Isolation and cloning of homologous glutathione S-transferase cDNAs from English sole and starry flounder liver

We are currently investigating hepatic glutathione S-transferase (GST) expr ession in English sole (Pleuronectes vetulus) and starry flounder (Platicht hys stellatus), two closely related benthic fish that exhibit striking diff erences in the prevalences of contaminant-associated liver neoplasia. Align ment of cDNA sequences encoding plaice (Pleuronectes platessa) liver GST (P LGST-A, a theta class GST, Leaver et al., 1993) and a GST-related lens crys tallins protein from octopus (Octopus vulgaris) revealed conserved regions for oligonucleotide primer design for polymerase chain reaction (PCR)-based studies of sole and flounder GSTs. Reverse-transcriptase-PCR (RT-PCR) anal ysis of sole and flounder hepatic total mRNA yielded 469 nucleotide cDNA fr agments that displayed extensive sequence homology (98%) to plaice GST-A. N orthern blotting analysis of English sole and starry flounder liver using t he PCR product from English sole as a probe detected a single band of appro ximately 1000 nucleotides that was highly expressed in English sole and in starry flounder liver. Biochemical and immunological studies using diagnost ic GST reference substrates and class specific polyclonal antibodies sugges ted little homology among English sole and starry flounder GSTs, and rodent alpha, mu, and pi class GSTs. Interestingly, English sole, the species exh ibiting a relatively high prevalence of contaminant-associated liver neopla sia, exhibited higher hepatic GST activity toward the environmental epoxide s aflatoxin B-1-8,9-epoxide (AFBO) and benzo[a]pyrene-trans-7,8-dihydrodiol -9,10-epoxide-anti (BPDE). BPDE, in particular, is a carcinogenic epoxide i ntermediate implicated as a possible causative agent in the etiology of the se hepatic lesions. In summary, our studies indicate that a GST related to non-mammalian theta class GST is conserved in marine flatfish. Further mech anistic studies are necessary to determine the role of GSTs in the observed susceptibility differences among English sole and starry flounder to sedim ent-associated carcinogens. (C) 1999 Elsevier Science B.V. All rights reser ved.