Arabidopsis CYP98A3 mediating aromatic 3-hydroxylation. Developmental regulation of the gene, and expression in yeast.
The general phenylpropanoid pathways generate a wide array of aromatic secondary metabolites that range from monolignols, which are ubiquitous in all plants, to sinapine, which is confined to crucifer seeds. The biosynthesis of these compounds involves hydroxylated and methoxylated cinnamyl acid, aldehyde, or alcohol intermediates. Of the three enzymes originally proposed to hydroxylate the 4-, 3-, and 5-positions of the aromatic ring, cinnamate 4-hydroxylase (C4H), which converts trans-cinnamic acid to p-coumaric acid, is the best characterized and is also the archetypal plant P450 monooxygenase. Ferulic acid 5-hydroxylase (F5H), a P450 that catalyzes 5-hydroxylation, has also been studied, but the presumptive 3-hydroxylase converting p-coumarate to caffeate has been elusive. We have found that Arabidopsis CYP98A3, also a P450, could hydroxylate p-coumaric acid to caffeic acid in vivo when expressed in yeast (Saccharomyces cerevisiae) cells, albeit very slowly. CYP98A3 transcript was found in Arabidopsis stem and silique, resembling both C4H and F5H in this respect. CYP98A3 showed further resemblance to C4H in being highly active in root, but differed from F5H in this regard. In transgenic Arabidopsis, the promoters of CYP98A3 and C4H showed wound inducibility and a comparable developmental regulation throughout the life cycle, except in seeds, where the CYP98A3 promoter construct was inactive while remaining active in silique walls. Within stem and root tissue, the gene product and the promoter activity of CYP98A3 were most abundant in lignifying cells. Collectively, these studies show involvement of CYP98A3 in the general phenylpropanoid metabolism, and suggest a downstream function for CYP98A3 relative to the broader and upstream role of C4H.
Nair
Ramesh B
RB
Plant Biotechnology Institute, National Research Council of Canada, 110 Gymnasium Place, Saskatoon, Saskatchewan, Canada S7N 0W9.
Xia
Qun
Q
Kartha
Cyril J
CJ
Kurylo
Eugen
E
Hirji
Rozina N
RN
Datla
Raju
R
Selvaraj
Gopalan
G
D017360:Arabidopsis / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D029681:Arabidopsis Proteins; D002109:Caffeic Acids / Q000378:metabolism; D003373:Coumaric Acids / Q000378:metabolism; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D018507:Gene Expression Regulation, Developmental; D018506:Gene Expression Regulation, Plant; D006900:Hydroxylation; D015151:Immunoblotting; D007150:Immunohistochemistry; D008031:Lignin / Q000378:metabolism; D006899:Mixed Function Oxygenases / Q000235:genetics / Q000378:metabolism*; D010666:Phenylpropionates / Q000378:metabolism; D018517:Plant Roots / Q000378:metabolism; D018547:Plant Stems / Q000378:metabolism; D030821:Plants, Genetically Modified; D011422:Propionates; D012441:Saccharomyces cerevisiae / Q000235:genetics; D050564:Trans-Cinnamate 4-Monooxygenase
D029681:Arabidopsis Proteins; D002109:Caffeic Acids; D003373:Coumaric Acids; D010666:Phenylpropionates; D011422:Propionates; D008031:Lignin; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; C438015:CYP98A3 protein, Arabidopsis; D050564:Trans-Cinnamate 4-Monooxygenase; C495469:p-coumaric acid; C040048:caffeic acid