Caffeoyl-coenzyme A 3-O-methyltransferase enzyme activity, protein and transcript accumulation in flax (Linum usitatissimum) stem during development

Flax (Linum usitatissimum) is a commercially important fiber crop in Europe. Lignification of its phloem fibers, although weak, causes a decrease in their commercial quality. In flax, fiber lignin mainly consists of guaiacyl (G) units in contrast to the mixed guaiacyl-syringyl (G-S) lignin type occurring in xylem fibers. G lignins are reported as more condensed polymers due to a higher frequency of 5-5 linkages, whereas the deposition of syringyl end groups in lignins increases the proportion of alky-aryl ether linkages as beta -O-4-bonds. The type of linkages within a lignin polymer depends on the methylation of either the 3-hydroxyl groups or both 3-OH and 5-OH groups, which is controlled by two enzymes: caffeate 3-O-methyttransferase (COMT) and caffeoyl-coenzyme A 3-O-methyltransferase (CCoAOMT). First, we measured the in vitro activity of both OMTs in the flax stem tissues during stem development. CCoAOMT activity varied in the same way as COMT, i.e. increased gradually with stem maturity, from the top to the bottom of the stem, was maximum at the flowering stage and was lower, but still scorable, in the outer fiber-bearing tissues than in the xylem cells. In a second step, we focused our studies on the characterization of CCoAOMT in order to understand the implication of this enzyme in the lignification of flax fiber cells. CCoAOMT activity appeared to be associated with the accumulation of an acidic 33-kDa polypeptide identified as a CCoAOMT after immunological cross-reactivity with a poplar CCoAOMT and microsequencing. The differential accumulation of the CCoAOMT protein was confirmed by immunolocalization on tissue prints and correlated with that of the transcripts, suggesting a transcriptional regulation of CCoAOMT in the flax stem.