University of Nebraska - Lincoln University of Nebraska - Lincoln
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U.S. Environmental Protection Agency Papers U.S. Environmental Protection Agency
2007
FundulusFundulus
as the premier teleost model in environmental biology: as the premier teleost model in environmental biology:
Opportunities for new insights using genomics Opportunities for new insights using genomics
Karen G. Burnett
College of Charleston
, burnettk@cofc.edu
Lisa J. Bain
Clemson University
William S. Baldwin
Clemson University
Gloria V. Callard
Boston University
Sarah Cohen
San Francisco State University
See next page for additional authors
Follow this and additional works at: https://digitalcommons.unl.edu/usepapapers
Part of the Earth Sciences Commons, Environmental Health and Protection Commons, Environmental
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Burnett, Karen G.; Bain, Lisa J.; Baldwin, William S.; Callard, Gloria V.; Cohen, Sarah; Di Giulio, Richard T.;
Evans, David H.; Gómez-Chiarri, Marta; Hahn, Mark E.; Hoover, Cindi A.; Karchner, Sibel I.; Katoh, Fumi;
MacLatchy, Deborah L.; Marshall, William S.; Meyer, Joel N.; Nacci, Diane E.; Oleksiak, Marjorie F.; Rees,
Bernard B.; Singer, Thomas D.; Stegeman, John J.; Towle, David W.; Van Veld, Peter A.; Vogelbein,
Wolfgang K.; Whitehead, Andrew; Winn, Richard N.; and Crawford, Douglas L., "
Fundulus
as the premier
teleost model in environmental biology: Opportunities for new insights using genomics" (2007).
U.S.
Environmental Protection Agency Papers
. 98.
https://digitalcommons.unl.edu/usepapapers/98
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Authors Authors
Karen G. Burnett, Lisa J. Bain, William S. Baldwin, Gloria V. Callard, Sarah Cohen, Richard T. Di Giulio, David
H. Evans, Marta Gómez-Chiarri, Mark E. Hahn, Cindi A. Hoover, Sibel I. Karchner, Fumi Katoh, Deborah L.
MacLatchy, William S. Marshall, Joel N. Meyer, Diane E. Nacci, Marjorie F. Oleksiak, Bernard B. Rees,
Thomas D. Singer, John J. Stegeman, David W. Towle, Peter A. Van Veld, Wolfgang K. Vogelbein, Andrew
Whitehead, Richard N. Winn, and Douglas L. Crawford
This article is available at DigitalCommons@University of Nebraska - Lincoln: https://digitalcommons.unl.edu/
usepapapers/98
Review
Fundulus as the premier teleost model in environmental biology:
Opportunities for new insights using genomics
Karen G. Burnett
a,
⁎
, Lisa J. Bain
b
, William S. Baldwin
b
, Gloria V. Callard
c
, Sarah Cohen
d
,
Richard T. Di Giulio
e
, David H. Evans
f
, Marta Gómez-Chiarri
g
, Mark E. Hahn
h
,
Cindi A. Hoover
c
, Sibel I. Karchner
h
, Fumi Katoh
j
, Deborah L. MacLatchy
i
,
William S. Marshall
j
, Joel N. Meyer
e
, Diane E. Nacci
k
, Marjorie F. Oleksiak
l
, Bernard B. Rees
m
,
Thomas D. Singer
n
, John J. Stegeman
h
, David W. Towle
o
, Peter A. Van Veld
p
,
Wolfgang K. Vogelbein
p
, Andrew Whitehead
q
, Richard N. Winn
r
, Douglas L. Crawford
l
a
Grice Marine Laboratory, College of Charleston, Charleston, SC 29412, USA
b
Clemson Institute of Environmental Toxicology, Clemson University; Pendleton, SC 29670, USA
c
Department of Biology, Boston University, Boston, MA, USA
d
Romberg Tiburon Center and Department of Biology, San Francisco State University, Tiburon, CA 94120, USA
e
Nicholas School of the Environment and Earth Sciences, Duke University, Durham, NC, USA
f
Department of Zoology, University of Florida, Gainesville, FL 32611, USA
g
Department of Fisheries, Animal and Veterinary Science, University of Rhode Island, Kingston, RI 02881, USA
h
Biology Department, Woods Hole Oceanographic Institution, Woods Hole, MA 02543, USA
i
Faculty of Science, Wilfred Laurier University, Waterloo, Ontario, Canada N2L 3C5
j
Department of Biology, St. Francis Xavier University, Antigonish, N.S., Canada B2G 2W5
k
US Environmental Protection Agency Office of Research and Development, Narragansett, RI 02882, USA
l
