Synthesis and Release of a-Amylase

The 8-h lag period after GA addition is followed by a 16-(or more) h period of rapid a-amylase synthesis (Fig. 7.1 A). More ultrastructural changes are said to occur in the aleurone cells during this time [66]. These include further proliferation of the RER, distention of the RER cisternae, continued reduction in the size of the aleurone grains, decreases in the number of oil bodies, an increase in the number of plastids, and loss of the phytin globoid. 

The barley aleurone system has attracted biochemists interested in determining if GA acts at the nuclear level by inducing transcription of the gene for a-amylase. Recent studies have taken advantage of the observation that many messenger RNAs in eukaryotic cells contain a covalently linked poly(adenylic acid) seg- 

The possibility of post-transcriptional control of enzyme synthesis has been raised by several workers, and it has been noted that methylation of purine residues of tRNA and heavy rRNA is enhanced in GA-treated aleurone tissue [18]. It is not clear how GA might influence this process, or why methylation of certain types of RNA should be essential for a-amylase synthesis. Other studies claiming to show post-transcriptional control by GA [16] have received no support [17]. 

The synthesis and secretion of a-amylase by aleurone cells is sharply reduced when GA is withdrawn, say after a 12-h treatment [22, 67]. If the presence of GA is required all the time for the continued production of enzyme, then it is necessary to determine if there is continued synthesis of a-amylase mRNA after 12 h of incubation on GA. If not (and results in Fig. 7.3A suggest not) then the requirement for the continued presence of GA must be to control some event in the synthesis of a-amylase other than the production of mRNA — perhaps the translation process. 

Finally, a brief word about cyclic AMP. Activation of the enzyme adenyl cyclase which catalyses cyclic 3, 5 -adenosine monophophate synthesis constitutes the initial molecular event in the target cells of several mammalian hormones. The nucleotide then initiates a series of events leading to a final response characteristic of the hormone. Despite numerous studies on the barley aleurone layer there is no convincing evidence that GA action is mediated via cyclic AMP [73] in fact the weight of evidence is against this nucleotide playing an important role in any plant tissue [7]. 

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Induction of de novo synthesis of a-amylase by GA in isolated aleurone layers is evident after a lag period of approximately 8 hr following administration of the hormone. In keeping with hormone responses generally, GA must be present continuously if the de novo synthesis of hydrolases is to be sustained. Synthesis of new RNA is essential to the GA-induction of de novo synthesis of hydrolases. Actinomycin D, an inhibitor of RNA synthesis, inhibits the synthesis and release of a-amylase if the inhibitor is presented during the first 7 to 8 hr after treatment. Inhibitors of protein synthesis, such as cycloheximide, also inhibit GA-induction of hydrolases. And, interestingly, abscisic acid, a growth-inhibiting hormone, inhibits GA-induced a-amylase synthesis as well. 

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