Cytochromes P450 CYP94C1 and CYP94B3 catalyze two successive oxidation steps of plant hormone Jasmonoyl-isoleucine for catabolic turnover.
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.
Institut de Biologie Moléculaire des Plantes, Unité Propre de Recherche 2357 CNRS, Université de Strasbourg, 12 rue du Général Zimmer, 67084 Strasbourg Cedex, France. thierry.heitz@ibmp-cnrs.unistra.fr
Thierry
T
Heitz
Emilie
E
Widemann
Raphaël
R
Lugan
Laurence
L
Miesch
Pascaline
P
Ullmann
Laurent
L
Désaubry
Emilie
E
Holder
Bernard
B
Grausem
Sylvie
S
Kandel
Michel
M
Miesch
Danièle
D
Werck-Reichhart
Franck
F
Pinot
D017360:Arabidopsis / Q000201:enzymology* / Q000235:genetics; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D003517:Cyclopentanes / Q000378:metabolism*; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D005838:Genotype; D007532:Isoleucine / Q000031:analogs & derivatives* / Q000378:metabolism; D008660:Metabolism / Q000502:physiology; D009713:Nucleotidyltransferases / Q000378:metabolism; D010084:Oxidation-Reduction; D010937:Plant Growth Regulators / Q000378:metabolism*; D018515:Plant Leaves / Q000201:enzymology; D015398:Signal Transduction / Q000502:physiology; D010453:Peptide Synthases
D029681:Arabidopsis Proteins; D003517:Cyclopentanes; D003577:Cytochrome P-450 Enzyme System; D007532:Isoleucine; D009713:Nucleotidyltransferases; D010937:Plant Growth Regulators; C112279:COI1 protein, Arabidopsis; C532883:jasmonoyl-isoleucine; C560423:cytochrome P-450 94B3, Arabidopsis; C571537:cytochrome P-450 94C1, Arabidopsis; C412831:AT2G46370 protein, Arabidopsis; D010453:Peptide Synthases