Arabidopsis CYP707As Encode (+)-Abscisic Acid 8-Hydroxylase, a Key Enzyme in the Oxidative Catabolism of Abscisic Acid

Abscisic acid (ABA) is involved in a number of critical processes in normal growth and development as well as in adaptive responses to environmental stresses. For correct and accurate actions, a physiologically active ABA level is controlled through fine-tuning of de novo biosynthesis and catabolism. The hydroxylation at the 8ʹ-position of ABA is known as the key step of ABA catabolism, and this reaction is catalyzed by ABA 8ʹ-hydroxylase, a cytochrome P450. Here, we demonstrate CYP707As as the P450 responsible for the 8ʹ-hydroxylation of (+)-ABA. First, all four CYP707A cDNAs were cloned from Arabidopsis and used for the production of the recombinant proteins in insect cells using a baculovirus system. The insect cells expressing CYP707A3 efficiently metabolized (+)-ABA to yield phaseic acid, the isomerized form of 8ʹ-hydroxy-ABA. The microsomes from the insect cells exhibited very strong activity of 8ʹ-hydroxylation of (+)-ABA (Km = 1.3 (mu)M and kcat = 15 min^-1). The solubilized CYP707A3 protein bound (+)-ABA with the binding constant Ks = 3.5 (mu)M, but did not bind (–)-ABA. Detailed analyses of the reaction products confirmed that CYP707A3 does not have the isomerization activity of 8ʹ-hydroxy-ABA to phaseic acid. Further experiments revealed that Arabidopsis CYP707A1 and CYP707A4 also encode ABA 8ʹ-hydroxylase. The transcripts of the CYP707A genes increased in response to salt, osmotic, and dehydration stresses as well as ABA. These results establish that the CYP707A family plays a key role in regulating the ABA level through the 8ʹhydroxylation of (+)-ABA.