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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 extends lifespan in a daf-16-dependent manner under mild thermal stress (25°C).Lifespan analyses (three trials) were completed at 25°C for three nematode strains (N2 wild-type (top panel), wild-type PCM-1 overexpressor strain (PL51) (middle panel), mutant PCM-1 overexpressor strain (PL54) (bottom panel)) grown in two conditions: control RNAi (solid lines) and daf-16 RNAi (dotted lines). L4 larvae were transferred (Day 0) to NGM plates streaked with RNAi bacteria and survival was scored every other day until all nematodes were dead. Statistical analysis of these data is given in Table 2.
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pone-0020850-g001: PCM-1 overexpression extends lifespan in a daf-16-dependent manner under mild thermal stress (25°C).Lifespan analyses (three trials) were completed at 25°C for three nematode strains (N2 wild-type (top panel), wild-type PCM-1 overexpressor strain (PL51) (middle panel), mutant PCM-1 overexpressor strain (PL54) (bottom panel)) grown in two conditions: control RNAi (solid lines) and daf-16 RNAi (dotted lines). L4 larvae were transferred (Day 0) to NGM plates streaked with RNAi bacteria and survival was scored every other day until all nematodes were dead. Statistical analysis of these data is given in Table 2.

Mentions: To test whether the PCM-1 overexpression effect was due to a modulation of the IIS system, we tested the dependence of this effect on the presence of DAF-16, the critical downstream transcription factor of the IIS. Specifically, we grew the N2, PL51, and PL54 strains on bacteria expressing scrambled control RNAi or daf-16 RNAi and used FUDR to prevent egg laying and cross generation sampling [29], [36]. We confirmed that PCM-1 overexpressing animals display on average an increase of 38% and 51% in median and maximum lifespan, respectively, compared to wild-type animals, and that no change in survival is observed between wild-type animals and nematodes expressing a catalytically inactive PCM-1 enzyme (Fig. 1, Table 2). In all strains tested, the lifespan values obtained in this study in the presence of FUDR were slightly reduced compared to those obtained previously in the absence of FUDR by Banfield et al. [17], but the same lifespan patterns were observed on control RNAi bacteria (Fig. 1, Table 2). All lifespan extensions (including the 50%, 25% and 10% survival values) measured for the overexpressor strain were statistically significant when compared to wild-type (N2) animals (Table 2).


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 extends lifespan in a daf-16-dependent manner under mild thermal stress (25°C).Lifespan analyses (three trials) were completed at 25°C for three nematode strains (N2 wild-type (top panel), wild-type PCM-1 overexpressor strain (PL51) (middle panel), mutant PCM-1 overexpressor strain (PL54) (bottom panel)) grown in two conditions: control RNAi (solid lines) and daf-16 RNAi (dotted lines). L4 larvae were transferred (Day 0) to NGM plates streaked with RNAi bacteria and survival was scored every other day until all nematodes were dead. Statistical analysis of these data is given in Table 2.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3113807&req=5

pone-0020850-g001: PCM-1 overexpression extends lifespan in a daf-16-dependent manner under mild thermal stress (25°C).Lifespan analyses (three trials) were completed at 25°C for three nematode strains (N2 wild-type (top panel), wild-type PCM-1 overexpressor strain (PL51) (middle panel), mutant PCM-1 overexpressor strain (PL54) (bottom panel)) grown in two conditions: control RNAi (solid lines) and daf-16 RNAi (dotted lines). L4 larvae were transferred (Day 0) to NGM plates streaked with RNAi bacteria and survival was scored every other day until all nematodes were dead. Statistical analysis of these data is given in Table 2.
Mentions: To test whether the PCM-1 overexpression effect was due to a modulation of the IIS system, we tested the dependence of this effect on the presence of DAF-16, the critical downstream transcription factor of the IIS. Specifically, we grew the N2, PL51, and PL54 strains on bacteria expressing scrambled control RNAi or daf-16 RNAi and used FUDR to prevent egg laying and cross generation sampling [29], [36]. We confirmed that PCM-1 overexpressing animals display on average an increase of 38% and 51% in median and maximum lifespan, respectively, compared to wild-type animals, and that no change in survival is observed between wild-type animals and nematodes expressing a catalytically inactive PCM-1 enzyme (Fig. 1, Table 2). In all strains tested, the lifespan values obtained in this study in the presence of FUDR were slightly reduced compared to those obtained previously in the absence of FUDR by Banfield et al. [17], but the same lifespan patterns were observed on control RNAi bacteria (Fig. 1, Table 2). All lifespan extensions (including the 50%, 25% and 10% survival values) measured for the overexpressor strain were statistically significant when compared to wild-type (N2) animals (Table 2).

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