Expression pattern of two paralogs encoding cinnamyl alcohol dehydrogenases in Arabidopsis. Isolation and characterization of the corresponding mutants.
Studying Arabidopsis mutants of the phenylpropanoid pathway has unraveled several biosynthetic steps of monolignol synthesis. Most of the genes leading to monolignol synthesis have been characterized recently in this herbaceous plant, except those encoding cinnamyl alcohol dehydrogenase (CAD). We have used the complete sequencing of the Arabidopsis genome to highlight a new view of the complete CAD gene family. Among nine AtCAD genes, we have identified the two distinct paralogs AtCAD-C and AtCAD-D, which share 75% identity and are likely to be involved in lignin biosynthesis in other plants. Northern, semiquantitative restriction fragment-length polymorphism-reverse transcriptase-polymerase chain reaction and western analysis revealed that AtCAD-C and AtCAD-D mRNA and protein ratios were organ dependent. Promoter activities of both genes are high in fibers and in xylem bundles. However, AtCAD-C displayed a larger range of sites of expression than AtCAD-D. Arabidopsis null mutants (Atcad-D and Atcad-C) corresponding to both genes were isolated. CAD activities were drastically reduced in both mutants, with a higher impact on sinapyl alcohol dehydrogenase activity (6% and 38% of residual sinapyl alcohol dehydrogenase activities for Atcad-D and Atcad-C, respectively). Only Atcad-D showed a slight reduction in Klason lignin content and displayed modifications of lignin structure with a significant reduced proportion of conventional S lignin units in both stems and roots, together with the incorporation of sinapaldehyde structures ether linked at Cbeta. These results argue for a substantial role of AtCAD-D in lignification, and more specifically in the biosynthesis of sinapyl alcohol, the precursor of S lignin units.
Sibout
Richard
R
Natural Resources Canada, Canadian Forest Service, Laurentian Forestry Centre, 1055 du PEPS, P.O. Box 3800, Quebec, Canada G1V 4C7.
Eudes
Aymerick
A
Pollet
Brigitte
B
Goujon
Thomas
T
Mila
Isabelle
I
Granier
Fabienne
F
Séguin
Armand
A
Lapierre
Catherine
C
Jouanin
Lise
L
D000429:Alcohol Oxidoreductases / Q000235:genetics*; D017360:Arabidopsis / Q000145:classification / Q000201:enzymology / Q000235:genetics*; D001483:Base Sequence; D017931:DNA Primers; D020869:Gene Expression Profiling; D015971:Gene Expression Regulation, Enzymologic; D018506:Gene Expression Regulation, Plant; D005966:Glucuronidase / Q000235:genetics; D008031:Lignin / Q000378:metabolism; D005810:Multigene Family; D016296:Mutagenesis; D010802:Phylogeny; D030821:Plants, Genetically Modified; D016133:Polymerase Chain Reaction
D017931:DNA Primers; D008031:Lignin; D000429:Alcohol Oxidoreductases; C018656:cinnamyl alcohol dehydrogenase; D005966:Glucuronidase