The dynamics of ubiquitin pools within cultured human lung fibroblasts.
The dynamics of ubiquitin pools within the cultured human lung fibroblast line IMR-90 were examined using solid phase immunochemical methods to quantitate free and conjugated polypeptide. Fetal calf serum was found to contain a nondialyzable factor that induced a transient accumulation of ubiquitin. During the induction, free and conjugated ubiquitin pools changed in concert so that the fraction conjugated remained constant. The induction of ubiquitin by the serum factor resulted from an enhanced rate of protein synthesis. Within experimental error no change in the first order rate constant for intracellular ubiquitin degradation was observed. Pulse-chase studies revealed ubiquitin to turn over with a half-life of 28-31 h in conditioned and freshly fed cultures. Withdrawal of serum from cultures led to a rapid decline in total ubiquitin during which the fractional level of conjugation remained constant. The accelerated ubiquitin turnover following removal of serum likely involves lysosomal autophagy since 10 mM NH4Cl led to an accumulation of the polypeptide. Since no similar effect of the lysosomotropic compound was observed in conditioned or freshly fed cultures, nonlysosomal processes are probably responsible for ubiquitin turnover under nutritional balance. The dynamics of these intracellular pools suggests that the ubiquitin ligation system is subject to regulatory constraints not previously suspected. The short half-life for ubiquitin is consistent with the apparent ability of cells to alter ubiquitin levels in response to external stimuli and stress.
D000643:Ammonium Chloride / Q000494:pharmacology; D000818:Animals; D002417:Cattle; D002460:Cell Line; D003470:Culture Media; D005312:Fetal Blood; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism*; D006207:Half-Life; D006801:Humans; D008168:Lung / Q000378:metabolism*; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D014452:Ubiquitins / Q000096:biosynthesis / Q000378:metabolism*
D003470:Culture Media; D014452:Ubiquitins; D000643:Ammonium Chloride