H₂S increases thermotolerance in wild-type C. elegans. (A) Animals exposed to H₂S survive longer than untreated controls at high temperature. Nematodes were moved to 35°C in the same gaseous atmosphere in which they had been cultured (SI Fig. 5). The mean survival time of animals grown in H₂S was 65.5 h (solid line; n = 136), compared with 9.1 h (n = 96) for untreated controls (dashed line). (B) Prior exposure to H₂S is required to survive high temperature in H₂S. All animals were grown in room air without H₂S and then moved to 35°C in the presence or absence of 50-ppm H₂S. Animals first exposed to H₂S at high temperature had a mean survival time of 2.1 h (solid line; n = 20), whereas the control group exposed in room air survived for 7.3 h (dashed line; n = 20). (C) The continuous presence of H₂S in the atmosphere is required for increased survival at high temperature. Animals were exposed to 35°C in room air. Animals grown in H₂S before heat shock survived 7.3 h (solid line; n = 20), which is not significantly longer than untreated controls (dashed line; 7.0 h; n = 20). Indicated P values were determined by log-rank analysis.

H₂S increases lifespan in C. elegans. (A) Animals grown in H₂S live longer than untreated controls. The lifespan of animals was monitored in the same conditions in which they had developed. The mean lifespan of animals in H₂S was 22.6 ± 1.0 days (solid line; n = 80), compared with 13.0 ± 1.0 days for untreated controls in room air (dashed line; n = 40). Maximum lifespan also was increased. (B) Exposure to H₂S beginning as L4 does not increase lifespan. All animals were from populations grown in room air. The lifespan of animals moved into H₂S-containing environments at the beginning of the lifespan experiment (solid line) is 14.8 ± 0.3 days (n = 73), which is slightly shorter than controls that remained in house air (dashed line; mean lifespan 18.2 ± 0.4 days; n = 48). (C) Increased lifespan requires continuous exposure to H₂S. The lifespan of all animals was monitored in room air. The lifespan of animals raised in H₂S until L4 (solid line; 12.8 ± 0.7 days; n = 52) was indistinguishable from untreated controls (dashed line; 13.2 ± 0.7 days; n = 59). All lifespan experiments were performed at room temperature.

Thermotolerance of canonical long-lived mutants is increased by H₂S. Just as H₂S increases thermotolerance of wild-type worms (Fig. 1 and SI Fig. 6), long-lived mutants in canonical pathways that influence lifespan (20) also are more thermotolerant when grown in H₂S. All animals grown in H₂S were challenged with high temperature in H₂S (solid line), whereas the thermotolerance of untreated controls was assayed in room air (dashed line). (A) H₂S effects are genetically independent of IIS. The thermotolerance of daf-2(e1370) animals can be enhanced by exposure to H₂S, and daf-16(m26) mutants, which are defective in IIS, become thermotolerant when grown in H₂S. To facilitate the experiments shown in this figure, strains that show intrinsic thermotolerance, such as daf-2(e1370) (30), were tested at a slightly higher temperature both in room air and H₂S. However, the thermotolerance at 35°C also is increased when the animals are grown in H₂S (data not shown). (B) H₂S-induced thermotolerance is observed in isp-1(gk267) and clk-1(qm30) animals that are long-lived as a result of mitochondrial dysfunction. (C) H₂S-induced thermotolerance is observed in eat-2(ad1116) mutant animals, which have defects in pharyngeal pumping that result in dietary restriction.

sir-2.1 is required for increased thermotolerance and lifespan in H₂S. (A) H₂S does not increase thermotolerance of animals that have a deletion in sir-2.1. The mean survival time of sir-2.1(ok434) animals grown in H₂S and exposed to high temperature in H₂S (solid line) is 9.8 ± 0.3 h (n = 20), which is not significantly longer than untreated controls in room air (dashed line; mean survival 9.6 ± 0.3 h; n = 20). (B) H₂S does not increase the lifespan of sir-2.1(ok434) animals. The lifespan of sir-2.1(ok434) animals raised in H₂S is 20.0 ± 1.6 days (solid line; n = 47), which is statistically indistinguishable from control animals in room air (dashed line; 22.2 ± 1.2 days; n = 26). Indicated P values were determined by log-rank analysis.