Isolation and characterisation of laccase cDNAs from meristematic and stem tissues of ryegrass (¤Lolium perenne¤)

Plant laccases are believed to be involved in dehydrogenative polymerisation of lignin. We report here the first cloning of monocot laccases. Five different laccase-encoding cDNA sequences were identified from ryegrass (Lolium perenne); four from stem and one from meristematic tissue. Three cDNA's contained entire coding sequences; LpLAC5-4 and LpLAC5-6 from stem, and LpLAC2-1 from meristem. LpLAC5-4 encodes a polypeptide of 610 amino acids with 14 potential N-glycosylation sites, and a calculated pI value of 5.0. Lack of an N-terminal signal peptide in LpLAC5-4 suggests that the mature protein is located intracellularly. A genomic clone, gLpLAC5-4, has been isolated and represents the first exon-intron structure and promoter sequence of a plant laccase gene. LpLAC5-6 cDNA encodes a protein of 579 amino acids with an N-terminal signal peptide characteristic of secreted proteins, 13 potential N-glycosylation sites, and a pI value of 9.0. LpLAC2-1 was the only laccase found in a meristematic library. The cDNA encodes a protein of 578 amino acids with a predicted pI of 5.8, 9 putative N-glycosylation sites, and an N-terminal signal peptide. Ryegrass laccases are differentially expressed and are under tight transcriptional control. Sequence analyses have shown that the ryegrass genome encodes more than 25 different laccase genes. Plant laccases are distributed in different groups rather than being closely intraspecies related. High heterogeneity of plant laccases indicates that gene duplication events have occurred continuously before and after divergence of seedplants into gymnosperms, monocots, and dicots. (C) 2002 Elsevier Science Ireland Ltd. All rights reserved.