Cytochromes P450 CYP94C1 and CYP94B3 Catalyze Two Successive Oxidation Steps of Plant Hormone Jasmonoyl-isoleucine for Catabolic Turnover

• Published in: The Journal of biological chemistry Vol. 287; no. 9; pp. 6296 - 6306
• Main Authors: Heitz, Thierry, Widemann, Emilie, Lugan, Raphaël, Miesch, Laurence, Ullmann, Pascaline, Désaubry, Laurent, Holder, Emilie, Grausem, Bernard, Kandel, Sylvie, Miesch, Michel, Werck-Reichhart, Danièle, Pinot, Franck
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 24.02.2012; American Society for Biochemistry and Molecular Biology
• Subjects: Arabidopsis; Arabidopsis - enzymology; Arabidopsis - genetics; Arabidopsis Proteins - genetics; Arabidopsis Proteins - metabolism; Biochemistry; Biochemistry, Molecular Biology; Botanics; Cyclopentanes - metabolism; CYP94; Cytochrome P-450 Enzyme System - genetics; Cytochrome P-450 Enzyme System - metabolism; Cytochrome P450; Genotype; Isoleucine - analogs & derivatives; Isoleucine - metabolism; Jasmonate Metabolism; Life Sciences; Metabolic Profiling; Metabolism; Metabolism - physiology; Nucleotidyltransferases - metabolism; Oxidation-Reduction; Plant Biology; Plant Defense; Plant Growth Regulators - metabolism; Plant Hormones; Plant Leaves - enzymology; Signal Transduction - physiology; Signaling; Vegetal Biology; Wound Response; Plant Hormones; Signaling; Jasmonate Metabolism; Arabidopsis; Cytochrome P450; Metabolic Profiling; Wound Response; Plant Defense; Metabolism; CYP94
• ISSN: 0021-9258; 1083-351X; 1083-351X
• DOI: 10.1074/jbc.M111.316364

The jasmonate hormonal pathway regulates important defensive and developmental processes in plants. Jasmonoyl-isoleucine (JA-Ile) has been identified as a specific ligand binding the COI1-JAZ co-receptor to relieve repression of jasmonate responses. Two JA-Ile derivatives, 12OH-JA-Ile and 12COOH-JA-Ile, accumulate in wounded Arabidopsis leaves in a COI1- and JAR1-dependent manner and reflect catabolic turnover of the hormone. Here we report the biochemical and genetic characterization of two wound-inducible cytochromes P450, CYP94C1 and CYP94B3, that are involved in JA-Ile oxidation. Both enzymes expressed in yeast catalyze two successive oxidation steps of JA-Ile with distinct characteristics. CYP94B3 performed efficiently the initial hydroxylation of JA-Ile to 12OH-JA-Ile, with little conversion to 12COOH-JA-Ile, whereas CYP94C1 catalyzed preferentially carboxy-derivative formation. Metabolic analysis of loss- and gain-of-function plant lines were consistent with in vitro enzymatic properties. cyp94b3 mutants were largely impaired in 12OH-JA-Ile levels upon wounding and to a lesser extent in 12COOH-JA-Ile levels. In contrast, cyp94c1 plants showed wild-type 12OH-JA-Ile accumulation but lost about 60% 12COOH-JA-Ile. cyp94b3cyp94c1 double mutants hyperaccumulated JA-Ile with near abolition of 12COOH-JA-Ile. Distinct JA-Ile oxidation patterns in different plant genotypes were correlated with specific JA-responsive transcript profiles, indicating that JA-Ile oxidation status affects signaling. Interestingly, exaggerated JA-Ile levels were associated with JAZ repressor hyperinduction but did not enhance durably defense gene induction, revealing a novel negative feedback signaling loop. Finally, interfering with CYP94 gene expression affected root growth sensitivity to exogenous jasmonic acid. These results identify CYP94B3/C1-mediated oxidation as a major catabolic route for turning over the JA-Ile hormone. Oxidized derivatives of the plant hormone jasmonoyl-isoleucine accumulate in wounded Arabidopsis leaves. Cytochromes P450 CYP94C1 and CYP94B3 cooperate to catalyze the formation of 12OH-JA-Ile and 12COOH-JA-Ile. CYP94C1 and CYP94B3 define a major route for JA-Ile catabolism. Elucidation of CYP94-mediated JA-Ile oxidation opens new avenues for understanding jasmonate metabolism and signaling.