Cytosolic glutathione S-transferases ofOesophagostomum dentatum

Abstract

Oesophagostomum dentatum stages were investigated for glutathione S-transferase (GST) expression at the protein and mRNA levels. GST activity was detected in all stages (infectious and parasitic stages including third- and fourth-stage larvae of different ages as well as males and females) and could be dose-dependently inhibited with sulfobromophthalein (SBP). Addition of SBP to in vitro larval cultures reversibly inhibited development from third- to fourth-stage larvae. Two glutathione-affinity purified proteins (23 and 25 kDa) were detected in lysates of exsheathed third-stage larvae by SDS-PAGE. PCR-primers were designed based on peptide sequences and conserved GST sequences of other nematodes for complete cDNA sequences (621 and 624 nt) of 2 isoforms, Od-GST1 and Od-GST2, with 72% nucleotide similarity and 75% for the deduced proteins. Genomic sequences consisted of 7 exons and 6 introns spanning 1296 bp for Od-GST1 and 1579 and 1606 bp for Od-GST2. Quantitative real-time-PCR revealed considerably elevated levels of Od-GST1 in the early parasitic stages and slightly reduced levels of Od-GST2 in male worms. Both Od-GSTs were most similar to GST of Ancylostoma caninum (nucleotides: 73 and 70%; amino acids: 80 and 73%). The first three exons (75 amino acids) corresponded to a synthetic prostaglandin D2 synthase (53% similarity). O. dentatum GSTs might be involved in intrinsic metabolic pathways which could play a role both in nematode physiology and in host-parasite interactions.