The MDT-15 subunit of mediator interacts with dietary restriction to modulate longevity and fluoranthene toxicity in Caenorhabditis elegans

Abstract

Dietary restriction (DR), the limitation of calorie intake while maintaining proper nutrition, has been found to extend life span and delay the onset of age-associated disease in a wide range of species. Previous studies have suggested that DR can reduce the lethality of environmental toxins. To further examine the role of DR in toxin response, we measured life spans of the nematode Caenorhabditis elegans treated with the mutagenic polyaromatic hydrocarbon, fluoranthene (FLA). FLA is a direct byproduct of combustion, and is one of U.S. Environmental Protection Agency's sixteen priority environmental toxins. Treatment with 5 µg/ml FLA shortened the life spans of ad libitum fed nematodes, and DR resulted in increased sensitivity to FLA. To determine the role of detoxifying enzymes in the toxicity of FLA, we tested nematodes with mutations in the gene encoding the MDT-15 subunit of mediator, a transcriptional coactivator that regulates genes involved in fatty acid metabolism and detoxification. Mutation of mdt-15 increased the life span of FLA treated animals compared to wild-type animals with no difference observed between DR and ad libitum fed mdt-15 animals. We also examined mutants with altered insulin-IGF-1-like signaling (IIS), which is known to modulate life span and stress resistance in C. elegans independently of DR. Mutation of the genes coding for the insulin-like receptor DAF-2 or the FOXO-family transcription factor DAF16 did not alter the animals' susceptibility to FLA compared to wild type. Taken together, our results suggest that certain compounds have increased toxicity when combined with a DR regimen through increased metabolic activation. This increased metabolic activation appears to be mediated through the MDT-15 transcription factor and is independent of the IIS pathway.

Figure 1. Dietary Restriction Shortens the Life Span of Fluoranthene Treated N2 Adult Animals.

(A) The chemical structure of fluoranthene (FLA). (B) Life span of N2 animals treated continuously with DMSO or FLA starting at adult day 4. Pooled data is shown, mean life span is shown in parentheses. Treatment with FLA shortened life span of N2 animals under control-fed conditions. Bacterial deprivation (BD) further shortened the life span of N2 animals compared to the control-fed. Summary data and statistics for both pooled and individual experiments are provided in Table S1.

Figure 2. Bacterial Deprivation and Fluoranthene Treatment Reduce Pharyngeal Pumping.

Pharyngeal pump rates of N2 and eat-2(ad1116) animals after 24 hours treatment with DMSO or FLA under control-fed (Fed) or bacterially deprived (BD) conditions. Pharyngeal pumping was reduced in both FLA treated and BD populations. *denotes p>0.0001 compared to DMSO treated samples. Rates are recorded as pumps per minute. Summary data and statistics are shown in Table 1 .

Figure 3. eat-2(ad1116) Animals are not Resistant to FLA Treatment.

Life spans of N2 and eat-2(ad1116) animals after continuous exposure to DMSO or FLA starting at day 4 of adulthood. Pooled data is shown, mean life span is shown in parentheses. Mutation of eat-2 increased life span in DMSO treated populations compared to N2. FLA treated N2 and eat-2(ad1116) populations were not significantly different. Summary data and statistics for both pooled and individual experiments are shown in Table S2.

Figure 4. FLA Induces Nuclear Localization of Daf-16.

Proportion of DAF-16::GFP animals with nuclear puncta after treatment with FLA, DMSO, daf-2(RNAi), or 2 hour incubation at 37°C (A). Treatment with FLA induced nuclear puncta similar to those observed daf-2(RNAi) with treatment. (B) Images of DAF-16::GFP animals treated with DMSO (i), heat-shock (ii), or FLA (iii). Summary data and statistics are shown in Table 2 .

Figure 5. Insulin/IGF-1 signaling does not influence FLA toxicity.

Life spans of N2, daf-2(e1370), and daf-16(mu86) animals after continuous treatment with DMSO or FLA under control-fed conditions starting at day 4 of adulthood. Mutation of daf-2 and daf-16 extended and shortened life span, respectively, compared to N2 in DMSO treated samples. N2, daf-2(e1370), and daf-16(mu86) animals displayed significantly shortened life spans when treated with FLA. Summary data and statistics for both pooled and individual experiments are shown in Table S3.

Figure 6. mdt-15(tm2182) mutants are resistant to FLA and improve response to BD.

Life spans of N2 and mdt-15(tm2182) animals after continuous treatment with DMSO or FLA under control fed or BD conditions starting at day 4 of adulthood. mdt-15(tm2182) animals had life spans similar to N2 under control fed conditions. Treatment with FLA shortened the life spans of mdt-15(tm2182) animals but not as severely as N2 animals. Control fed and BD mdt-15(tm2182) animals treated with FLA did not have significantly different life spans. Summary data and statistics for both pooled and individual experiments are shown in Table S4.