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The interplay between protein L-isoaspartyl methyltransferase activity and insulin-like signaling to extend lifespan in Caenorhabditis elegans.

Khare S, Linster CL, Clarke SG - PLoS ONE (2011)

Bottom Line: We demonstrate that reducing the levels of the DAF-16 downstream transcriptional effector of the IIS pathway by RNA interference reduces the lifespan extension resulting from PCM-1 overexpression.Using quantitative real-time PCR analysis, we show the up-regulation of DAF-16-dependent stress response genes in the PCM-1 overexpressor animals compared to wild-type and pcm-1 mutant nematodes under mild thermal stress conditions.Although we observe a higher accumulation of damaged proteins in pcm-1 mutant nematodes, the basal level of isoaspartyl residues detected in wild-type animals was not reduced by PCM-1 overexpression.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Biochemistry, University of California-Los Angeles, Los Angeles, California, United States of America.

ABSTRACT
The protein L-isoaspartyl-O-methyltransferase functions to initiate the repair of isomerized aspartyl and asparaginyl residues that spontaneously accumulate with age in a variety of organisms. Caenorhabditis elegans nematodes lacking the pcm-1 gene encoding this enzyme display a normal lifespan and phenotype under standard laboratory growth conditions. However, significant defects in development, egg laying, dauer survival, and autophagy have been observed in pcm-1 mutant nematodes when deprived of food and when exposed to oxidative stress. Interestingly, overexpression of this repair enzyme in both Drosophila and C. elegans extends adult lifespan under thermal stress. In this work, we show the involvement of the insulin/insulin-like growth factor-1 signaling (IIS) pathway in PCM-1-dependent lifespan extension in C. elegans. We demonstrate that reducing the levels of the DAF-16 downstream transcriptional effector of the IIS pathway by RNA interference reduces the lifespan extension resulting from PCM-1 overexpression. Using quantitative real-time PCR analysis, we show the up-regulation of DAF-16-dependent stress response genes in the PCM-1 overexpressor animals compared to wild-type and pcm-1 mutant nematodes under mild thermal stress conditions. Additionally, similar to other long-lived C. elegans mutants in the IIS pathway, including daf-2 and age-1 mutants, PCM-1 overexpressor adult animals display increased resistance to severe thermal stress, whereas pcm-1 mutant animals survive less long under these conditions. Although we observe a higher accumulation of damaged proteins in pcm-1 mutant nematodes, the basal level of isoaspartyl residues detected in wild-type animals was not reduced by PCM-1 overexpression. Our results support a signaling role for the protein L-isoaspartyl methyltransferase in lifespan extension that involves the IIS pathway, but that may be independent of its function in overall protein repair.

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PCM-1 overexpression enhances resistance to severe heat stress (37°C).Survival assays (three trials) were completed for four nematode strains (N2 wild-type, pcm-1 mutant (qa201), PCM-1 overexpressor strain (PL51), mutant PCM-1 overexpressor strain (PL54)). Shown are representative survival curves from one trial. L4 larvae were transferred to NGM + OP50 plates and were allowed to grow at 20°C overnight. The next day, animals were transferred to 37°C and survival was scored every two hours for the first four hours and every hour afterwards until all nematodes were dead. Statistical analysis of these data is given in Table 3 for the three replicates of this experiment.
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pone-0020850-g002: PCM-1 overexpression enhances resistance to severe heat stress (37°C).Survival assays (three trials) were completed for four nematode strains (N2 wild-type, pcm-1 mutant (qa201), PCM-1 overexpressor strain (PL51), mutant PCM-1 overexpressor strain (PL54)). Shown are representative survival curves from one trial. L4 larvae were transferred to NGM + OP50 plates and were allowed to grow at 20°C overnight. The next day, animals were transferred to 37°C and survival was scored every two hours for the first four hours and every hour afterwards until all nematodes were dead. Statistical analysis of these data is given in Table 3 for the three replicates of this experiment.

Mentions: In addition to displaying extended adult lifespans, daf-2 pathway mutants also show resistance to a variety of stressors, including severe thermal stress [32]. In testing the PL51 PCM-1 overexpressor strain for resistance to heat stress, we found that day 1 adults displayed increased thermotolerance at 37°C compared to wild-type (N2) animals, most significantly at the 25% survival time point (Fig. 2, Table 3). PL51 adult animals displayed 25% survival at 4.5±0.5 h compared to 3.2±0.8 h in N2 animals. The enhanced thermotolerance appeared to be at least partially dependent on a functional methyltransferase cofactor-binding domain, as the PL54 animals did not show significantly increased survival at any noted time point. Furthermore, pcm-1 mutant animals displayed a significant decrease in thermotolerance when compared to PL51 animals at all 50%, 25%, and 10% survival time points (Table 3).


