Dataset Information

Gene expression patterns in rainbow trout, Oncorhynchus mykiss, exposed to a suite of model toxicants.

ABSTRACT: The increased availability and use of DNA microarrays has allowed the characterization of gene expression patterns associated with exposure to different toxicants. An important question is whether toxicant induced changes in gene expression in fish are sufficiently diverse to allow for identification of specific modes of action and/or specific contaminants. In theory, each class of toxicant may generate a gene expression profile unique to its mode of toxic action. In this study, isogenic (cloned) rainbow trout Oncorhynchus mykiss were exposed to sublethal levels of a series of model toxicants with varying modes of action, including ethynylestradiol (xeno-estrogen), 2,2,4,4'-tetrabromodiphenyl ether (BDE-47, thyroid active), diquat (oxidant stressor), chromium VI, and benzo[a]pyrene (BaP) for a period of 1-3 weeks. An additional experiment measured trenbolone (anabolic steroid; model androgen) induced gene expression changes in sexually mature female trout. Following exposure, fish were euthanized, livers removed and RNA extracted. Fluorescently labeled cDNA were generated and hybridized against a commercially available Atlantic Salmon/Trout array (GRASP project, University of Victoria) spotted with 16,000 cDNA's. The slides were scanned to measure abundance of a given transcript in each sample relative to controls. Data were analyzed via Genespring (Silicon Genetics) to identify a list of up- and downregulated genes, as well as to determine gene clustering patterns that can be used as "expression signatures". The results indicate each toxicant exposure caused between 64 and 222 genes to be significantly altered in expression. Most genes exhibiting altered expression responded to only one of the toxicants and relatively few were co-expressed in multiple treatments. For example, BaP and Diquat, both of which exert toxicity via oxidative stress, upregulated 28 of the same genes, of over 100 genes altered by either treatment. Other genes associated with steroidogenesis, p450 and estrogen responsive genes appear to be useful for selectively identifying toxicant mode of action in fish, suggesting a link between gene expression profile and mode of toxicity. Our array results showed good agreement with quantitative real time polymerase chain reaction (qRT PCR), which demonstrates that the arrays are an accurate measure of gene expression. The specificity of the gene expression profile in response to a model toxicant, the link between genes with altered expression and mode of toxic action, and the consistency between array and qRT PCR results all suggest that cDNA microarrays have the potential to screen environmental contaminants for biomarkers and mode of toxic action.

SUBMITTER: Hook SE

PROVIDER: S-EPMC2494855 | biostudies-literature | 2006 May

REPOSITORIES: biostudies-literature

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Gene expression patterns in rainbow trout, Oncorhynchus mykiss, exposed to a suite of model toxicants.

Hook Sharon E SE Skillman Ann D AD Small Jack A JA Schultz Irvin R IR

Aquatic toxicology (Amsterdam, Netherlands) 20060220 4

The increased availability and use of DNA microarrays has allowed the characterization of gene expression patterns associated with exposure to different toxicants. An important question is whether toxicant induced changes in gene expression in fish are sufficiently diverse to allow for identification of specific modes of action and/or specific contaminants. In theory, each class of toxicant may generate a gene expression profile unique to its mode of toxic action. In this study, isogenic (cloned ...[more]

PMID: 16488489

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Project description:The pollution of the environment with microplastics has been recognized as an emerging threat worldwide. Due to an exponential increase in production of plastic over the last eight decades and its longevity in the environment, accumulating amounts of microplastic are polluting rivers, lakes and the ocean. Their entry pathways are diverse and still only incompletely understood. Since microplastics are usually defined smaller than 5 mm, it can be ingested by a wide range of aquatic organisms including teleost fish. There are different approaches to study the detrimental effects of pollutants on aquatic organisms. On the one hand, generic baseline parameters such as growth and mortality are regularly considered, often accompanied by established stress parameters such as cortisol, heat shock proteins or lipid oxidation. The conflicting findings to date suggest that these parameters might not be sensitive enough to indicate the physiological effects of environmentally relevant microplastic concentrations. For this reason, more sophisticated biological approaches could provide new insights into whether and how microplastics harm fish. To date, proteomic approaches have been used only sporadically when investigating the effects of microplastic exposure on aquatic organisms. So far, this approach has not been used to address potential microplastic impacts in fish. In the present study, a proteomic approach was trialed alongside established methods in an investigation of fish experiencing long-term exposure to environmentally relevant concentrations of microplastics. Two groups of rainbow trout (Oncorhynchus mykiss were exposed to microplastic concentrations and sizes currently encountered in wild fish and an increased concentration, expected to occur in the near future. These groups where compared to a control group maintained in MP free conditions. Five fish of each treatment were sampled at three time points (week 1, week 4, week 17). The experiments were performed in triplicates, resulting in 45 samples used in the proteomic analysis.

