A cDNA encoding S-adenosyl-L-methionine:caffeic acid 3-O-methyltransferase from Eucalyptus.

In angiosperms, OMTs (EC 2.1.1.6) are bispecific and catalyze the methylation of caffeic acid to ferulic acid and 5hydroxy-ferulic acid to sinapic acid using S-adenosyl-L-Met as methyl donor. The resulting products, ferulic acid and sinapic acid, may subsequently be converted to the corresponding alcohols that are incorporated in lignins. In gymnosperms, where syringyl lignins are absent, OMTs are monospecific for caffeic acid. Because of their differential substrate specificity, OMTs are believed to play a role in the regulation of the monomeric composition of lignins and therefore are considered as target genes for manipulating lignins through recombinant DNA technology. We have cloned an OMT cDNA from eucalyptus, a woody angiosperm of crucial importance in the pulp industry. A cDNA library from young, developing xylem tissue of Eucalyptus gunnii, constructed in the vector XZapII (Stratagene), was screened with an OMT I cDNA clone from tobacco (Jaeck et al., 1992). A full-length cDNA clone, designated pEuOMT, was isolated containing a continuous open reading frame of 1098 nucleotides encoding a polypeptide of 366 amino acid residues (Table I). The eucalyptus OMT sequence is highly homologous (identity/similarity) at the amino acid leve1 with caffeic acid/5hydroxyferulic O-methyltransferases from other angiosperms: 84.4/9.9% with aspen (Bugos et al., 1991) and poplar (Dumas et al., 1992) OMTs, 80.8/10.7%, 76.4/11.8%, 62.6/ 15.1% with, respectively, alfafa (Gowri et al., 1991), tobacco (Jaeck et al., 1992), and maize (Collazo et al., 1992) OMTs. A high percentage of homology (50.4% identity, 22.5% similarity) is also found with a myo-inositol O-methylhansferase induced by osmotic stress in the facultative halophyte Mesenbryanthemum cystallinum (Vernon and Bohnert, 1992). The alignment of a11 of these OMT sequences with ZRP4, an mRNA putatively coding for an OMT in maize roots (Held et al., 1993), reveals a significant homology (30% identity, 20% similarity) over the entire amino acid sequence. The highest amino acid identity is found in the carboxy half of the protein containing the three regions previously shown to occur in many enzymes requiring S-adenosyl-L-Met as a substrate (Bugos et al., 1991). Surprisingly, no extended ho-