Human diploid fibroblasts (IMR-90) regulate their overall rates of proteolysis in response to the composition of t h e culture medium and the ambient temperature. The magnitude and, in some cases, t h e direction of t h e response depend on the half-lives of the cellular proteins that are radioactively labeled and t h e time chosen for measurements of protein degradation. Fetal calf serum, insulin, fibroblast growth factor, epidermal growth factor, and amino acids selectively regulate catabolism of long-lived proteins without affecting degradation of short-lived proteins. Fetal calf serum reduces degradative rates of long-lived proteins and is maximally effective at a concentration of 20%, but the effect of serum on proteolysis is evident only for the first 24 hr. Insulin inhibits degradation of long-lived proteins in the presence or absence of glucose and amino acids in the medium, but is maximally effective only at high concentrations M). Amino acid deprivation increases degradative rates of long-lived proteins for t h e first 6 hr, but then decreases their catabolism for the subsequent 20 h r . Lowered temperature is the only condition tested that significantly alters degradative rates of short-lived proteins. Although cells incubated at 27Β°C have reduced rates of degradation for both short-lived and long-lived proteins compared to cells at 37"C, lowered temperature reduces catabolism of long-lived proteins to a greater extent.