Fig. 3. UCP4C is necessary for period mutant lifespan and sufficient to extend wild-type lifespan.

Relative to controls (gray), per⁰¹ mutants (green) exhibited: a higher expression of Ucp4B and Ucp4C but not Ucp4A; and constitutively high expression of b Ucp4B and c Ucp4C, both of which are circadian-regulated in controls. Relative to controls (gray), per⁰¹ mutants (green) also exhibited the following phenotypes, which were reverted by suppression of Ucp4B/C expression, comparing flies with (dashed lines) or without (solid lines) piggyback mutation of Ucp4B/C: d increased leak respiration by purified mitochondria; e decreased mitochondrial membrane potential; f faster cold shock recovery; and g increased lifespan. Relative to vehicle-fed controls (gray), daGS > UAS-Ucp4C flies fed RU486 to induce constitutive UCP4C overexpression (magenta) exhibited: h higher leak respiration (p < 0.001); i lower mitochondrial membrane potential (p < 0.001); and j increased lifespan (p < 0.0001). k Ubiquitous overexpression of UCP4C in otherwise wild-type flies was sufficient to extend lifespan (gray, dashed) relative to driver-only controls (gray, solid). In per⁰¹ mutants (green, solid), overexpression of UCP4C did not further extend the lifespan of per⁰¹ mutants (green, dashed). See Supplementary Table 1 for n and statistical analysis of lifespans, particularly multicurve comparisons; ^n.s.p > 0.05, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.001; p values were obtained by unpaired two-tailed t-test (a, h, i), ANOVA followed by Tukey’s post-hoc test (d, e), and log-rank analysis (f, g, j, k); error bars represent SEM.