Chaperoning extended life

Abstract

The capacity to moderate internal and external stress is arguably the central function regulating senescence in whole-animal ageing^1,2,3. During ageing, molecular chaperones such as heat-shock proteins are thought to combat stress-related senescent dysfunction^4,5. In transgenic Drosophila melanogaster, with varying copy numbers of the gene hsp70 encoding heat-shock protein hsp70, we found that heat-induced expression of hsp70 increased lifespan at normal temperatures. Only a brief, low level of expression was required to obtain a long-term improvement in survival.

Main

Thermally conditioned D. melanogaster and Caenorhabditis elegans exhibit greater longevity when mildly heated adults are returned to normal temperatures⁶,⁷. In D. melanogaster, such extended longevity is caused by an immediate increase in age-specific survival that persists for several weeks⁶. This change in survival is not easily explained by the removal of mortality costs of reproduction because these levels of thermal conditioning do not reduce egg laying. Rather, improved longevity may represent the modulation of age-related stress by the induced expression of heat-shock proteins.

We used transgenic D. melanogaster strains (from S. Lindquist), which varied in the number of copies of the inducible hsp70 gene⁸,⁹, to determine the effect of hsp70 protein on survival during ageing at normal temperatures. These allelic strains control for the effects of insertional mutagenesis. The ‘excision’ strain carried the wild-type complement of constitutive hsp70 and heat-inducible hsp70 genes. The ‘extra-copy’ strain added to the second chromosome a total of 12 additional copies of heat-inducible hsp70. The excision strain carried only a remnant P-element construct at the same integration site.

Four days after emergence of adult males, we heat-treated each strain with a brief, non-lethal 36 °C pulse of varying duration, and measured the subsequent expression of induced hsp70 protein, age-specific survival and remaining life expectancy at 24 °C. In independent trials we replicated these results, characterized the time-course of hsp70 protein induction, and observed the effects of longer heat treatments.

The presence of hsp70 protein increased subsequent survival at normal temperatures when expression exceeded 10-12% of standard (Fig. 1). The improvement in survival reached a plateau with higher levels of expressed protein. ‘Extra-copy’ but not ‘excision’ flies heated for 10 or 15 min expressed hsp70 and had improved survival over the two-week period after heat shock. Under these conditions, life expectancy increased by as much as 7.9% in ‘extra-copy’ flies, which is a substantial rise given the prevailing low mortality rates of young adults. Short heat treatments failed to induce hsp70 or to improve survival in the excision flies. These observations show that hsp70 moderates survival during subsequent ageing.

Figure 1: Effect of hsp70 expression on relative survivalin ‘extra-copy’ and ‘excision’ strains of Drosophila.

Within each strain, relative survival is calculated over the 14 days after heat shock compared with a control cohort of unheated flies (see ref. 6). Life tables were constructed from cohorts of about 1,200 flies per treatment. Heat treatments varied from 10 to 30 min, and durations that produced significant increases in survival are marked with asterisks. Levels of hsp70 protein were measured by enzyme-linked immunosorbent assay (ELISA)9. Arrow shows the background hsp70 level in unheated flies. Standard errors are shown for estimates of both variables.

Heat treatments of 20 and 30 min induced hsp70 to levels greater than 30% of standard in both strains and improved survival by similar amounts in each. Because the relationship between improved survival and hsp70 is similar in ‘extra-copy’ and ‘excision’ strains, we infer that the strains differ solely in their ability to express hsp70 protein for a given heat dose. This functional similarity also suggests that hsp70 copy number, and not merely differences in insert size, causes the improved survival after heat shock.

The whole-fly titre of induced hsp70 protein is transient, but its effect on age-specific survival is persistent. A transient expression of a molecular chaperone may increase age-specific survival through its ability to renature, assemble and disassemble many non-heat-shock proteins¹⁰, and to interact with other stress-response mechanisms such as superoxide dismutase⁵. It is also possible that hsp70 persists at high levels in some critical and specific tissue over the period of increased survival. Although hsp70 and heat-shock proteins in general are tightly regulated¹⁰, transient but effective levels of hsp70 could be present when stress is routinely encountered, and hsp70 may repair and restore higher order cell functions which themselves would otherwise accelerate senescence.