Project description:Omics approaches are broadly used to explore endocrine and toxicity-related pathways and functions. Nevertheless, there is still a significant gap in knowledge in terms of understanding the endocrine system and its numerous connections and intricate feedback loops, especially in non-model organisms. The fathead minnow (Pimephales promelas) is a widely used small fish model for aquatic toxicology and regulatory testing, particularly in North America. A draft genome has been published but the amount of available genomic or transcriptomic information is still far behind that of other more broadly studied species, such as the zebrafish. Here, we surveyed the tissue-specific proteome and transcriptome profiles in adult male fathead minnow. To do so, we generated a draft transcriptome using short and long sequencing reads. We also performed RNA sequencing and proteomics analysis on the telencephalon, hypothalamus, liver, and gut of male fish. The main purpose of this analysis was to generate tissue-specific omics data in order to support future aquatic ecotoxicogenomic and endocrine-related studies as well as to improve our understanding of the fathead minnow as an ecological model.
Project description:Fathead minnow and zebrafish are among the most intensively studied fish species in environmental toxicogenomics. To aid the assessment and interpretation of subtle transcriptomic effects from treatment conditions of interest, there needs to be a better characterization and understanding of the natural variation in gene expression among fish individuals within populations. Little effort, however, has been made in this area. Leveraging the transcriptomics data from a number of our toxicogenomics studies conducted over the years, we conducted a meta-analysis of nearly 600 microarrays generated from the ovary tissue of untreated, reproductively mature fathead minnow and zebrafish samples. As expected, there was considerable batch-to-batch transcriptomic variation; this “batch-effect” appeared to impact the fish transcriptomes randomly. The overall level of variation within-batch was quite low in fish ovary tissue, making it a suitable system for studying chemical stressors with subtle biological effects. The within-batch variation, however, differed considerably among individual genes and molecular pathways. This difference in variability is probably both technical and biological, thus suggesting a need to take into account both the expression levels and variance in evaluating and interpreting the transcriptional impact on genes and pathways by experimental conditions. There was significant conservation of both the genomes and transcriptomes between fathead minnow and zebrafish. The conservation to such a degree would enable not only a comparative biology approach in studying the mechanisms of action underlying environmental stressors, but also effective sharing of a large amount of existing public transcriptomics data for future development of toxicogenomics applications. total RNA from the ovary tissue of treated or control fish labeled in single color was hybridized to Agilent fathead minnow microarray (design 019597)
Project description:Production, usage and disposal of the munitions constituent (MC) cyclotrimethylenetrinitramine (RDX) has led to environmental releases on military facilities. The chemical attributes of RDX are conducive for leaching to surface water which may put aquatic organisms at risk of exposure. Because RDX has been observed to cause aberrant neuromuscular effects across a wide range of animal phyla, we assessed the effects of RDX on central nervous system (CNS) function in the representative aquatic ecotoxicological model species, fathead minnow (Pimephales promelas). A brain-tissue based cDNA library enriched for transcripts differentially expressed in response to RDX exposure was developed for fathead minnow and was transitioned to custom cDNA-based microarrays. All 4,128 cDNAs were sequenced, quality filtered and assembled yielding 3,018 unique sequences and 945 significant blastx matches (E ≤ 10-5). Bioassays were conducted exposing fathead minnows to RDX at 0.625, 1.25, 2.5, 5, 10 mg/L or an acetone-spike control for 10d. Overt toxicity of RDX in fathead minnow occurred only at the highest exposure concentration resulting in 50% mortality. Conversely, Bayesian analysis of microarray data indicated significant changes in transcript expression in fathead minnow brain tissue at RDX concentrations as low as 0.625 mg/L. In total, 154 microarray targets representing 44 unique transcript identities were differentially expressed in RDX exposures, the majority of which were validated by RT-qPCR. Investigation of molecular pathways, gene ontology and individual gene functions indicated that RDX exposures affected metabolic processes involved in: oxygen transport, neurological function, calcium binding / signaling, energy metabolism, cell cycle / cell proliferation, oxidative stress and ubiquitination. In total, our study indicated that RDX exposure affected molecular processes critical to CNS function in fathead minnow.