Rosenstiel School of Marine & Atmospheric Science, University of Miami, Miami, FL 33149, USA
m
Department of Biological Sciences, University of New Orleans, New Orleans, LA 70148, USA
n
School of Optometry, University of Waterloo, Waterloo, ON, Canada N2L 3G1
o
Center for Marine Functional Genomics, Mount Desert Island Biological Laboratory, Salsbury Cove, Maine 04672, USA
p
The College of William and Mary, Virginia Institute of Marine Science, Gloucester Point, VA 23062, USA
q
Department of Biological Sciences, Louisiana State University, Baton Rouge, LA 70803, USA
r
Aquatic Biotechnology and Environmental Laboratory, University of Georgia, Athens, GA 30602, USA
Received 25 April 2007; received in revised form 31 August 2007; accepted 1 September 2007
Available online 8 September 2007
A
vailable online at www.sciencedirect.com
Comparative Biochemistry and Physiology, Part D 2 (2007) 257 – 286
www.elsevier.com/locate/cbpd
Abbreviations: AHR, Aryl hydrocarbon receptor; AHRR, AHR repressor; ANF, α-naphthoflavone; ARNT, AHR nuclear translocator; BAC, Bacterial artificial
chromosome; BAP, Benzo[a]pyrene; bHLH-PAS, Basic helix-loop-helix Per-ARNT-Sim; BNF, β naphthoflavone; CAR, Constitutive androstane receptor; CFTR,
Cystic fibrosis transmembrane conductance regulator; COX, Cyclooxygenase; CYP, Cytochrome P450; DDT, Dichloro-diphenyl-trichloroethane; DLC, Dioxin-like
compound; DO, Dissolved oxygen; ECOTOX, Environmental Protection Agency's database of aquatic toxicity testing; ENU, N-ethyl-N-nitrosourea; ER, Estrogen
receptor; ERR, Estrogen receptor-related receptor; EST, Expressed sequence tag; FGC, Fundulus Genomics Consortium; FXR, Farnesoid X-receptor; GO, Gene
Ontologies; GR, Glutocorticoid receptor; GST, Glutathione S-transferase; HAH, Halogenated aromatic hydrocarbon; HIF, Hypoxia-inducible factor; KEGG, Kyoto
Encyclopedia of Genes and Genomes; LDH, Lactate dehydrogenase; LXR, Liver X-receptor; MH, Major histocompatibility; MIS, Maturation induction steroid; MR,
Mineralocorticoid receptor; NBH, New Bedford Harbor; NCBI, National Center for Biotechnology Information; NR, Nuclear receptor; NKCC, Na
+
,K
+
,2Cl
−
cotransporter; NHE, Na
+
–H
+
-exchangers; P450arom, Cytochrome P450 aromatase; PAH, Polycyclic aromatic hydrocarbon; PCB, Polychlorinated biphenyl; P
crit
,
critical oxygen pressure; PO
2
, Oxygen partial pressure; PCR, Polymerase chain reaction; PXR, Pregnane X-receptor; SET protein, Phosphatase 2A inhibitor I2PP2A;
SNP, Single nucleotide polymorphism; TCDD, 2,3,7,8-tetrachlorodibenzo-p-dioxin; VDR, Vitamin D receptor.
⁎
Corresponding author. Tel.: +1 843 762 8933; fax: +1 843 762 8737.
E-mail address: burnettk@cofc.edu (K.G. Burnett).
1744-117X/$ - see front matter © 2007 Elsevier Inc. All rights reserved.
doi:10.1016/j.cbd.2007.09.001
This article is a U.S. government work, and is not subject to copyright in the United States.
Abstract
A strong foundation of basic and applied research documents that the estuarine fish Fundulus heteroclitus and related species are unique
laboratory and field models for understanding how individuals and populations interact with their environment. In this paper we summarize an
extensive body of work examining the adaptive responses of Fundulus species to environmental conditions, and describe how this research has
contributed importantly to our understanding of physiology, gene regulation, toxicology, and ecological and evolutionary genetics of teleosts and
other vertebrates. These explorations have reached a critical juncture at which advancement is hindered by the lack of genomic resources for these
species. We suggest that a more complete genomics toolbox for F. heteroclitus and related species will permit researchers to exploit the power of
this model organism to rapidly advance our understanding of fundamental biological and pathological mechanisms among vertebrates, as well as
ecological strategies and evolutionary processes common to all living organisms.
