Identification & Characterization of Methylmercury (MeHg) Tolerance

We are actively engaged in unbiased whole-genome screens using the Drosophila (fruit fly) model to identify novel candidate genes and pathways that function in development and moderate MeHg toxicity. Using a transcriptomic approach we have resolved a role for Phase I metabolism genes of the Cytochrome P450 (CYP) family as tolerance genes in MeHg toxicity. The strongest candidate, Drosophila CYP6g1, is the homolog of human CYP3A genes (CYP3A4, 3A5, and 3A7), which are the foremost xenobiotic metabolizing enzymes in the human liver. We are actively characterizing the role of both human and Drosophila CYP genes in several contexts of development, including the central nervous system and other known MeHg target organs in the developing embryo. These studies have also identified a beneficial role for caffeine, a known inducer of CYP activity, in inducing MeHg tolerance.

Pictured Right: Global changes in transcripts in developing brains of flies selected for MeHg tolerance.  Scatter plots illustrating the pair-wise comparisons of the entire Affymetrix Genechip (Drosophila 2) probeset intensity data using the criteria of ≥1.5-fold change (black dots, 1.5-fold boundary marked by solid lines) and  p ≤ 0.05.  (Axes values are log2 of probe intensity) (A) MeHg exposure to S0 (non MeHg-selected) strain leads to differential expression of 362 transcripts with the majority (90%) down-regulated (below the diagonal). (B) Basal expression in the S20 (20µM MeHg tolerance selected) strain versus S0 strains shows a change in 246 transcripts and majority of them (72%) are down-regulated in the S20 due to selection. (C) MeHg exposure in S20 strains shows differential expression of 249 transcripts with 44% up-regulated (above the diagonal). (D) S20 vs S0 upon MeHg exposure shows differential expression of 233 transcripts with 74% showing up-regulation. (Values in the parentheses are differential expression with criteria of ≥ 2 fold change and p-value ≤ 0.05). (From: Mahapatra et al., Toxicol. Sci. 104:163, 2010)

A second round of screening, using a SNP-based genome-wide association (GWA) approach, is underway and is exploiting the unique power of the recently released Drosophila Genetics Reference Panel (DGRP) (MacKay et al., 2012 Nature 482:173). The DGRP consists of a collection of more than 200 lines of inbred wild-derived flies with fully sequenced genomes. More than 2,500,000 SNPs can be analyzed in GWA for the phenotypic trait of tolerance or susceptibility to MeHg during development. Characterization of this panel of flies is currently resolving several new candidate genes and revealing new insights into pathways intrinsic to both neural and non-neural targets of methylmercury during development. These studies are being extended to examine conserved mechanisms in mammals through collaborations with Environmental Health Center researchers at U of R. We anticipate our unique genomic approaches to characterizing MeHg toxicity mechanisms will advance the understanding of the fundamental basis of MeHg insult in people and contribute much needed scientific integrity to the principles guiding risk assessment for MeHg exposure in the future.

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