Page 218 - The Toxicology of Fishes

198 The Toxicology of Fishes

1000bp
1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

GST –A2P GST–A1 GST–A
3 genes of 6 exons, spanning 9.5kb
FIGURE 4.15 Structure of the plaice Rho-class gene complex.

Many laboratory studies of GST induction in fish have typically been conducted in conjunction with
studies of CYP1A induction and have therefore involved AhR inducers, such as 3-methylcholanthrene
(3-MC) and β-naphthoflavone (BNF) (George, 1994). Studies of GST induction in rodents indicate that
antioxidant compounds such as ethoxyquin and butylated hydroxyanisole (BHA) are more effective
inducers of GST expression than are AhR agonists (Buetler et al., 1995; Hayes and Pulford, 1995). This
may not be the case in fish; for example, the level of induction in GST–CDNB activity in bullhead liver
(1.6-fold) by ethoxyquin is consistent with previous studies of GST induction by other agents in other
fish species. As an example, approximately twofold inductions of GST–CDNB activity have been
reported in trout exposed to BNF (Celander et al., 1993) and in plaice (Pleuronectes platessa) exposed
to BHA or trans-stilbene oxide (Leaver et al., 1992). Equivocable results have been obtained in studies
on the induction of fish GSTs by xenobiotics, partially due to their high constitutive levels and large
inter-animal variations in activities in wild populations, but primarily due to the use of substrates (e.g.,
HNE, ETHA) that are conjugated by several isoforms or the general substrate CDNB.
Promoter sequences targeted by prototypical GST antioxidant-inducing agents such as BHA not only
include antioxidant response elements (AREs) but also electrophile response elements (EpREs), which
confer inducibility by the monofunctional phenolic antioxidants to several rat GST and other phase II
genes (Li and Jaiswal, 1993; Rushmore et al., 1991). The growing body of studies of GST induction in
fish suggests that ARE-mediated GST gene induction may be conserved among many fish species. The
GSTA1 gene (Figure 4.15) contains two peroxisomal proliferator response elements (PPREs) that coor-
dinately upregulate expression of both GSTA and GSTA1 after exposure to peroxisomal proliferators
(hyperlipidemic drugs, but potentially also environmental contaminants with binding ability such as
synthetic lubricants, phthalate ester plasticizers, PCBs, certain pesticides, and alkylphenols). One of these
PPREs is associated with an estrogen response element (ERE) that may produce cross-talk and thus
explain the sex-dependent differences in expression that are observed. The GSTA gene contains four
upstream antioxidant response elements that upregulate expression by compounds which in vivo include
β-naphthoflavone (or probably a metabolite) and trans-stilbene oxide (Leaver et al., 1993). Two of these
AREs are associated with CCAAT boxes, which may confer tissue specificity of response (Figure 4.16).
Attempts have been made to utilize GST induction in several invertebrates as biomarkers of xenobiotic
exposure; indeed, the pi-class enzymes of Mollusca and Crustacea do show small (one- to twofold)
increases in CDNB conjugating activity to many compounds, especially pesticides (Table 4.16). Several
studies have also reported increased fish GST activities in fish inhabiting polluted environments
(Armknecht et al., 1998; Otto and Moon, 1996). Accordingly, induction of GST activity has been
proposed as a biomarker of exposure to environmental pollutants under field conditions. Again, careful
characterization of the inducibility of individual fish GST isozymes under laboratory conditions and
with respect to physiological status is necessary to interpret field data of GST expression.

GST A
CAAT CAAT
ARE ARE ARE ARE
5ʼ
ERE ERE–like GST A1
PPRE PPRE ERE–like
(RXRE)
FIGURE 4.16 Promoter regions of the plaice Rho-class genes.