Fundulus as the premier teleost model in environmental biology : opportunities for new insights using genomics

Author Posting. © Elsevier B.V., 2007. This is the author's version of the work. It is posted here by permission of Elsevier B.V. for personal use, not for redistribution. The definitive version was published in Comparative Biochemistry and Physiology Part D: Genomics and Proteomics 2 (2007): 257-286, doi:10.1016/j.cbd.2007.09.001.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.This material is based on work supported by grants from the National Science Foundation DBI-0420504 (LJB), OCE 0308777 (DLC, RNW, BBR), BES-0553523 (AW), IBN 0236494 (BBR), IOB-0519579 (DHE), IOB-0543860 (DWT), FSML-0533189 (SC); National Institute of Health NIEHS P42-ES007381(GVC, MEH), P42-ES10356 (RTD), ES011588 (MFO); and NCRR P20 RR-016463 (DWT); Natural Sciences and Engineering Research Council of Canada Discovery (DLM, TDS, WSM) and Collaborative Research and Development Programs (DLM); NOAA/National Sea Grant NA86RG0052 (LJB), NA16RG2273 (SIK, MEH,GVC, JJS); Environmental Protection Agency U91620701 (WSB), R82902201(SC) and EPA’s Office of Research and Development (DEN)

BMC Molecular Biology BioMed Central Research article

Characterization of housekeeping genes in zebrafish: male-female differences and effects of tissue type, developmental stage and chemical treatmen

Time course Analysis of Gene expression patterns in ZebrafIsh Eye during Optic Nerve Regeneration

It is well-established that neurons in the adult mammalian central nervous system (CNS) are terminally differentiated and, if injured, will be unable to regenerate their connections. In contrast to mammals, zebrafish and other teleosts display a robust neuroregenerative response. Following optic nerve crush (ONX), retinal ganglion cells (RGC) regrow their axons to synapse with topographically correct targets in the optic tectum, such that vision is restored in ~21 days. What accounts for these differences between teleostean and mammalian responses to neural injury is not fully understood. A time course analysis of global gene expression patterns in the zebrafish eye after ONX can help to elucidate cellular and molecular mechanisms that contribute to a successful neuroregeneration. To define different phases of regeneration after ONX, alpha tubulin 1 ( tuba1 ) and growth-associated protein 43 ( gap43 ), markers previously shown to correspond to morphophological events, were measured by real time quantitative PCR (qPCR). Microarray analysis was then performed at defined intervals (6 hours, 1, 4, 12, and 21 days) post-ONX and compared to SHAM. Results show that optic nerve damage induces multiple, phase-related transcriptional programs, with the maximum number of genes changed and highest fold-change occurring at 4 days. Several functional groups affected by optic nerve regeneration, including cell adhesion, apoptosis, cell cycle, energy metabolism, ion channel activity, and calcium signaling, were identified. Utilizing the whole eye allowed us to identify signaling contributions from the vitreous, immune and glial cells as well as the neural cells of the retina. Comparisons between our dataset and transcriptional profiles from other models of regeneration in zebrafish retina, heart and fin revealed a subset of commonly regulated transcripts, indicating shared mechanisms in different regenerating tissues. Knowledge of gene expression patterns in all components of the eye in a model of successful regeneration provides an entry point for functional analyses, and will help in devising hypotheses for testing normal and toxic regulatory factors

Characterization of housekeeping genes in zebrafish: male-female differences and effects of tissue type, developmental stage and chemical treatment

