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Genome-wide association analysis of tolerance to methylmercury toxicity in Drosophila implicates myogenic and neuromuscular developmental pathways.

Montgomery SL, Vorojeikina D, Huang W, Mackay TF, Anholt RR, Rand MD - PLoS ONE (2014)

Bottom Line: We found significant genetic variation in the effects of MeHg on development, measured by eclosion rate, giving a broad sense heritability of 0.86.We found that caffeine counteracts the deleterious effects of MeHg in the majority of lines, and there is significant genetic variance in the magnitude of this effect, with a broad sense heritability of 0.80.Furthermore, our observations that caffeine can ameliorate the toxic effects of MeHg show that nutritional factors and dietary manipulations may offer protection against the deleterious effects of MeHg exposure.

View Article: PubMed Central - PubMed

Affiliation: Department of Environmental Medicine, University of Rochester School of Medicine and Dentistry, Rochester, New York, United States of America.

ABSTRACT
Methylmercury (MeHg) is a persistent environmental toxin present in seafood that can compromise the developing nervous system in humans. The effects of MeHg toxicity varies among individuals, despite similar levels of exposure, indicating that genetic differences contribute to MeHg susceptibility. To examine how genetic variation impacts MeHg tolerance, we assessed developmental tolerance to MeHg using the sequenced, inbred lines of the Drosophila melanogaster Genetic Reference Panel (DGRP). We found significant genetic variation in the effects of MeHg on development, measured by eclosion rate, giving a broad sense heritability of 0.86. To investigate the influence of dietary factors, we measured MeHg toxicity with caffeine supplementation in the DGRP lines. We found that caffeine counteracts the deleterious effects of MeHg in the majority of lines, and there is significant genetic variance in the magnitude of this effect, with a broad sense heritability of 0.80. We performed genome-wide association (GWA) analysis for both traits, and identified candidate genes that fall into several gene ontology categories, with enrichment for genes involved in muscle and neuromuscular development. Overexpression of glutamate-cysteine ligase, a MeHg protective enzyme, in a muscle-specific manner leads to a robust rescue of eclosion of flies reared on MeHg food. Conversely, mutations in kirre, a pivotal myogenic gene identified in our GWA analyses, modulate tolerance to MeHg during development in accordance with kirre expression levels. Finally, we observe disruptions of indirect flight muscle morphogenesis in MeHg-exposed pupae. Since the pathways for muscle development are evolutionarily conserved, it is likely that the effects of MeHg observed in Drosophila can be generalized across phyla, implicating muscle as an additional hitherto unrecognized target for MeHg toxicity. Furthermore, our observations that caffeine can ameliorate the toxic effects of MeHg show that nutritional factors and dietary manipulations may offer protection against the deleterious effects of MeHg exposure.

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Genome-wide association analysis of eclosion on MeHg with and without caffeine supplementation.Single marker analyses using ANOVA of 2,180,555 (MeHg alone) and 2,357,353 (MeHg+caffeine) polymorphic alleles across 167 (MeHg alone) and 139 (MeHg+caffeine) DGRP lines, respectively, resolved (A) 350 and (B) 239 polymorphic markers (p<10−4, MAF>3%). Depicted is a heat map for linkage disequilibrium (LD) based on r2 values where the black bars represent the five major Drosophila chromosome arms. Red indicates high LD, while blue indicates low LD. The black dots represent polymorphic marker associations for eclosion on MeHg or MeHg+caffeine. p values (log10(p)), effect size (a/σP), and the minor allele frequency (MAF) are shown. (C,D) MAF vs. effect size. All 350 (C) or 239 (D) polymorphic markers associated with phenotypic variation for eclosion on MeHg or MeHg+caffeine, respectively, are depicted. a/σp indicates effect size ([mean of major allele class – mean of minor allele class]/2), where negative a/σp indicates the minor allele is associated with increased MeHg tolerance with respect to eclosion.
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pone-0110375-g003: Genome-wide association analysis of eclosion on MeHg with and without caffeine supplementation.Single marker analyses using ANOVA of 2,180,555 (MeHg alone) and 2,357,353 (MeHg+caffeine) polymorphic alleles across 167 (MeHg alone) and 139 (MeHg+caffeine) DGRP lines, respectively, resolved (A) 350 and (B) 239 polymorphic markers (p<10−4, MAF>3%). Depicted is a heat map for linkage disequilibrium (LD) based on r2 values where the black bars represent the five major Drosophila chromosome arms. Red indicates high LD, while blue indicates low LD. The black dots represent polymorphic marker associations for eclosion on MeHg or MeHg+caffeine. p values (log10(p)), effect size (a/σP), and the minor allele frequency (MAF) are shown. (C,D) MAF vs. effect size. All 350 (C) or 239 (D) polymorphic markers associated with phenotypic variation for eclosion on MeHg or MeHg+caffeine, respectively, are depicted. a/σp indicates effect size ([mean of major allele class – mean of minor allele class]/2), where negative a/σp indicates the minor allele is associated with increased MeHg tolerance with respect to eclosion.

