Mode of Action of Abscisic Acid in Barley Aleurone Layers: Induction of Proteins by Abscisic Acid

198 p.Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 1987.Abscisic acid (ABA) induces at least 16 polypeptides in barley aleurone layers as determined by two-dimensional gel electrophoretic analysis. The induction seems to be at the mRNA level because the message levels of the induced proteins closely parallel the protein levels. None of the ABA-induced proteins are secreted. The most abundant ABA-induced protein, p29, is localized in the 1,000g pellet and is sensitive to thioprotease degradation. The 23 kD ABA-induced protein cross-reacts with antibody against an $\alpha$-amylase/subtilisin inhibitor, and the 35/36 kD doublet cross-reacts with antibody against a barley lectin specific for free amino sugars. The most abundant induced protein, p29, can be greatly enriched by a combination of low pH, low concentrations of isopropanol, high temperature, and reducing conditions. Phaseic acis (PA), the first stable metabolite of ABA, is probably the active component of the hormone in inhibiting GA-induced $\alpha$-amylase synthesis when ABA is added 20 hours after GA, based on studies with two monooxygenase inhibitors, paclobutrazol and tetcyclacis, which block the conversion of ABA to PA. However, when ABA is added at the same time as GA, ABA itself appears to be active, at least toward the low pI $\alpha$-amylase isozymes. A number of stress conditions, including heat shock, chemical stress (arsenite and cadmium), chilling stress, water stress (imposed by PEG 400, PEG 8,000, mannitol, or sorbitol), and salt stress (imposed by NaCl, KCl, or CaCl$\sb2$), were tested for the induction of proteins. Only salinity (NaCl) stress and water stress imposed by sugar alcohols induced the same ABA-inducible proteins. The induction of these proteins by salt and sorbitol is blocked by the ABA biosynthesis inhibitor fluridone, indicating the involvement of ABA. Determination of ABA content by ELISA indicates that sorbitol induces ABA proteins through an increase in intracellular ABA content, whereas salt induces ABA proteins without any apparent ABA increase. A model based on the compartmentation of intracellular ABA is proposed to explain the induction of ABA proteins by salt.U of I OnlyRestricted to the U of I community idenfinitely during batch ingest of legacy ETD