulation by Gibberellic Acid and Calcium in Barley Aleurone Layers

Abstract

Pulse-labeling of barley (Hordeum vulgare L. cvHimalaya) aleurone layers incubated for 13 hours in 2.5 micromolar gibberellic acid (GA3) with or without 5 millimolar CaCI2 shows that a-amylase isozymes 3 and 4 are notsynthesized in vivo in the absence of Ca2. A cDNA clone for a-amylase was isolated and used to measure a-amylase mRNA levels in aleurone layers incubated in the presence and absence of Ca2'. No difference was observed in a-amylase mRNA levels between layers incubated for 12 hours in 2.5 micromolar GA3 with 5 millimolar CaC12 and layers incubated in GA3 alone. RNA isolated from layers incubated for 12 hours in GA3 with and without Ca21 was translated in vitro and was found to produce the same complement of translation products regardless of the presence of Ca2' in the incubation medium. Immuno-precipitation of translation products showed that the RNA for a-amylase synthesized in Ca2-deprived aleurone layers was translatable. Ca2' is required for the synthesis of a-amylase isozymes 3 and 4 at a step after mRNA accumulation and processing. Gibberellic acid induces the synthesis and secretion of a-amylase in barley aleurone layers (for reviews, see 11, 12, and 15). The accumulation of a-amylase mRNA is controlled by GA3 in barley (4, 9, 10, 25-29) and wheat (1) aleurone although whether this control is at the level of transcription or of mRNA stability is still undetermined.