Identification of Methylmercury Tolerance Gene Candidates in Drosophila

Department of Anatomy and Neurobiology, College of Medicine, University of Vermont, Burlington, Vermont 05405, USA.
Toxicological Sciences (Impact Factor: 3.85). 04/2010; 116(1):225-38. DOI: 10.1093/toxsci/kfq097
Source: PubMed


Methylmercury (MeHg) is a ubiquitous environmental contaminant that preferentially targets the developing nervous system.
Variable outcomes of prenatal MeHg exposure within a population point to a genetic component that regulates MeHg toxicity.
We therefore sought to identify fundamental MeHg tolerance genes using the Drosophila model for genetic and molecular dissection
of a MeHg tolerance trait. We observe autosomal dominance in a MeHg tolerance trait (development on MeHg food) in both wild-derived
and laboratory-selected MeHg-tolerant strains of flies. We performed whole-genome transcript profiling of larval brains of
tolerant (laboratory selected) and nontolerant (control) strains in the presence and absence of MeHg stress. Pairwise transcriptome
comparisons of four conditions (+/−selection and +/−MeHg) identified a “down-down-up” expression signature, whereby MeHg alone
and selection alone resulted in a greater number of downregulated transcripts, and the combination of selection + MeHg resulted
in a greater number of upregulated transcripts. Functional annotation cluster analyses showed enrichment for monooxygenases/oxidoreductases,
which include cytochrome P450 (CYP) family members. Among the 10 CYPs upregulated with selection + MeHg in tolerant strains,
CYP6g1, previously identified as the dichlorodiphenyl trichloroethane resistance allele in flies, was the most highly expressed
and responsive to MeHg. Among all the genes, Turandot A (TotA), an immune pathway–regulated humoral response gene, showed the greatest upregulation with selection + MeHg. Neural-specific
transgenic overexpression of TotA enhanced MeHg tolerance during pupal development. Identification of TotA and CYP genes as MeHg tolerance genes is an inroad to investigating the conserved function of immune signaling and phase I metabolism
pathways in MeHg toxicity and tolerance in higher organisms.

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Available from: David M. Rand, Dec 17, 2013
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    • "Expression levels in a pooled sample of 20 larvae were determined by qRT-PCR, normalized to RP49 and expressed relative to Canton S at the 0M MeHg exposure. bility (Magnusson and Ramel, 1986; Mahapatra et al., 2010). "
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    ABSTRACT: Methylmercury (MeHg) is a ubiquitous and persistent neurotoxin that poses a risk to human health. While the mechanisms of MeHg toxicity are not fully understood, factors that contribute to susceptibility are even less well known. Studies of human gene polymorphisms have identified a potential role for the multidrug resistance-like protein (MRP/ABCC) family, ATP-dependent transporters, in MeHg susceptibility. MRP transporters have been shown to be important for MeHg excretion in adult mouse models, but their role in moderating MeHg toxicity during development has not been explored. We therefore investigated effects of manipulating expression levels of MRP using a Drosophila development assay. Drosophila MRP (dMRP) is homologous to human MRP1-4 (ABCC1-4), sharing 50% identity and 67% similarity with MRP1. A greater susceptibility to MeHg is seen in dMRP mutant flies, demonstrated by reduced rates of eclosion on MeHg-containing food. Furthermore, targeted knockdown of dMRP expression using GAL4>UAS RNAi methods demonstrates a tissue-specific function for dMRP in gut, Malpighian tubules, and the nervous system in moderating developmental susceptibility to MeHg. Using X-ray synchrotron fluorescence imaging (XFI), these same tissues were also identified as the highest Hg-accumulating tissues in fly larvae. Moreover, higher levels of Hg are seen in dMRP mutant larvae compared to a control strain fed an equivalent dose of MeHg. In sum, these data demonstrate that dMRP expression, both globally and within Hg-targeted organs, has a profound effect on susceptibility to MeHg in developing flies. Our findings point to a potentially novel and specific role for dMRP in neurons in the protection against MeHg. Finally, this experimental system provides a tractable model to evaluate human polymorphic variants of MRP and other gene variants relevant to genetic studies of mercury-exposed populations.
    Full-text · Article · May 2014 · Toxicological Sciences
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    • "In contrast, embryos are soaked in MeHg medium allowing for more direct contact with embryonic cells. Yet, at the doses used, larvae show similar lethality as embryos (Mahapatra et al., 2010). Assuming that embryo-specific induction of E(spl)mδ is not related to the route of MeHg administration it may help elucidate the mechanism for the specific neurodevelopmental toxicity in mice and humans. "
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    ABSTRACT: Methylmercury (MeHg) is a ubiquitous toxicant that targets the developing fetal nervous system. MeHg interacts with the Notch signaling pathway, a highly-conserved intercellular signaling mechanism required for normal development. Notch signaling is conveyed by activation of the genes in the enhancer of split (E(spl)) locus in Drosophila. We have previously shown that acute high doses of MeHg upregulate several E(spl) genes in Drosophila neural-derived C6 cells. Furthermore, MeHg induction of E(spl) can occur independent of the Notch receptor itself. We now show that MeHg, unlike inorganic mercury (HgCl2), preferentially upregulates E(spl)mδ and E(spl)mγ in Drosophila C6 cells. This is distinct from Delta ligand-induced Notch signaling in which no induction of E(spl)mδ is seen. MeHg is also seen to specifically upregulate E(spl)mδ in Drosophila embryos where HgCl2 showed no such effect. Additionally, treatment of embryos with MeHg caused a consistent failure in axonal outgrowth of the intersegmental nerve (ISN). This ISN phenotype was partially replicated by genetic activation of the Notch pathway, but was not replicated by increasing expression of E(spl)mδ. These data suggest a role for Notch signaling and the E(spl)mδ target gene in MeHg toxicity, however, the site of action for E(spl)mδ in this system remains to be elucidated.
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