Transcriptome alterations following developmental atrazine exposure in zebrafish are associated with disruption of neuroendocrine and reproductive system function, cell cycle, and carcinogenesis.

Atrazine, a herbicide commonly applied to agricultural areas and a common contaminant of potable water supplies, is implicated as an endocrine-disrupting chemical (EDC) and potential carcinogen. Studies show that EDCs can cause irreversible changes in tissue formation, decreased reproductive potential, obesity, and cancer. The U.S. Environmental Protection Agency considers an atrazine concentration of ≤ 3 ppb in drinking water safe for consumption. The specific adverse human health effects associated with a developmental atrazine exposure and the underlying genetic mechanisms of these effects are not well defined. In this study, zebrafish embryos were exposed to a range of atrazine concentrations to establish toxicity. Morphological, transcriptomic, and protein alterations were then assessed at 72h postfertilization following developmental atrazine exposure at 0, 0.3, 3, or 30 ppb. A significant increase in head length was observed in all three atrazine treatments. Transcriptomic profiles revealed 21, 62, and 64 genes with altered expression in the 0.3, 3, and 30 ppb atrazine treatments, respectively. Altered genes were associated with neuroendocrine and reproductive system development, function, and disease; cell cycle control; and carcinogenesis. There was a significant overlap (42 genes) between the 3 and 30 ppb differentially expressed gene lists, with two of these genes (CYP17A1 and SAMHD1) present in all three atrazine treatments. Increased transcript levels were translated to significant upregulation in protein expression. Overall, this study identifies genetic and molecular targets altered in response to a developmental atrazine exposure to further define the biological pathways and mechanisms of toxicity.

Identification of copper-responsive genes in an early life stage of the fathead minnow Pimephales promelas.

While physiological changes associated with copper toxicity have been studied in adult fathead minnow, Pimephales promelas, little is known about the effect of copper on newly hatched larvae. As a result we initiated an investigation on the mechanism of copper toxicity in 24 h post-hatch larvae using gene expression changes to identify responsive genes. Fish were exposed to copper concentrations of 0, 50, 125 and 200 mug/L in a 48 h toxicity test. Total RNA from survivors was used in a differential display assay to screen for differentially expressed gene products. Altogether, 654 copper-responsive differentially expressed bands were collected. Database searches found homology for 261 sequences. One hundred and sixty-one bands were homologous to NCBI genes of known function, of which 69 were individual genes. The most abundant categories of functional genes responding to copper were involved in protein synthesis/translational machinery and contractile proteins. Twenty-one dose-responsive genes were measured for expression changes using real-time quantitative PCR. Differential gene expression was validated for 11 of 13 genes, when a 1.2 times qPCR difference between the copper and control samples was observed. Transcripts identified as titin, cytochrome b, fast muscle specific heavy myosin chain 4, fast muscle troponin I, proteasome 26S subunit and troponin T3a were induced over twofold. Differential display bands identified as 60S ribosomal proteins L27 and L12 were repressed approximately threefold. We conclude that copper exposure affects several cellular pathways in larval fathead minnows with protein synthesis, ribosome structure, and muscle contractile proteins being the most sensitive to this stress.

Sexual dimorphism in hepatic gene expression and the response to dietary carbohydrate manipulation in the zebrafish (Danio rerio).

In this study, we tested for the presence of sexual dimorphism in the hepatic transcriptome of the adult zebrafish and examined the effect of long term manipulation of dietary carbohydrate on gene expression in both sexes. Zebrafish were fed diets comprised of 0%, 15%, 25%, or 35% carbohydrate from the larval stage through sexual maturity, then sampled for hepatic tissue, growth, proximate body composition, and retention efficiencies. Using Affymetrix microarrays and qRT-PCR, we observed substantial sexual dimorphism in the hepatic transcriptome. Males up-regulated genes associated with oxidative metabolism, carbohydrate metabolism, energy production, and amelioration of oxidative stress, while females had higher expression levels of genes associated with translation. Restriction of dietary carbohydrate (0% diet) significantly affected hepatic gene expression, growth performance, retention efficiencies of protein and energy, and percentages of moisture, lipid, and ash. The response of some genes to dietary manipulation varied by sex; with increased dietary carbohydrate, males up-regulated genes associated with oxidative metabolism (e.g. hadhbeta) while females up-regulated genes associated with glucose phosphorylation (e.g. glucokinase). Our data support the use of the zebrafish model for the study of fish nutritional genomics, but highlight the importance of accounting for sexual dimorphism in these studies.