Project description:BackgroundGenetic maps characterizing the inheritance patterns of traits and markers have been developed for a wide range of species and used to study questions in biomedicine, agriculture, ecology and evolutionary biology. The status of rainbow trout genetic maps has progressed significantly over the last decade due to interest in this species in aquaculture and sport fisheries, and as a model research organism for studies related to carcinogenesis, toxicology, comparative immunology, disease ecology, physiology and nutrition. We constructed a second generation genetic map for rainbow trout using microsatellite markers to facilitate the identification of quantitative trait loci for traits affecting aquaculture production efficiency and the extraction of comparative information from the genome sequences of model fish species.ResultsA genetic map ordering 1124 microsatellite loci spanning a sex-averaged distance of 2927.10 cM (Kosambi) and having 2.6 cM resolution was constructed by genotyping 10 parents and 150 offspring from the National Center for Cool and Cold Water Aquaculture (NCCCWA) reference family mapping panel. Microsatellite markers, representing pairs of loci resulting from an evolutionarily recent whole genome duplication event, identified 180 duplicated regions within the rainbow trout genome. Microsatellites associated with genes through expressed sequence tags or bacterial artificial chromosomes produced comparative assignments with tetraodon, zebrafish, fugu, and medaka resulting in assignments of homology for 199 loci.ConclusionThe second generation NCCCWA genetic map provides an increased microsatellite marker density and quantifies differences in recombination rate between the sexes in outbred populations. It has the potential to integrate with cytogenetic and other physical maps, identifying paralogous regions of the rainbow trout genome arising from the evolutionarily recent genome duplication event, and anchoring a comparative map with the zebrafish, medaka, tetraodon, and fugu genomes. This resource will facilitate the identification of genes affecting traits of interest through fine mapping and positional cloning of candidate genes.

Project description:BackgroundMaternal transcripts are accumulated in the oocyte during oogenesis to provide for protein synthesis from oocyte maturation through early embryonic development, when nuclear transcription is silenced. The maternal mRNAs have short poly(A) tails after undergoing post-transcriptional processing necessary for stabilizing them for storage. The transcripts undergo cytoplasmic polyadenylation when they are to be translated. Transcriptome analyses comparing total mRNA and elongated poly(A) mRNA content among eggs of different quality can provide insight into molecular mechanisms affecting egg developmental competence in rainbow trout. The present study used RNA-seq to compare transcriptomes of unfertilized eggs of rainbow trout females yielding different eyeing rates, following rRNA removal and poly(A) retention for construction of the libraries.ResultsThe percentage of embryos to reach the 32-cell stage at 24 h post fertilization was significantly correlated to family eyeing rate, indicating that inviable embryos were developmentally compromised before zygotic genome activation. RNA sequencing identified 2 differentially expressed transcripts (DETs) from total mRNA sequencing comparing females with low-quality (< 5% eyeing), medium-quality (30-50% eyeing), and high-quality (> 80% eyeing) eggs. In contrast, RNA sequencing from poly(A) captured transcripts identified 945 DETs between low- and high-quality eggs, 1012 between low- and medium-quality eggs, and only 2 between medium- and high-quality eggs. The transcripts of mitochondrial genes were enriched with polyadenylated transcript sequencing and they were significantly reduced in low-quality eggs. Similarly, mitochondrial DNA was reduced in low-quality eggs compared with medium- and high-quality eggs. The functional gene analysis classified the 945 DETs between low- and high-quality eggs into 31 functional modules, many of which were related to ribosomal and mitochondrial functions. Other modules involved transcription, translation, cell division, apoptosis, and immune responses.ConclusionsOur results indicate that differences in egg quality may be derived from differences in maternal nuclear transcript activation and cytoplasmic polyadenylation before ovulation, as opposed to accumulation and storage of maternal nuclear transcripts during oogenesis. Transcriptome comparisons suggest low-quality eggs suffered from impaired oxidative phosphorylation and translation. The DETs identified in this study provide insight into developmental competence in rainbow trout eggs.

Dataset Information

Gene expression patterns in rainbow trout, Oncorhynchus mykiss, exposed to a suite of model toxicants.

Publications

Gene expression patterns in rainbow trout, Oncorhynchus mykiss, exposed to a suite of model toxicants.

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