© 2007 Elsevier Inc. All rights reserved.
Keywords: Fundulus heteroclitus; Physiological genomics; Ecological genomics; Evolutionary genomics; Toxicogenomics; Environmental genomics
Contents
1. Introduction .............................................................. 259
2. Physiology............................................................... 259
2.1. Osmoregulation ........................................................ 259
2.1.1. Seawater ....................................................... 259
2.1.2. Freshwater ...................................................... 260
2.1.3. Kidney and intestine ................................................. 260
2.2. Responses to hypoxia ..................................................... 261
2.3. Temperature adaptations .................................................... 262
2.3.1. Ldh-B ......................................................... 262
2.3.2. Glycolysis ....................................................... 262
2.4. Reproduction and development ................................................ 263
2.5. Sensory systems ........................................................ 264
3. Gene regulation ............................................................ 264
3.1. Receptors and other ligand-activated transcription factors ................................... 265
3.1.1. Nuclear receptors ................................................... 265
3.1.2. Basic helix-loop-helix Per-Arnt-Sim family of proteins................................ 265
3.2. Cytochrome P450s ....................................................... 266
4. Toxicology ............................................................... 267
4.1. Reproductive and developmental effects ............................................ 267
4.2. Effects on immunocompetence ................................................ 268
4.3. Carcinogenesis ......................................................... 268
4.4. Toxicant-altered gene expression ................................................ 269
5. Ecological and evolutionary genetics ................................................. 269
5.1. Evolved tolerance to polluted habitats ............................................. 270
5.1.1. Tolerance to mercury and TCDD (Newark, NJ, USA) ................................ 270
5.1.2. Tolerance to PCBs (New Bedford Harbor, MA, USA) ................................ 270
5.1.3. Tolerance to PAHs (Elizabeth River, VA, USA) ................................... 271
5.2. Evolution of host–parasite interactions ............................................ 272
5.3. Evolutionary variation in glycolytic enzyme expression .................................... 272
5.4. Evolutionary variation in messenger RNA expression ..................................... 273
5.5. Adaptive patterns of DNA variation .............................................. 273
6. Fundulus genomics: current status .................................................. 274
6.1. ESTs .............................................................. 274
6.2. Microarrays .......................................................... 274
6.3. Genomes ............................................................ 275
6.4. Natural and constructed genetic modifications ......................................... 276
6.4.1. Transgenics ...................................................... 276
6.4.2. Clones ........................................................ 276
6.4.3. Morpholinos ..................................................... 276
6.4.4. Targeted mutagenesis: knockout mutants ....................................... 276
7. Summary ............................................................... 276
Acknowledgements ............................................................. 277
References ................................................................. 277
258 K.G. Burnett et al. / Comparative Biochemistry and Physiology, Part D 2 (2007) 257–286
1. Introduction
Fundulus is a diverse and widespread genus of small teleost
fishes, with species inhabiting coastal marshes from New
Brunswick and the north shore of the St. Lawrence to Florida
and the Gulf Coast from Florida to Texas, as well as inland
systems of North America (Bigelow and Schroeder, 1953; Lee
et al., 1980; Scott and Crossman, 1998). Fundulus heteroclitus,
also called the killifish, mud minnow or mummichog, is one of the
most abundant intertidal marsh fishes along the east coast of
North America, where they play a dominant role as both piscivore
and prey for a variety of birds, fishes and invertebrates (reviewed
in Able, 2002; Able et al., 2007; Kimball and Able, 2007).
F. heteroclitus is non-migratory (Skinner et al., 2005), with
local sub-populations exhibiting summer home ranges on the
order of 30–40 m (Lotrich, 1975) and greatly restricted winter
movements (Fritz et al., 1975). This broad distribution and limited
home range have made F. heteroclitus a powerful field model for
examining biological and ecological responses to natural
environmental changes, such as the wide variations in salinity,
oxygen, pH, and temperature, that routinely occur in estuarine
ecosystems. F. heteroclitus thrive in highly populated coastal
areas and chemically-polluted sites where they have evolved
mechanisms to tolerate some toxic chemicals. Abundance and
accessibility make F. heteroclitus easy to collect; their small size
and adaptability to a range of environmental conditions make
them easy to maintain in the laboratory. All life stages are hardy
and amenable to experimental manipulation. These attributes
make F. heteroclitus a valuable laboratory model for the study of
physiological processes such as osmoregulation and reproduction
in aquatic vertebrates. Consequently, these organisms are used
both in laboratory and field studies to examine basic disease
processes and toxicological mechanisms, as well as ecological
responses associated with chemical pollutants and other anthro-
pogenic stressors. Finally, the many related species of Fundulus
distributed over great geographic distances provide a powerful
framework for investigating fundamental ecological and evolu-
tionary processes.
