Gene set enrichment analysis of microarray data from Pimephales promelas (Rafinesque), a non-mammalian model organism

<p>Abstract</p> <p>Background</p> <p>Methods for gene-class testing, such as Gene Set Enrichment Analysis (GSEA), incorporate biological knowledge into the analysis and interpretation of microarray data by comparing gene expression patterns to pathways, systems and emergent phenotypes. However, to use GSEA to its full capability with non-mammalian model organisms, a microarray platform must be annotated with human gene symbols. Doing so enables the ability to relate a model organism's gene expression, in response to a given treatment, to potential human health consequences of that treatment. We enhanced the annotation of a microarray platform from a non-mammalian model organism, and then used the GSEA approach in a reanalysis of a study examining the biological significance of acute and chronic methylmercury exposure on liver tissue of fathead minnow (<it>Pimephales promelas</it>). Using GSEA, we tested the hypothesis that fathead livers, in response to methylmercury exposure, would exhibit gene expression patterns similar to diseased human livers.</p> <p>Results</p> <p>We describe an enhanced annotation of the fathead minnow microarray platform with human gene symbols. This resource is now compatible with the GSEA approach for gene-class testing. We confirmed that GSEA, using this enhanced microarray platform, is able to recover results consistent with a previous analysis of fathead minnow exposure to methylmercury using standard analytical approaches. Using GSEA to compare fathead gene expression profiles to human phenotypes, we also found that fathead methylmercury-treated livers exhibited expression profiles that are homologous to human systems & pathways and results in damage that is similar to those of human liver damage associated with hepatocellular carcinoma and hepatitis B.</p> <p>Conclusions</p> <p>This study describes a powerful resource for enabling the use of non-mammalian model organisms in the study of human health significance. Results of microarray gene expression studies involving fathead minnow, typically used for aquatic ecological toxicology studies, can now be used to generate hypotheses regarding consequences of contaminants and other stressors on humans. The same approach can be used with other model organisms with microarray platforms annotated in a similar manner.</p

Gene Expression Changes Related to Endocrine Function and Decline in Reproduction in Fathead Minnow (Pimephales promelas) after Dietary Methylmercury Exposure

BACKGROUND: Methylmercury (MeHg) is a known neurotoxic agent, but the mechanisms by which MeHg may act on reproductive pathways are relatively unknown. Several studies have indicated potential changes in hormone levels as well as declines in vertebrates with increasing dietary MeHg exposure. OBJECTIVES: The purpose of this study was to identify alterations in gene expression associated with MeHg exposure, specifically those associated with previously observed changes in reproduction and reproductive biomarkers. Fathead minnows, Pimephales promelas, were fed one of three diets that were similar to documented concentrations of MeHg in the diets of wild invertivorous and piscivorous fish. We used a commercial macroarray in conjunction with quantitative polymerase chain reaction to examine gene expression in fish in relation to exposure to these environmentally relevant doses of MeHg. RESULTS: Expression of genes commonly associated with endocrine disruption was altered with Hg exposure. Specifically, we observed a marked up-regulation in vitellogenin mRNA in individual Hg-exposed males and a significant decline in vitellogenin gene expression in female fish with increasing Hg concentrations. Other genes identified by the macroarray experiment included those associated with egg fertilization and development, sugar metabolism, apoptosis, and electron transport. We also observed differences in expression patterns between male and female fish not related to genes specifically associated with reproduction, indicating a potential physiological difference in the reaction of males and females to MeHg. CONCLUSION: Gene expression data may provide insight into the mechanisms by which MeHg affects reproduction in fish and indicate how MeHg differs in its effect from other heavy metals and endocrine-disrupting compounds

Transcriptome signatures of wastewater effluent exposure in larval zebrafish vary with seasonal mixture composition in an effluent-dominated stream

