title: Cloning, functional expression, and characterization of CYP709C1, the first sub-terminal hydroxylase of long chain fatty acid in plants. Induction by chemicals and methyl jasmonate.
abstract: We cloned and characterized CYP709C1, a new plant cytochrome P450 belonging to the P450 family, that so far has no identified function except for clustering with a fatty acid metabolizing clade of P450 enzymes. We showed here that CYP709C1 is capable of hydroxylating fatty acids at the omega-1 and omega-2 positions. This work was performed after recoding and heterologous expression of a full-length cDNA isolated from a wheat cDNA library in an engineered yeast strain. Investigation on substrate specificity indicates that CYP709C1 metabolizes different fatty acids varying in their chain length (C12 to C18) and unsaturation. CYP709C1 is the first identified plant cytochrome P450 that can catalyze sub-terminal hydroxylation of C18 fatty acids. cis-9,10-Epoxystearic acid is metabolized with the highest efficiency, i.e. K((m)(app)) of 8 microM and V(max(app)) of 328 nmol/min/nmol P450. This, together with the fact that wheat possesses a microsomal peroxygenase able to synthesize this compound from oleic acid, strongly suggests that it is a physiological substrate. Hydroxylated fatty acids are implicated in plant defense events. We postulated that CYP709C1 could be involved in plant defense by producing such compounds. This receives support from the observation that (i) sub-terminal hydroxylation of 9,10-epoxystearic acid is induced (15-fold after 3 h) in microsomes of wheat seedlings treated with the stress hormone methyl jasmonate and (ii) CYP709C1 is enhanced at the transcriptional level by this treatment. CYP709C1 transcript also accumulated after treatment with a combination of the safener naphthalic acid anhydride and phenobarbital. This indicates a possible detoxifying function for CYP709C1 that we discussed.
authors.affiliation: Département Réponse Métabolique à l'Environnement Biotique, IBMP-CNRS, UPR 2357, 28 Rue Goethe, F-67083 Strasbourg Cedex, France.
authors.forename: Sylvie
authors.initials: S
authors.lastname: Kandel
authors.forename: Marc
authors.initials: M
authors.lastname: Morant
authors.forename: Irène
authors.initials: I
authors.lastname: Benveniste
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Blée
authors.forename: Danièle
authors.initials: D
authors.lastname: Werck-Reichhart
authors.forename: Franck
authors.initials: F
authors.lastname: Pinot
mesh_terms: D000085:Acetates / Q000494:pharmacology*; D015152:Blotting, Northern; D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer; D003001:Cloning, Molecular; D003517:Cyclopentanes / Q000494:pharmacology*; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry* / Q000235:genetics*; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005227:Fatty Acids / Q000378:metabolism*; D008401:Gas Chromatography-Mass Spectrometry; D015723:Gene Library; D018745:Genome, Plant; D007700:Kinetics; D007850:Lauric Acids / Q000378:metabolism; D008861:Microsomes / Q000378:metabolism; D008956:Models, Chemical; D009281:Naphthalenes / Q000378:metabolism; D054883:Oxylipins; D010634:Phenobarbital / Q000378:metabolism; D010940:Plant Proteins / Q000737:chemistry*; D013229:Stearic Acids / Q000378:metabolism; D013379:Substrate Specificity; D013997:Time Factors; D014908:Triticum / Q000235:genetics
chemical_list: D000085:Acetates; D003517:Cyclopentanes; D018076:DNA, Complementary; D005227:Fatty Acids; D007850:Lauric Acids; D009281:Naphthalenes; D054883:Oxylipins; D010940:Plant Proteins; D013229:Stearic Acids; C030358:lauric acid; C013519:9,10-epoxystearic acid; C012514:1,8-naphthalenedicarboxylic acid anhydride; C072239:methyl jasmonate; D003577:Cytochrome P-450 Enzyme System; D010634:Phenobarbital