title: Protein-protein and protein-membrane associations in the lignin pathway.
abstract: Supramolecular organization of enzymes is proposed to orchestrate metabolic complexity and help channel intermediates in different pathways. Phenylpropanoid metabolism has to direct up to 30% of the carbon fixed by plants to the biosynthesis of lignin precursors. Effective coupling of the enzymes in the pathway thus seems to be required. Subcellular localization, mobility, protein-protein, and protein-membrane interactions of four consecutive enzymes around the main branch point leading to lignin precursors was investigated in leaf tissues of Nicotiana benthamiana and cells of Arabidopsis thaliana. CYP73A5 and CYP98A3, the two Arabidopsis cytochrome P450s (P450s) catalyzing para- and meta-hydroxylations of the phenolic ring of monolignols were found to colocalize in the endoplasmic reticulum (ER) and to form homo- and heteromers. They moved along with the fast remodeling plant ER, but their lateral diffusion on the ER surface was restricted, likely due to association with other ER proteins. The connecting soluble enzyme hydroxycinnamoyltransferase (HCT), was found partially associated with the ER. Both HCT and the 4-coumaroyl-CoA ligase relocalized closer to the membrane upon P450 expression. Fluorescence lifetime imaging microscopy supports P450 colocalization and interaction with the soluble proteins, enhanced by the expression of the partner proteins. Protein relocalization was further enhanced in tissues undergoing wound repair. CYP98A3 was the most effective in driving protein association.
authors.affiliation: Institute of Plant Molecular Biology of Centre National de la Recherche Scientifique, Unité Propre de Recherche 2357, University of Strasbourg, F-67000 Strasbourg, France.
authors.forename: Jean-Etienne
authors.initials: JE
authors.lastname: Bassard
authors.forename: Ludovic
authors.initials: L
authors.lastname: Richert
authors.forename: Jan
authors.initials: J
authors.lastname: Geerinck
authors.forename: Hugues
authors.initials: H
authors.lastname: Renault
authors.forename: Frédéric
authors.initials: F
authors.lastname: Duval
authors.forename: Pascaline
authors.initials: P
authors.lastname: Ullmann
authors.forename: Martine
authors.initials: M
authors.lastname: Schmitt
authors.forename: Etienne
authors.initials: E
authors.lastname: Meyer
authors.forename: Jerôme
authors.initials: J
authors.lastname: Mutterer
authors.forename: Wout
authors.initials: W
authors.lastname: Boerjan
authors.forename: Geert
authors.initials: G
authors.lastname: De Jaeger
authors.forename: Yves
authors.initials: Y
authors.lastname: Mely
authors.forename: Alain
authors.initials: A
authors.lastname: Goossens
authors.forename: Danièle
authors.initials: D
authors.lastname: Werck-Reichhart
mesh_terms: D000214:Acyl Coenzyme A / Q000378:metabolism; D000217:Acyltransferases / Q000378:metabolism; D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D003066:Coenzyme A Ligases / Q000378:metabolism; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism; D049452:Green Fluorescent Proteins; D062385:Hydroxybenzoates / Q000378:metabolism; D006900:Hydroxylation; D008031:Lignin / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D018515:Plant Leaves / Q000235:genetics / Q000378:metabolism; D030821:Plants, Genetically Modified; D025941:Protein Interaction Mapping; D055503:Protein Multimerization; D011993:Recombinant Fusion Proteins; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism*; D050564:Trans-Cinnamate 4-Monooxygenase / Q000235:genetics / Q000378:metabolism*; D019076:Transgenes
chemical_list: D000214:Acyl Coenzyme A; D029681:Arabidopsis Proteins; C493362:CYP73A5 protein, Arabidopsis; D062385:Hydroxybenzoates; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; C485184:enhanced green fluorescent protein; C058644:4-coumaroyl-coenzyme A; D049452:Green Fluorescent Proteins; D008031:Lignin; D003577:Cytochrome P-450 Enzyme System; C438015:CYP98A3 protein, Arabidopsis; D050564:Trans-Cinnamate 4-Monooxygenase; D000217:Acyltransferases; D003066:Coenzyme A Ligases; C017616:phenolic acid