<p>Abstract</p> <p>Background</p> <p>Research using the zebrafish model has experienced a rapid growth in recent years. Although real-time reverse transcription PCR (QPCR), normalized to an internal reference ("housekeeping") gene, is a frequently used method for quantifying gene expression changes in zebrafish, many commonly used housekeeping genes are known to vary with experimental conditions. To identify housekeeping genes that are stably expressed under different experimental conditions, and thus suitable as normalizers for QPCR in zebrafish, the present study evaluated the expression of eight commonly used housekeeping genes as a function of stage and hormone/toxicant exposure during development, and by tissue type and sex in adult fish.</p> <p>Results</p> <p>QPCR analysis was used to quantify mRNA levels of <it>bactin1, tubulin alpha 1(tuba1), glyceraldehyde-3-phosphate dehydrogenase (gapdh), glucose-6-phosphate dehydrogenase (g6pd), TATA-box binding protein (tbp), beta-2-microglobulin (b2m), elongation factor 1 alpha (elfa)</it>, and <it>18s ribosomal RNA (18s) </it>during development (2 – 120 hr postfertilization, hpf); in different tissue types (brain, eye, liver, heart, muscle, gonads) of adult males and females; and after treatment of embryos/larvae (24 – 96 hpf) with commonly used vehicles for administration and agents that represent known environmental endocrine disruptors. All genes were found to have some degree of variability under the conditions tested here. Rank ordering of expression stability using geNorm analysis identified <it>18s</it>, <it>b2m</it>, and <it>elfa </it>as most stable during development and across tissue types, while <it>gapdh, tuba1</it>, and <it>tpb </it>were the most variable. Following chemical treatment, <it>tuba1, bactin1</it>, and <it>elfa </it>were the most stably expressed whereas <it>tbp, 18s</it>, and <it>b2m </it>were the least stable. Data also revealed sex differences that are gene- and tissue-specific, and treatment effects that are gene-, vehicle- and ligand-specific. When the accuracy of QPCR analysis was tested using different reference genes to measure suppression of <it>cyp19a1b </it>by an estrogen receptor antagonist and induction of <it>cyp1a </it>by an arylhydrocarbon receptor agonist, the direction and magnitude of effects with stable and unstable genes differed.</p> <p>Conclusion</p> <p>This study provides data that can be expected to aid zebrafish researchers in their initial choice of housekeeping genes for future studies, but underlines the importance of further validating housekeeping genes for each new experimental paradigm and fish species.</p

Multiple structurally distinct ERa mRNA variants in zebrafish are differentially expressed by tissue type, stage of development and estrogen exposure

It is well established that estrogen-like environmental chemicals interact with the ligand-binding site of estrogen receptors (ER) to disrupt transcriptional control of estrogen responsive targets. Here we investigate the possibility that estrogens also impact splicing decisions on estrogen responsive genes, such as that encoding ERα itself. Targeted PCR cloning was applied to identify six ERα mRNA variants in zebrafish. Sequencing revealed alternate use of transcription and translation start sites, multiple exon deletions, intron retention and alternate polyadenylation. As determined by quantitative (q)PCR, N-terminal mRNA variants predicting long (ERαL) and short (ERαS) isoforms were differentially expressed by tissue-type, sex, stage of development and estrogen exposure. Whereas ERαL mRNA was diffusely distributed in liver, brain, heart, eye, and gonads, ERαS mRNA was preferentially expressed in liver (female > male) and ovary. Neither ERαL nor ERαS transcripts varied significantly during development, but 17β-estradiol selectively increased accumulation of ERαS mRNA (~170-fold by 120 hpf), an effect mimicked by bisphenol-A and diethylstilbestrol. Significantly, a C-truncated variant (ERαS-Cx) lacking most of the ligand binding and AF-2 domains was transcribed exclusively from the short isoform promoter and was similar to ERαS in its tissue-, stage- and estrogen inducible expression. These results support the idea that promoter choice and alternative splicing of the esr1 gene of zebrafish are part of the autoregulatory mechanism by which estrogen modulates subsequent ERα expression, and further suggest that environmental estrogens could exert some of their toxic effects by altering the relative abundance of structurally and functionally distinct ERα isoforms.2.674 JCR (2013) Q3, 64/123 Endocrinology & metabolis

Brain Aromatase in Fishes: Historical Perspective and Comparative Approaches

The ability to convert androgens to estrogens is an ancient and highly conserved characteristic of the vertebrate brain but is lacking in amphioxus and all other invertebrates, indicating that a functional form of the enzyme first evolved in the brain ofNo data (2012)UE