Wastewater treatment plant (WWTP) effluent-dominated streams provide critical habitat for aquatic and terrestrial organisms but also continually expose them to complex mixtures of pharmaceuticals that can potentially impair growth, behavior, and reproduction. Currently, few biomarkers are available that relate to pharmaceutical-specific mechanisms of action. In the experiment reported in this paper, zebrafish (Danio rerio) embryos at two developmental stages were exposed to water samples from three sampling sites (0.1 km upstream of the outfall, at the effluent outfall, and 0.1 km below the outfall) during base-flow conditions from two months (January and May) of a temperate-region effluent-dominated stream containing a complex mixture of pharmaceuticals and other contaminants of emerging concern. RNA-sequencing identified potential biological impacts and biomarkers of WWTP effluent exposure that extend past traditional markers of endocrine disruption. Transcriptomics revealed changes to a wide range of biological functions and pathways including cardiac, neurological, visual, metabolic, and signaling pathways. These transcriptomic changes varied by developmental stage and displayed sensitivity to variable chemical composition and concentration of effluent, thus indicating a need for stage-specific biomarkers. Some transcripts are known to be associated with genes related to pharmaceuticals that were present in the collected samples. Although traditional biomarkers of endocrine disruption were not enriched in either month, a high estrogenicity signal was detected upstream in May and implicates the presence of unidentified chemical inputs not captured by the targeted chemical analysis. This work reveals associations between bioeffects of exposure, stage of development, and the composition of chemical mixtures in effluent-dominated surface water. The work underscores the importance of measuring effects beyond the endocrine system when assessing the impact of bioactive chemicals in WWTP effluent and identifies a need for non-targeted chemical analysis when bioeffects are not explained by the targeted analysis

Mapping the Anthocyaninless (anl) Locus in Rapid-Cycling Brassica rapa (RBr) to Linkage Group R9

<p>Abstract</p> <p>Background</p> <p>Anthocyanins are flavonoid pigments that are responsible for purple coloration in the stems and leaves of a variety of plant species. <it>Anthocyaninless </it>(<it>anl</it>) mutants of <it>Brassica rapa </it>fail to produce anthocyanin pigments. In rapid-cycling <it>Brassica rapa</it>, also known as Wisconsin Fast Plants, the anthocyaninless trait, also called non-purple stem, is widely used as a model recessive trait for teaching genetics. Although anthocyanin genes have been mapped in other plants such as <it>Arabidopsis thaliana</it>, the <it>anl </it>locus has not been mapped in any <it>Brassica </it>species.</p> <p>Results</p> <p>We tested primer pairs known to amplify microsatellites in <it>Brassicas </it>and identified 37 that amplified a product in rapid-cycling <it>Brassica rapa</it>. We then developed three-generation pedigrees to assess linkage between the microsatellite markers and <it>anl</it>. 22 of the markers that we tested were polymorphic in our crosses. Based on 177 F₂offspring, we identified three markers linked to <it>anl </it>with LOD scores ≥ 5.0, forming a linkage group spanning 46.9 cM. Because one of these markers has been assigned to a known <it>B. rapa </it>linkage group, we can now assign the <it>anl </it>locus to <it>B. rapa </it>linkage group R9.</p> <p>Conclusion</p> <p>This study is the first to identify the chromosomal location of an anthocyanin pigment gene among the <it>Brassicas</it>. It also connects a classical mutant frequently used in genetics education with molecular markers and a known chromosomal location.</p

Mercury toxicity in livers of northern pike (Esox lucius) from Isle Royale, USA

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 C: Toxicology & Pharmacology 147 (2008): 331-338, doi:10.1016/j.cbpc.2007.12.003.Many laboratory studies have documented that mercury can be toxic to fish, but it is largely unknown if mercury is toxic to fish in their natural environments. The objective of our study was to investigate the toxic effects of mercury on northern pike (Esox lucius) at Isle Royale, Michigan. In 124 northern pike from eight inland lakes, concentrations of total mercury in skin-on fillets ranged from 0.069 to 0.622 µg/g wet wt. Concentrations of total mercury in livers increased exponentially compared with concentrations in fillets, to a maximum of 3.1 µg/g wet wt. Methylmercury constituted a majority of the mercury in livers with total mercury concentrations <0.5 µg/g wet wt, but declined to 28-51% of the mercury in livers with total mercury concentrations >0.5 µg/g wet wt. Liver color (absorbance at 400 nm) varied among northern pike and was positively related to liver total mercury concentration. The pigment causing variation in liver color was identified as lipofuscin, which results from lipid peroxidation of membranous organelles. An analysis of covariance revealed lipofuscin accumulation was primarily associated with mercury exposure, and this association obscured any normal accumulation from aging. We also documented decreased lipid reserves in livers and poor condition factors of northern pike with high liver total mercury concentrations. Our results suggest (i) northern pike at Isle Royale are experiencing toxicity at concentrations of total mercury common for northern pike and other piscivorous fish elsewhere in North America and (ii) liver color may be useful for indicating mercury exposure and effects in northern pike at Isle Royale and possibly other aquatic ecosystems and other fish species.Financial support was provided by the U.S. Environmental Protection Agency STAR Graduate Fellowship Program to P.E.D

