Cytochrome Is an Obligate Electron Shuttle Protein for Syringyl Lignin Biosynthesis in Arabidopsis.

Angiosperms have evolved the metabolic capacity to synthesize -hydroxyphenyl, guaiacyl (G), and syringyl (S) lignin subunits in their cell walls to better adapt to the harsh terrestrial environment. The structural characteristics of lignin subunits are essentially determined by three cytochrome P450-catalzyed reactions. NADPH-dependent cytochrome P450 oxidoreductase (CPR) is commonly regarded as the electron carrier for P450-catalyzed reactions during monolignol biosynthesis. Here, we show that cytochrome isoform D (CB5D) is an indispensable electron shuttle protein specific for S-lignin biosynthesis. Arabidopsis () CB5D localizes to the endoplasmic reticulum membrane and physically associates with monolignol P450 enzymes. Disrupting in Arabidopsis resulted in a >60% reduction in S-lignin subunit levels but no impairment in G-lignin formation compared with the wild type, which sharply contrasts with the impaired G- and S-lignin synthesis observed after disrupting , encoding Arabidopsis CPR. The defective S-lignin synthesis in mutants was rescued by the expression of the gene encoding CB5D but not with mutant CB5D devoid of its electron shuttle properties. Disrupting suppressed the catalytic activity of both cinnamic acid 4-hydroxylase and ferulate 5-hydroxylase (F5H), but eliminating specifically depleted the latter's activity. Therefore, CB5D functions as an obligate electron shuttle intermediate that specifically augments F5H-catalyzed reactions, thereby controlling S-lignin biosynthesis.