This paper reviews selected key advances in physiology,
gene regulation, toxicology, and evolutionary genetics that have
relied on the distinctive, often unique , attributes of Fundulidae.
Given the scope of research that relies on Fundulus models, this
review is not comprehensive; rather, emphasis is placed on
research in which genomics-based technologies have enhanced
our knowledge of fundamental biological processes. Within this
perspective, we summarize genomics reso urces that are
currently available for F. heteroclitus. Finally, we argue that
further dev elopment o f such res ources will p osition F.
heteroclitus and related species as a powerful model system
for testing hypotheses regarding biological responses to
environmental change across levels of biological organization
from molecules to ecosystems.
2. Physiology
Physiological studies in F. heteroclitus have been aided by
the species tolerance to a range of abiotic factors, ease of
capture, and adaptation to laboratory conditions. As described
here, research on F. heteroclitus has significantly increased our
fundamental knowledge on the mechanisms by which fish adapt
to key environmental challenges such as changes in salinity
(2.1), oxygen levels (2.2) and temperature (2.3). In addition,
they have been used to better understand selected integrated
systems, such as reproduction and development (2.4) and vision
(2.5), among other physiological processes. Many of these
studies have led to important new knowledge regarding gene
regulation (3.0) and respon ses to toxicants (4.0).
2.1. Osmoregulation
F. heteroclitus is renowned for its euryhaline capabilities,
readily adapting to environments ranging from ion-poor to
hypersaline conditions as high as 120‰ (Griffith, 1974). Based
upon this attribute, F. heteroclitus has been and continues to be
an important model organism for understanding mechanisms of
teleost osmoregulation, as documented in two major reviews:
Karnaky (1986) focused on chloride cell structure and function
and Wood and Marshall (1994) compared in vitro and in vivo
approaches to und erstandin g euryhalinity in this speci es.
Current models for teleost ion transport and acid–base
regulation in gills (Evans et al., 2004, 2005) and other major
osmoregulatory organs (Marshall and Grosell, 2005) also rely
extensively on research conducted with F. heteroclitus.
2.1.1. Seawater
Keys and Willmer (1932) identified a morphologically-
distinct cell type in eel gills that was rich in mitochondria that
they reasoned was responsible for NaCl secretion by marine
teleost gills. Philpott and Copeland (1963) recogni zed a field of
these cells in the gills, skin and buccal epithelium of F.
heteroclitus and described the curious ultrastructure, with a
hugely expanded basolateral membrane surface in serpentine
tubules that formed a mesh among the well-organized
mitochondria. Na
+
,K
+
-ATPase, localized specifically on the
basolateral membrane of these “chloride cells” (Karnaky et al.,
1976), displayed higher activity in the gills of F. heteroclitus
adapted to seawater than to fresh water, and higher in both
conditions than in fish adapted to brackish water resembling the
ionic composition of the blood (Epstein et al., 1967; Towle et al.,
1977). These observations of Na
+
,K
+
-ATPase activity explained
the transepithelial secretion of Na
+
, but not Cl
−
exit from the
animal into seawater (Silva et al., 1977).
An explanation for the movement of Cl
−
against both
electrical and concentration gradients, the “pump-leak” path-
way, relied on two additional discoveries: a transport er that used
the Na
+
gradient to drive Cl
−
accumulation intracellularly and
an anion channel in the apical membrane that allowed the Cl
−
to
leak out into the environment. Using the patch-clamp technique,
Marshall et al. (1993) showed that the F. heteroclitus chloride
cell has a channel that is similar to the mammalian cystic
fibrosis transmembrane conductance regulator (CFTR). Singer
et al. (1998) cloned this CFTR homolog in F. heteroclitus,
showing that its expression increased after transfer to seawater,
and that cAMP activated anion conductance by the channel
259K.G. Burnett et al. / Comparative Biochemistry and Physiology, Part D 2 (2007) 257–286