Page 273 - The Toxicology of Fishes

Receptor-Mediated Mechanisms of Toxicity 253

expression profiling has also provided clues to the mechanism by which TCDD inhibits fin regeneration
in zebrafish. In addition to induction of genes encoding xenobiotic-metabolizing enzymes, regenerating
fins of zebrafish exposed to TCDD displayed dramatically reduced expression of genes encoding extra-
cellular matrix components and of sox9b, which may be an important regulator of the initial stages of
regeneration (Andreasen et al., 2006). Together, these examples illustrate the emerging value of gene
expression profiling in mechanistic toxicology.
In contrast to the rapidly increasing application of microarrays in fish toxicology, the use of proteomic
techniques has lagged until recently (Bosworth et al., 2005; Denslow et al., 2005; Knoll-Gellida et al.,
2006; Link et al., 2006a,b; Walker et al., 2007). As with DNA microarrays, proteomic studies have the
power to illuminate mechanisms as well as to identify markers of exposure or effect in a way that
complements RNA- and DNA-based methods. Proteomics and metabolomics also can be used to com-
plement histopathological analyses, in a phenotypic anchoring approach that parallels that described
above for microarrays (Stentiford et al., 2005; Ward et al., 2006).

The Aryl Hydrocarbon Receptor Signaling Pathway

Of all the receptors that are known as targets of environmental chemicals in fishes, two have been studied
in greatest detail: aryl hydrocarbon receptors and estrogen receptors. Here, to illustrate the role of
receptors in fish toxicology, we provide an overview of the AhR signaling pathway, AhR diversity, and
studies demonstrating the mechanistic role of AhRs in the toxicity of planar halogenated aromatic
hydrocarbons (PHAHs, or dioxin-like compounds) and polynuclear aromatic hydrocarbons (PAHs) in
fishes. Additional details can be found in recent reviews of the AhR pathway in mammals (Ma, 2001;
Nebert et al., 2004; Puga et al., 2005; Schmidt and Bradfield, 1996) and fishes (Hahn, 2002; Hahn et
al., 2005, 2006a; Tanguay et al., 2003).
Much of what we know about the AhR and its associated signal transduction pathway has come from
studies in mammalian systems—primarily murine and human cells and tissues. Although not as extensive,
studies in fishes have shown that the essential features of AhR signaling in mammals are conserved
(Pollenz and Necela, 1998; Pollenz et al., 2002; Wentworth et al., 2004). AhR proteins are localized
primarily in the cytoplasm of cells, in association with Hsp90 and other proteins. Upon binding of ligands,
AhR proteins are preferentially translocated to (or retained in) the nucleus, where they form dimers with
AhR nuclear translocator (ARNT) proteins. The ligand–AhR–ARNT complex interacts with AhR response
elements (AhREs; also known as XREs or DREs) to activate or repress gene expression from target genes.
Diagrams of the AhR pathway can be found in recent publications (Hahn et al., 2005, 2006a).
Fish AhRs appear to have ligand structure–activity relationships similar (but not identical) to those of
mammalian receptors, with high-affinity binding of TCDD, non-ortho-substituted PCBs, and some PAHs
and lower affinity binding of other halogenated and nonhalogenated ligands such as indoles (Abnet et
al., 1999b; Hestermann et al., 2000, unpublished results). Similarly, the AhR recognition sequence
(AhRE) in fish is similar to that of mammals, as suggested by the ability of fish AhRs to recognize
mammalian AhREs. Overlap also occurs in the identity of at least some AhR target genes in fish and
mammals. CYP1A, CYP1B, and certain other genes encoding xenobiotic-metabolizing enzymes are
inducible by TCDD in both groups (Stegeman and Hahn, 1994). Also inducible in mammals and fish is
the AhR repressor (AhRR) gene, which encodes a repressor of AhR function (Evans et al., 2005; Karchner
et al., 2002; Mimura et al., 1999; Roy et al., 2006).
Despite the mechanistic similarities, the piscine AhR pathway differs from that of mammals in some
fundamental ways. One key distinction is in the number of AhR isoforms. Mammals have a single AhR,
whereas most fish possess multiple (two to six) distinct AhR genes (Hahn, 2002; Hahn et al., 2006a).
Phylogenetic analysis of teleost AhRs shows that there are two types, AhR1 and AhR2, which arose by
a gene duplication occurring early in vertebrate evolution, prior to the divergence of fish and mammalian
lineages (Hahn et al., 1997; Karchner et al., 1999). Fish AhR1 forms are orthologous (or co-orthologous)
to mammalian AhRs, whereas AhR2 forms are found in fish (and birds) but not in mammals. Fish AhR1
and AhR2 have different tissue-specific patterns of expression, suggesting different functional roles. This