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. Author manuscript; available in PMC: 2020 Feb 19.
Published in final edited form as: Nat Metab. 2019 Aug 19;1(8):790–810. doi: 10.1038/s42255-019-0097-9

Figure 8. Release of PGE2 by GSCs promotes cold-induced longevity.

Figure 8

a, PGE2 levels in fer-15;fem-1 control sterile worms at day 6 of adulthood (8.4 mg worms/ml, mean ± s.e.m. from 3 independent experiments). Two-tailed Student’s t-test for unpaired samples: * (P<0.05), ****(P<0.0001), NS= not significant (P=0.0519). b, Western blot of day 6-adult control sterile worms with antibodies to PGES-2 and α-tubulin loading control. The graph represents PGES-2 relative percentage values (corrected for α-tubulin) to 15°C (mean ± s.e.m., 4 independent experiments). Two-tailed Student’s t-test for unpaired samples: ***(P<0.001). c, Immunostaining with PGES-2 antibody. White and red arrows indicate intestine and germ line of day 6-adult wild-type worms, respectively. On the right, higher magnification of germ cells. Nuclei stained with DAPI. Scale bar, 20 μm. Images representative of three independent experiments. d, pges-2(ok3316) are short lived at cold temperature compared with wild-type animals (pges-2 15°C mean ± s.e.m: 30.10 ± 0.52, wild-type 15°C: 26.09 ± 0.66, P<0.0001), but do not live shorter at 20°C (pges-2 20°C: 18.13 ± 0.41, wild-type 20°C 19.11 ± 0.37, P=0.1303). e, cbs-1p::GFP in adult wild-type and pges-2(ok3316) mutant worms (day 5). DIC, differential interference contrast. Scale bar, 1000 μm. Images representative of three independent experiments. f, Quantification of cbs-1p::GFP in day 5-adult animals (mean ± s.e.m. relative to 15°C wild-type worms, 30 worms/condition from 3 independent experiments). Student’s t-test for unpaired samples: ****(P<0.0001), NS (P=0.0519). g, cbs-1p::GFP in day 8 adult-worms at 15°C. Exogenous PGE2 rescues low expression of cbs-1 in pges-2(ok3316) mutants. Scale bar, 200 μm. Images representative of two independent experiments. h, Quantification of cbs-1p::GFP in day 8-adult animals at 15°C (mean ± s.e.m. relative to non-treated wild-type worms. Wild-type (n=12) and pges-2 (n=15) from 2 independent experiments). Two-tailed Student’s t-test for unpaired samples: ****(P<0.0001), NS= wild-type versus wild-type + PGE2 (P=0.6614), wild-type versus pges-2 + PGE2 (P=0.4073). i, Exogenous PGE2 extends the short lifespan of pges-2(ok3316) mutants at 15°C (pges-2 versus pges-2 +PGE2, P<0.0001). Wild-type mean ± s.e.m: 28.07 ± 0.59; wild-type + PGE2: 28.60 ± 0.58; pges-2: 22.43 ± 0.73; pges-2 + PGE2: 26.54 ± 0.95. j, mrp-6 RNAi decreases lifespan of wild-type worms at 15°C (Vector RNAi mean ± s.e.m: 29.64 ± 0.59; mrp-6: 26.66 ± 0.61, P<0.0001), but not at 20°C (Vector: 21.23 ± 0.58; mrp-6: 21.58 ± 0.64, P=0.3707). RNAi was initiated during adulthood. k, Exogenous PGE2 does not prolong cold-induced longevity of control worms (Vector RNAi mean ± s.e.m: 29.85 ± 0.61 versus Vector + PGE2: 29.98 ± 0.59, P=0.9106), but extends the short lifespan of mrp-6 RNAi-treated worms (mrp-6: 26.37 ± 0.82 versus mrp-6 + PGE2: 31.43 ± 0.60, P<0.0001). l, Neuronal-specific mrp-6 RNAi (Vector RNAi: 23.87 ± 0.56, mrp-6: 24.18 ± 0.61, P=0.5119). m, Intestinal-specific mrp-6 RNAi (Vector RNAi: 24.86 ± 0.51, mrp-6: 26.16 ± 0.57, P=0.0531). n, When the germ line is sensitive to RNAi, mrp-6 RNAi reduces cold-induced longevity (Vector RNAi: 27.77 ± 1.03; mrp-6: 23.32 ± 0.78, P=0.0007). Lifespan experiments were analyzed by two-sided log-rank test, n= 96 worms/condition. Supplementary Data 3 contains replicate lifespan experiments.