Sir2 mediates longevity in the fly through a pathway related to calorie restriction

Abstract

Calorie restriction can extend life span in a variety of species including mammals, flies, nematodes, and yeast. Despite the importance of this nearly universal effect, little is understood about the molecular mechanisms that mediate the life-span-extending effect of calorie restriction in metazoans. Sir2 is known to be involved in life span determination and calorie restriction in yeast mother cells. In nematodes increased Sir2 can extend life span, but a direct link to calorie restriction has not been demonstrated. We now report that Sir2 is directly involved in the calorie-restriction life-span-extending pathway in Drosophila. We demonstrate that an increase in Drosophila Sir2 (dSir2) extends life span, whereas a decrease in dSir2 blocks the life-span-extending effect of calorie reduction or rpd3 mutations. These data lead us to propose a genetic pathway by which calorie restriction extends life span and provides a framework for genetic and pharmacological studies of life span extension in metazoans.

Fig. 1.

Increased expression of dSir2 extends life span in the fly. Life span extension is seen in female (a, c, e, and g) and male (b, d, f, and h) flies in which dSir2 expression is increased (red line) in a ubiquitous manner (a–d) or in the nervous system (e–h) compared with genetically matched controls (blue line). Survivorship curves for genetically matched controls and tubulin-GAL4 driver/dSir2^EP2300 flies (a and b), tubulin-GAL4 driver/dSir2^EP2384 flies (c and d), ELAV-GAL4 driver/dSir2^EP2300 flies (e and f), and RU-486-ELAV-GeneSwitch driver/dSir2^EP2300 flies (g and h). Genetically matched controls are flies derived from a second-generation cross of the offspring from the cross that generated the experimental flies in a–f. Controls in a–f are flies containing a mix of the chromosomes found in the experimental flies but which do not contain the GAL4 chromosome, the UAS-containing chromosome, or any balancer chromosome. Controls in g and h are flies from the same cohort fed only diluent and not RU-486. Each life span included at least 149 male and 159 female flies (16).

Fig. 2.

Calorie-reduction life span extension is mediated by dSir2. (a and b) The expected life span extension with calorie reduction was blocked by mutations in dSir2. Survivorship curves of dSir2-null [dSir2^4.5/dSir2^5.26 (29)] (a) and dSir2-hypomorphic (dSir2^KG00871/dSir2^KG00871) (b) female flies on normal (red filled triangles) and low-calorie (green open boxes) food. Male flies mutant for dSir2 also had no life span extension in response to caloric reduction. (c–f) The life span of long-lived dSir2-overexpressing flies was not further extended by calorie reduction. Survivorship curves of long-lived ELAV-GAL4/dSir2^EP2300 (c and d) and tubulin-GAL4/dSir2^EP2300 (e and f) male (c and e) and female (d and f) flies on normal (red filled triangles) and low-calorie (green open boxes) food and those of genetically matched controls (c and d) on normal (blue open circles) and low-calorie (black X's) food. Low-calorie food was as described in ref. and Materials and Methods. Each life span included at least 179 male and 171 female flies (16).

Fig. 3.

Calorie-restriction/Rpd3/Sir2 pathway for life span extension in the fly. In this model a stimulus of calorie reduction results in a decrease in Rpd3 (16). The reduction in Rpd3 activity decreases the inhibition of Sir2, causing an increase in Sir2 (16). The increase in Sir2, in conjunction with the decrease in Rpd3, effects life span extension.

Fig. 4.

Mutations in dSir2 block the life-span-extending effect of rpd3 mutants. The extension in life span seen with rpd3 mutation was prevented by mutations in dSir2. Flies labeled as “rpd3” are heterozygous for the rpd3^def24-null allele of rpd3, whereas flies labeled as “dSir2 and rpd3” possess one copy of the life-span-extending rpd3-null mutation and one copy of either the dSir2¹⁷-null (28) or dSir2^EP2300-hypomorphic allele. (a) dSir2¹⁷. Flies labeled “rpd3,” “dSir2 and rpd3,” and “control” were each backcrossed to a Canton-S background. (b) dSir2^EP2300. Flies labeled “rpd3,” “dSir2 and rpd3,” and “control” were collected from the offspring of crosses between the rpd3-null allele, rpd3^def24, and dSir2^EP2300. Flies labeled “control” were derived from an additional cross to remove the rpd3 and dSir2^EP2300 alleles and any balancer chromosomes. Each life span included at least 200 male and 200 female flies (16).