Altered degradation of intracellular proteins in aging human fibroblasts.

Confluent cultures of fibroblasts at different population doubling levels were incubated with [14C]leucine for 2 days and with [3H]leucine for 2 h to label long-lived and short-lived proteins, respectively. Proteolysis was then measured in the presence of excess unlabeled leucine to prevent reutilization of the isotope. Catabolism of long-lived proteins was reduced in senescent cells when measured in media without fetal bovine serum, insulin, fibroblast growth factor, or dexamethasone. In contrast, degradation of short-lived proteins was increased in senescent cells but only when measured in the presence of serum, hormones, and growth factors. Further experiments with cells of varying ages indicate that in unsupplemented medium half-lives of long-lived proteins lengthened by as much as 20 min per population doubling and in supplemented media half-lives of short-lived proteins decreased by 4 min per population doubling. The reduced catabolism of long-lived proteins in senescent cells cannot be explained by age-related changes in protein secretion or cell death during degradation measurements. These alterations in proteolysis may have major effects on protein content and composition in senescent cells.