Page 373 - The Toxicology of Fishes

Liver Toxicity 353

hepatic functions. With certain large gaps in information, a qualitatively similar story is unfolding in
fish. Importantly, livers of oviparous species have an additional hepatic function as an integral component
of the reproductive axis. In response to estrogen, hepatocytes in these organisms synthesize vitellogenin,
a yolk precursor protein, and choriogenin, a component of the outer egg envelope. Thus, hepatic NRs
of fishes will likely perform key roles, as above, but we need to know more about the molecular
mechanisms of NR action, their roles in critical liver functions, and the molecular mechanisms associated
with gene regulation.

Mammalian Liver Nuclear Receptors
Nuclear receptors are ligand-dependent transcription factors that bind lipophilic signaling molecules and
result in the control and expression of target genes. Mammalian NRs facilitate cellular responses to
endogenous and exogenous ligands by coordinating complex transcriptional responses (Mangelsdorf and
Evans, 1995). The NR superfamily includes receptors for multiple endobiotics, such as steroid hormones,
retinoids, thyroid hormone, vitamin D, bile acids, and prostaglandins. Also of importance are exogenous
ligands, including dietary components and xenobiotics.
Nuclear receptors commonly share a conserved N-terminal DNA-binding domain (DBD) and a
C-terminal ligand-binding domain (LBD). The DBD contains two zinc-finger motifs that form a single
structural domain containing an α-helix reading head that controls specific DNA sequence recognition
(Lin and White, 2004). The LBD confers ligand specificity and also contains a ligand-inducible trans-
activation function (AF2) essential for transcriptional activation. In the absence of a ligand, NRs are
associated with a nuclear receptor corepressor complex, resulting in inhibition of the basal transcription
activity of the associated promoter (Mangelsdorf and Evans, 1995; Ordentlich et al., 2001). Corepressor
proteins (SMART, NCoR) couple non-liganded, DNA-bound NR to enzymes with histone deacetylase
activity, resulting in chromatin condensation and a subsequent repression of gene expression (Polly et
al., 2000). Ligand binding causes a conformational change within the carboxy-terminal LBD (Rochel
et al., 2000), resulting in the release of corepressors and facilitating protein–protein interactions with
dimerization partners, coactivators, and mediator proteins (MEDs) (Glass and Rosenfeld, 2000). Coac-
tivators couple ligand-activated NRs to enzymes displaying histone acetyltransferase activity, thereby
facilitating chromatin remodeling. In subsequent steps, ligand-activated NRs interact with additional
MED family members to form a bridge to the RNA polymerase II (Pol II), resulting in transcription
activation. NRs bind to DNA as homodimers or heterodimers (usually with RXR) to hexameric response
elements (HREs) (5′-AG(G/T)TCA-3′), arranged as direct repeats (DRx), inverted repeats (IRx), or
everted repeats (ERx) separated by “x” base pairs.
The NR superfamily in mammals is composed of approximately 50 functional genes, 48 in humans,
47 in rats, and 49 in mice (Zhang et al., 2004). Interestingly, teleost fishes have a larger complement of
NR genes (68 in pufferfish) due to whole genome duplication events (Maglich et al., 2003). The current
official nomenclature for NRs divides the superfamily into seven families (NR0–6), with further subclas-
sification designated by alphanumeric characters representing sequence similarities (Bertrand et al., 2004).

Fish Nuclear Receptors with an Emphasis on Teleosts
Genomics efforts in pufferfish, zebrafish, and medaka have revealed orthologs for most mammalian NRs,
including those for steroid hormones, orphan receptors, and members of all seven NR families. In fact,
most NRs from teleosts, some cartilaginous fishes, and other lower vertebrates exhibit strong homologies
to mammalian sequences; however, certain structural modifications suggest functional differences that
may reflect adaptations arising with the movement of vertebrates from aquatic to terrestrial environments.
In addition, genome duplication resulted in a greater number of NRs in teleosts than in most mammals
(Venkatesh, 2003; Volff, 2005); for example, those NRs for which duplicate genes exist include the
vitamin D receptor (VDR), farnesol X receptor (FXR), glucocorticoid receptor (GR), and estrogen
receptor (ER), among others (Maglich et al., 2003). As a result, questions regarding molecular function
and diversification of paralogus sequences remain. Gene duplication events have different outcomes.
Formation of a nonfunctional, duplicate gene may result. Subfunctionalization of duplicated genes may
occur followed by degenerate mutations that modify expression or activity patterns of the single ancestral