Mentions: In Drosophila, we can use GWA analysis as an exploratory hypothesis-generating tool, and test hypotheses more rigorously in secondary screens using mutations or targeting RNAI for candidate genes implicated by the GWA analysis. Therefore, we used a lenient reporting threshold of p<10−4 for both GWA analyses (Tables S1, S2). We identified 350 polymorphisms in or near 145 genes associated with variation in eclosion rates on MeHg food (Fig. 3A) and 239 polymorphisms in or near 106 genes (Fig. 3B) associated with the modulatory effect of caffeine on MeHg treatment. Most polymorphisms associated with the two analyses had MAF <0.15, and, as expected, we found inverse relationships of effect size and allele frequency (Figs. 3C, 3D). Rarer alleles were associated with greater tolerance to MeHg and modulation of the effect by caffeine with respect to larva-adult viability for a majority of the polymorphisms (Figs. 3C, 3D), suggesting these alleles have other deleterious effects on fitness. There was little linkage disequilibrium between the most significant polymorphisms (Figs. 3A, 3B).


Genome-wide association analysis of tolerance to methylmercury toxicity in Drosophila implicates myogenic and neuromuscular developmental pathways.

Montgomery SL, Vorojeikina D, Huang W, Mackay TF, Anholt RR, Rand MD - PLoS ONE (2014)

Genome-wide association analysis of eclosion on MeHg with and without caffeine supplementation.Single marker analyses using ANOVA of 2,180,555 (MeHg alone) and 2,357,353 (MeHg+caffeine) polymorphic alleles across 167 (MeHg alone) and 139 (MeHg+caffeine) DGRP lines, respectively, resolved (A) 350 and (B) 239 polymorphic markers (p<10−4, MAF>3%). Depicted is a heat map for linkage disequilibrium (LD) based on r2 values where the black bars represent the five major Drosophila chromosome arms. Red indicates high LD, while blue indicates low LD. The black dots represent polymorphic marker associations for eclosion on MeHg or MeHg+caffeine. p values (log10(p)), effect size (a/σP), and the minor allele frequency (MAF) are shown. (C,D) MAF vs. effect size. All 350 (C) or 239 (D) polymorphic markers associated with phenotypic variation for eclosion on MeHg or MeHg+caffeine, respectively, are depicted. a/σp indicates effect size ([mean of major allele class – mean of minor allele class]/2), where negative a/σp indicates the minor allele is associated with increased MeHg tolerance with respect to eclosion.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4215868&req=5