Transcriptome sequencing and microarray development for the Manila clam, Ruditapes philippinarum: genomic tools for environmental monitoring

Abstract Background The Manila clam, Ruditapes philippinarum, is one of the major aquaculture species in the world and a potential sentinel organism for monitoring the status of marine ecosystems. However, genomic resources for R. philippinarum are still extremely limited. Global analysis of gene expression profiles is increasingly used to evaluate the biological effects of various environmental stressors on aquatic animals under either artificial conditions or in the wild. Here, we report on the development of a transcriptomic platform for global gene expression profiling in the Manila clam. Results A normalized cDNA library representing a mixture of adult tissues was sequenced using a ultra high-throughput sequencing technology (Roche 454). A database consisting of 32,606 unique transcripts was constructed, 9,747 (30%) of which could be annotated by similarity. An oligo-DNA microarray platform was designed and applied to profile gene expression of digestive gland and gills. Functional annotation of differentially expressed genes between different tissues was performed by enrichment analysis. Expression of Natural Antisense Transcripts (NAT) analysis was also performed and bi-directional transcription appears a common phenomenon in the R. philippinarum transcriptome. A preliminary study on clam samples collected in a highly polluted area of the Venice Lagoon demonstrated the applicability of genomic tools to environmental monitoring. Conclusions The transcriptomic platform developed for the Manila clam confirmed the high level of reproducibility of current microarray technology. Next-generation sequencing provided a good representation of the clam transcriptome. Despite the known limitations in transcript annotation and sequence coverage for non model species, sufficient information was obtained to identify a large set of genes potentially involved in cellular response to environmental stress.This work was partially supported by a grant from European Union-funded Network of Excellence "Marine Genomics Europe". CS wishes to acknowledge additional funding from the Ministry of Education and Science (Spain) through grant AGL2007-60049. MM had a PhD scholarship from the University of Florence, Italy. RL was recipient of PhD fellowship SFRH/BD/30112/2006, from the Portuguese Science and Technology Foundation (FCT) and LC and RL acknowledge a grant from FCT project ISOPERK (PTDC/CVT/72083/2006).Peer Reviewe

Psychoactive Pharmaceuticals Induce Fish Gene Expression Profiles Associated with Human Idiopathic Autism

Idiopathic autism, caused by genetic susceptibility interacting with unknown environmental triggers, has increased dramatically in the past 25 years. Identifying environmental triggers has been difficult due to poorly understood pathophysiology and subjective definitions of autism. The use of antidepressants by pregnant women has been associated with autism. These and other unmetabolized psychoactive pharmaceuticals (UPPs) have also been found in drinking water from surface sources, providing another possible exposure route and raising questions about human health consequences. Here, we examined gene expression patterns of fathead minnows treated with a mixture of three psychoactive pharmaceuticals (fluoxetine, venlafaxine & carbamazepine) in dosages intended to be similar to the highest observed conservative estimates of environmental concentrations. We conducted microarray experiments examining brain tissue of fish exposed to individual pharmaceuticals and a mixture of all three. We used gene-class analysis to test for enrichment of gene sets involved with ten human neurological disorders. Only sets associated with idiopathic autism were unambiguously enriched. We found that UPPs induce autism-like gene expression patterns in fish. Our findings suggest a new potential trigger for idiopathic autism in genetically susceptible individuals involving an overlooked source of environmental contamination

Digging for names in the mountains: Combined person name recognition and reference resolution for German alpine texts

In this paper we introduce a module that combines person name recognition and reference resolution for German. Our data consisted of a corpus of Alpine texts. This text type poses special challenges because of a multitude of toponyms, some of which interfere with person names. Our reference resolution algorithm outputs person entities based on their last names and first names along with their associated features (jobs, addresses, academic titles)