Abstract
Pulse-labeling of barley (Hordeum vulgare L. cv Himalaya) aleurone layers incubated for 13 hours in 2.5 micromolar gibberellic acid (GA(3)) with or without 5 millimolar CaCl(2) shows that alpha-amylase isozymes 3 and 4 are not synthesized in vivo in the absence of Ca(2+). A cDNA clone for alpha-amylase was isolated and used to measure alpha-amylase mRNA levels in aleurone layers incubated in the presence and absence of Ca(2+). No difference was observed in alpha-amylase mRNA levels between layers incubated for 12 hours in 2.5 micromolar GA(3) with 5 millimolar CaCl(2) and layers incubated in GA(3) alone. RNA isolated from layers incubated for 12 hours in GA(3) with and without Ca(2+) was translated in vitro and was found to produce the same complement of translation products regardless of the presence of Ca(2+) in the incubation medium. Immunoprecipitation of translation products showed that the RNA for alpha-amylase synthesized in Ca(2+)-deprived aleurone layers was translatable. Ca(2+) is required for the synthesis of alpha-amylase isozymes 3 and 4 at a step after mRNA accumulation and processing.