pone-0110375-g003: Genome-wide association analysis of eclosion on MeHg with and without caffeine supplementation.Single marker analyses using ANOVA of 2,180,555 (MeHg alone) and 2,357,353 (MeHg+caffeine) polymorphic alleles across 167 (MeHg alone) and 139 (MeHg+caffeine) DGRP lines, respectively, resolved (A) 350 and (B) 239 polymorphic markers (p<10−4, MAF>3%). Depicted is a heat map for linkage disequilibrium (LD) based on r2 values where the black bars represent the five major Drosophila chromosome arms. Red indicates high LD, while blue indicates low LD. The black dots represent polymorphic marker associations for eclosion on MeHg or MeHg+caffeine. p values (log10(p)), effect size (a/σP), and the minor allele frequency (MAF) are shown. (C,D) MAF vs. effect size. All 350 (C) or 239 (D) polymorphic markers associated with phenotypic variation for eclosion on MeHg or MeHg+caffeine, respectively, are depicted. a/σp indicates effect size ([mean of major allele class – mean of minor allele class]/2), where negative a/σp indicates the minor allele is associated with increased MeHg tolerance with respect to eclosion.
Mentions: In Drosophila, we can use GWA analysis as an exploratory hypothesis-generating tool, and test hypotheses more rigorously in secondary screens using mutations or targeting RNAI for candidate genes implicated by the GWA analysis. Therefore, we used a lenient reporting threshold of p<10−4 for both GWA analyses (Tables S1, S2). We identified 350 polymorphisms in or near 145 genes associated with variation in eclosion rates on MeHg food (Fig. 3A) and 239 polymorphisms in or near 106 genes (Fig. 3B) associated with the modulatory effect of caffeine on MeHg treatment. Most polymorphisms associated with the two analyses had MAF <0.15, and, as expected, we found inverse relationships of effect size and allele frequency (Figs. 3C, 3D). Rarer alleles were associated with greater tolerance to MeHg and modulation of the effect by caffeine with respect to larva-adult viability for a majority of the polymorphisms (Figs. 3C, 3D), suggesting these alleles have other deleterious effects on fitness. There was little linkage disequilibrium between the most significant polymorphisms (Figs. 3A, 3B).

Bottom Line: We found significant genetic variation in the effects of MeHg on development, measured by eclosion rate, giving a broad sense heritability of 0.86.We found that caffeine counteracts the deleterious effects of MeHg in the majority of lines, and there is significant genetic variance in the magnitude of this effect, with a broad sense heritability of 0.80.Furthermore, our observations that caffeine can ameliorate the toxic effects of MeHg show that nutritional factors and dietary manipulations may offer protection against the deleterious effects of MeHg exposure.

View Article: PubMed Central - PubMed

Affiliation: Department of Environmental Medicine, University of Rochester School of Medicine and Dentistry, Rochester, New York, United States of America.

ABSTRACT
Methylmercury (MeHg) is a persistent environmental toxin present in seafood that can compromise the developing nervous system in humans. The effects of MeHg toxicity varies among individuals, despite similar levels of exposure, indicating that genetic differences contribute to MeHg susceptibility. To examine how genetic variation impacts MeHg tolerance, we assessed developmental tolerance to MeHg using the sequenced, inbred lines of the Drosophila melanogaster Genetic Reference Panel (DGRP). We found significant genetic variation in the effects of MeHg on development, measured by eclosion rate, giving a broad sense heritability of 0.86. To investigate the influence of dietary factors, we measured MeHg toxicity with caffeine supplementation in the DGRP lines. We found that caffeine counteracts the deleterious effects of MeHg in the majority of lines, and there is significant genetic variance in the magnitude of this effect, with a broad sense heritability of 0.80. We performed genome-wide association (GWA) analysis for both traits, and identified candidate genes that fall into several gene ontology categories, with enrichment for genes involved in muscle and neuromuscular development. Overexpression of glutamate-cysteine ligase, a MeHg protective enzyme, in a muscle-specific manner leads to a robust rescue of eclosion of flies reared on MeHg food. Conversely, mutations in kirre, a pivotal myogenic gene identified in our GWA analyses, modulate tolerance to MeHg during development in accordance with kirre expression levels. Finally, we observe disruptions of indirect flight muscle morphogenesis in MeHg-exposed pupae. Since the pathways for muscle development are evolutionarily conserved, it is likely that the effects of MeHg observed in Drosophila can be generalized across phyla, implicating muscle as an additional hitherto unrecognized target for MeHg toxicity. Furthermore, our observations that caffeine can ameliorate the toxic effects of MeHg show that nutritional factors and dietary manipulations may offer protection against the deleterious effects of MeHg exposure.

Show MeSH
Related in: MedlinePlus