Molecular characterisation of Ltchi7, a gene encoding a Class III endochitinase induced by drought stress in Lotus spp

Abstract

Chitinases are enzymes that digest chitin molecules, present principally in insects and fungi. In plants, these enzymes play an important role in defence against pathogen attack, although they have also been described as induced by mechanical damage, ozone, heavy metals, cold, salinity, etc. Using an annealing control primer, we isolated a gene fragment whose translated sequence has high homology with a class III endochitinase. The gene, named Ltchi7, consisted of one ORF of 1005 bp, which codes for a peptide of 334 amino acids, including a deduced signal peptide of 27 amino acid that directs protein to the extracellular space. Phylogenetic analysis suggests that Ltchi7 is within a cluster that includes Sesbania rostrata, Medicago sativa and Glycine max class III endochitinases. This group is differentiated from other species of endochitinases by the presence of an additional extension in carboxy-terminal region. Moreover, in comparison with the majority of chitinases, Ltchi7 has two additional cysteine residues, which, according to 3D modelling studies, are very close. Gene expression analysis showed enhanced transcript abundance of this gene during drought stress in Lotus tenuis and Lotus japonicus, compared with growth under normal conditions. Furthermore, its expression is restricted to nodules and roots. Expression of this gene was also induced by salt stress, hydrogen peroxide and weakly with abscisic acid.