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Developmental Expression and Substrate Specificities of Alfalfa Caffeic Acid 3-O-Methyltransferase and Caffeoyl Coenzyme A 3-O-Methyltransferase in Relation to Lignification1

Kentaro Inoue2, Vincent J.H. Sewalt2, 3, G. Murray Ballance4, Weiting Ni5, Cornelia Stürzer6, and Richard A. Dixon*

Plant Biology Division, Samuel Roberts Noble Foundation, 2510 Sam Noble Parkway, Ardmore, Oklahoma 73401

The biosynthesis of monolignols can potentially occur via two parallel pathways involving free acids or their coenzyme A (CoA) esters. Caffeic acid 3-O-methyltransferase (COMT) and caffeoyl CoA 3-O-methyltransferase (CCOMT) catalyze functionally identical reactions in these two pathways, resulting in the formation of mono- or dimethoxylated lignin precursors. The activities of the two enzymes increase from the first to the sixth internode in stems of alfalfa (Medicago sativa L.), preceding the deposition of lignin. Alfalfa CCOMT is highly similar at the amino acid sequence level to the CCOMT from parsley, although it contains a six-amino acid insertion near the N terminus. Transcripts encoding both COMT and CCOMT are primarily localized to vascular tissue in alfalfa stems. Alfalfa CCOMT expressed in Escherichia coli catalyzes O-methylation of caffeoyl and 5-hydroxyferuloyl CoA, with preference for caffeoyl CoA. It has low activity against the free acids. COMT expressed in E. coli is active against both caffeic and 5-hydroxyferulic acids, with preference for the latter compound. Surprisingly, very little extractable O-methyltransferase activity versus 5-hydroxyferuloyl CoA is present in alfalfa stem internodes, in which relative O-methyltransferase activity against 5-hy-droxyferulic acid increases with increasing maturity, correlating with increased lignin methoxyl content.

1   This work was supported by the Samuel Roberts Noble Foundation.
2   These authors contributed equally to this work.
3   Present address: Research Center, Pioneer Hi-Bred International, 7300 N.W. 62nd Avenue, Johnston, IA 50131.
4   Permanent address: Department of Plant Science, University of Manitoba, Winnipeg, Manitoba, Canada R3T 2N2.
5   Present address: U.S. Department of Agriculture-Agricultural Research Service, Department of Agronomy and Genetics, University of Minnesota, St. Paul, MN 55108.
6   Present address: Technical University Carolo Wilhelmina, Braunschweig, Germany.
*   Corresponding author; e-mail radixon{at}noble.org; fax 1-580-221-7380.

Plant Physiol. (1998) 117: 761-770
Copyright Clearance Center:   0032-0889/98/117/0761/10
© 1998 American Society of Plant Physiologists




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