The interplay between protein L-isoaspartyl methyltransferase activity and insulin-like signaling to extend lifespan in Caenorhabditis elegans.

Khare S, Linster CL, Clarke SG - PLoS ONE (2011)

PCM-1 overexpression enhances resistance to severe heat stress (37°C).Survival assays (three trials) were completed for four nematode strains (N2 wild-type, pcm-1 mutant (qa201), PCM-1 overexpressor strain (PL51), mutant PCM-1 overexpressor strain (PL54)). Shown are representative survival curves from one trial. L4 larvae were transferred to NGM + OP50 plates and were allowed to grow at 20°C overnight. The next day, animals were transferred to 37°C and survival was scored every two hours for the first four hours and every hour afterwards until all nematodes were dead. Statistical analysis of these data is given in Table 3 for the three replicates of this experiment.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3113807&req=5

pone-0020850-g002: PCM-1 overexpression enhances resistance to severe heat stress (37°C).Survival assays (three trials) were completed for four nematode strains (N2 wild-type, pcm-1 mutant (qa201), PCM-1 overexpressor strain (PL51), mutant PCM-1 overexpressor strain (PL54)). Shown are representative survival curves from one trial. L4 larvae were transferred to NGM + OP50 plates and were allowed to grow at 20°C overnight. The next day, animals were transferred to 37°C and survival was scored every two hours for the first four hours and every hour afterwards until all nematodes were dead. Statistical analysis of these data is given in Table 3 for the three replicates of this experiment.
Mentions: In addition to displaying extended adult lifespans, daf-2 pathway mutants also show resistance to a variety of stressors, including severe thermal stress [32]. In testing the PL51 PCM-1 overexpressor strain for resistance to heat stress, we found that day 1 adults displayed increased thermotolerance at 37°C compared to wild-type (N2) animals, most significantly at the 25% survival time point (Fig. 2, Table 3). PL51 adult animals displayed 25% survival at 4.5±0.5 h compared to 3.2±0.8 h in N2 animals. The enhanced thermotolerance appeared to be at least partially dependent on a functional methyltransferase cofactor-binding domain, as the PL54 animals did not show significantly increased survival at any noted time point. Furthermore, pcm-1 mutant animals displayed a significant decrease in thermotolerance when compared to PL51 animals at all 50%, 25%, and 10% survival time points (Table 3).

Bottom Line: We demonstrate that reducing the levels of the DAF-16 downstream transcriptional effector of the IIS pathway by RNA interference reduces the lifespan extension resulting from PCM-1 overexpression.Using quantitative real-time PCR analysis, we show the up-regulation of DAF-16-dependent stress response genes in the PCM-1 overexpressor animals compared to wild-type and pcm-1 mutant nematodes under mild thermal stress conditions.Although we observe a higher accumulation of damaged proteins in pcm-1 mutant nematodes, the basal level of isoaspartyl residues detected in wild-type animals was not reduced by PCM-1 overexpression.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Biochemistry, University of California-Los Angeles, Los Angeles, California, United States of America.

ABSTRACT
The protein L-isoaspartyl-O-methyltransferase functions to initiate the repair of isomerized aspartyl and asparaginyl residues that spontaneously accumulate with age in a variety of organisms. Caenorhabditis elegans nematodes lacking the pcm-1 gene encoding this enzyme display a normal lifespan and phenotype under standard laboratory growth conditions. However, significant defects in development, egg laying, dauer survival, and autophagy have been observed in pcm-1 mutant nematodes when deprived of food and when exposed to oxidative stress. Interestingly, overexpression of this repair enzyme in both Drosophila and C. elegans extends adult lifespan under thermal stress. In this work, we show the involvement of the insulin/insulin-like growth factor-1 signaling (IIS) pathway in PCM-1-dependent lifespan extension in C. elegans. We demonstrate that reducing the levels of the DAF-16 downstream transcriptional effector of the IIS pathway by RNA interference reduces the lifespan extension resulting from PCM-1 overexpression. Using quantitative real-time PCR analysis, we show the up-regulation of DAF-16-dependent stress response genes in the PCM-1 overexpressor animals compared to wild-type and pcm-1 mutant nematodes under mild thermal stress conditions. Additionally, similar to other long-lived C. elegans mutants in the IIS pathway, including daf-2 and age-1 mutants, PCM-1 overexpressor adult animals display increased resistance to severe thermal stress, whereas pcm-1 mutant animals survive less long under these conditions. Although we observe a higher accumulation of damaged proteins in pcm-1 mutant nematodes, the basal level of isoaspartyl residues detected in wild-type animals was not reduced by PCM-1 overexpression. Our results support a signaling role for the protein L-isoaspartyl methyltransferase in lifespan extension that involves the IIS pathway, but that may be independent of its function in overall protein repair.

Show MeSH
Related in: MedlinePlus