Mechanisms of intracellular protein catabolism. Intracellular fate of microinjected polypeptides translated in vitro

Abstract

Erythrocyte-mediated microinjection was used to introduce [35S]polypeptides translated in vitro into 3T3-L1 cells. Such [35S]polypeptides are not degraded after loading into erythrocytes and are stable for the first 2 h after microinjection into growing 3T3-L1 cells. Similarly, little or no degradation of microinjected [35S]polypeptides is observed in either growing or confluent 3T3-L1 cells over a 70 h period. Microinjection of reticulocyte lysate alone does not affect the rate of degradation of long-lived endogenous protein. Reductively [3H]methylated lysate haemoglobin is degraded after microinjection by a cytosolic mechanism. Microinjected 125I-labelled bovine serum albumin is rapidly degraded by a cytosolic mechanism at the same rate in the absence or presence of reticulocyte lysate. The data do not support the notion that the observed lack of degradation of microinjected [35S]polypeptides translated in vitro is due to the presence of proteolytic inhibitors in reticulocyte lysates which can inhibit the degradation of microinjected or cellular proteins.