9723408 Zeevaart Biosynthesis of the plant growth substance abscisic acid (ABA) is stimulated during water stress so that there is a large increase in the ABA content during drought. Following rehydration, ABA is rapidly degraded to phaseic acid (PA). This project deals with both the biosynthesis and catabolism of ABA. The research is focused on two key reactions: 1) the oxidative cleavage of epoxycarotenoids (C40) by a dioxygenase to form xanthoxin (Cl5), a precursor of ABA, and C25- epoxy-apocarotenoids; and 2) the oxidative deactivation of ABA to PA, catalyzed by ABA 8'- hydroxylase, a cytochrome P450 monooxygenase. The research is based on the hypothesis that the cleavage reaction is the key regulatory step in ABA biosynthesis. Experiments will be conducted to test this hypothesis at the mRNA and protein levels in water-stressed bean leaves and ripening avocado fruits, two systems that accumulate large amounts of ABA. Genes encoding the cleavage enzyme will be cloned by the polymerase chain reaction with degenerate primers of conserved domains in dioxygenases. Leaves will also be treated with ABA or ABA analogs prior to wilting to determine whether expression of the gene encoding the cleavage enzyme is subject to negative feed-back regulation. The strategy to clone ABA 8'-hydroxylase is based on differential expression of the 8'-hydroxylase in control and ABA-treated tissue. Putative cDNA clones for 8'-hydroxylase will be expressed in yeast and assayed for their ability to convert ABA to PA. Effects of tissue water status and ABA levels on expression of 8'-hydroxylase will be investigated. Abscisic acid (ABA) is a plant hormone that is involved in a number of physiological processes, such as closure of stomata, seed maturation and dormancy, and amelioration of environmental stresses (drought, salinity, low temperature). The cloning of two genes for key steps in ABA metabolism will provide powerful tools to manipulate ABA levels in plants. By raising their ABA levels, plants may be bett er adapted to grow under stressful conditions. This could be used to increase crop yields in unfavorable climates.
