Page 310 - The Toxicology of Fishes

290 The Toxicology of Fishes

Cell membrane
Dietary antioxidants
Prooxidant xenobiotics
Extracellular ROS Mitochondrial electron transport chain
Microsomal electron transport chain
Phase I metabolites
Intracellular ROS

Intracellular ROS and sulfhydryl-
reactive compounds
Nuclear membrane
PKC MAPK PI3K
P
P
SH SH GST, QR1, GCL, HO-1
Nrf2:small Maf c-jun, Keap1, others
Nrf2:small Maf
Keap1:Nrf2
SH
ARE
Actin filament

FIGURE 6.4 A working model of ARE-related signaling and gene regulation in mammals, based on references cited in
the text. Extracellular and intracellular ROS from any of a number of sources either act directly on the actin-bound Keap1
protein, oxidizing sulfhydryl groups and causing release of Nrf2, or act indirectly via kinase signaling cascades to phos-
phorylate Nrf2, leading to dissociation of Nrf2 from Keap1 and protection of Nrf2 from proteasomic degradation. Free
Nrf2 translocates to the nucleus, heterodimerizes with another protein such as c-Jun or a small Maf protein, and binds to
the ARE. This binding facilitates transcription of ARE-driven genes, such as those listed (see also Table 6.1). Note that
upregulation of c-jun and Keap1, if they occur, may result in positive or negative (respectively) feedback regulation of
ARE-mediated gene induction.

platessa) (Leaver and George, 1996; Leaver et al., 1997), and the expression of that gene was inducible
by treatment with trans-stilbene oxide, β-naphthoflavone, and perfluorooctanoic acid (a peroxisome-
proliferating agent) (Leaver et al., 1993, 1997). Three additional lines of evidence support the possi-
bility that some piscine genes are regulated by ARE sequences. First, some of the genes known or
hypothesized to be induced by electrophiles in mammals via the ARE have been shown to be inducible
by electrophiles in fish species at the levels of mRNA, protein, and catalytic activity. Some examples
are presented in Table 6.2. One complication with interpreting these data is the fact that the isoforms
of these genes present in fish have not been fully described, making it difficult to know whether the
genes seen to be inducible in fish correspond to those known to be ARE regulated in mammals,
particularly in the case of catalytic assays. A second line of evidence supporting the likelihood of
ARE regulation of endogenous gene expression in fish comes from studies demonstrating the func-
tionality of reporter genes driven by mammalian ARE promoter regions (murine GSTA1, murine QR1,
and possibly human QR1) in zebrafish and topminnow cells exposed to a variety of prooxidant
chemicals (Carvan et al., 2000, 2001; Rau et al., 2004). A final, and particularly convincing, line of
evidence is the demonstration that Nrf2 and Keap1 are present in zebrafish and regulate the expression
of zebrafish genes known to be ARE regulated in mammals (GSTP, QR1, and GCLH) (Kobayashi et
al., 2002); therefore, the transcription factors necessary to drive ARE-mediated gene transcription in
mammals are present and functional in fish, are able to recognize a mammalian ARE sequence, and
regulate the expression of endogenous zebrafish genes that are electrophile inducible. These results,
although not quite definitive, very strongly suggest that ARE-regulated genes exist in fish as well as
mammals and that there is considerable evolutionary conservation of function of ARE-mediated gene
regulation between mammals and fish.