Metabolic properties of the products of mitochondrial protein synthesis in HeLa cells.
The metabolic behavior of the mitochondrial protein synthesis products has been investigated in HeLa cells. Particular attention was given to the four major electrophoretic components (designated as Nos. 2, 3, 5, and 8) of the 10 previously identified as organelle-specific products. Inhibition of cytoplasmic protein synthesis with emetine or cycloheximide causes a rapid decline in the rate of mitochondrial protein synthesis, with an estimated half-life of 1 to 2 h, affecting in a parallel way all the discrete components. About 30% of the original synthetic activity appears to be resistant to emetine treatment for at least 24 h; however, all the polypeptides synthesized after the first 4 h of cell exposure to emetine are metabolically unstable, possibly because of lack of integration into the inner mitochondrial membrane. An analysis of the stability of newly synthesized products of mitochondrial protein synthesis pulse-labeled in the presence of cycloheximide and then chased in the absence of the drug (i.e. under conditions of resumed cytoplasmic protein synthesis) has revealed marked differences among the various discrete components. In particular, about three-fourths of the radioactivity associated with components 3 and 5 decays within 4 h of chase, the remainder being substantailly stable afterwards; by contrast, the radioactivity in components 2 and 8 shows only a slow decline during a 3-day chase. If the chase is carried out under conditions of a persistent block of cytoplasmic protein synthesis, as is the situation after a pulse labeling in the presence of emetine, all newly synthesized components appear to be destablized in various degrees, with the exception of component 5, which is to a great extent stabilized. Inhibition of mitochondrial protein synthesis with chloramphenicol has a progressive stabilizing effect on most of the discrete components newly synthesized after removal of the drug; this effect is especially striking in the case of component 5 which, in experiments of continuous labeling in the presence of emetine after prolonged chloramphenicol treatment, becomes, after 24 h of labeling or more, the only recognizable peak in the electrophoretic pattern of the sodium dodecyl sulfate-lysed mitochondrial fraction. The results of the kinetic experiments described here are interpreted in terms of two roles of cytoplasmically synthesized proteins, one required for the synthesis of polypeptides within the organelles, the other necessary for the stabilization of the mitochondrial products.
D002455:Cell Division / Q000187:drug effects; D003513:Cycloheximide / Q000494:pharmacology; D006367:HeLa Cells / Q000187:drug effects / Q000378:metabolism*; D007700:Kinetics; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism*; D009363:Neoplasm Proteins / Q000378:metabolism*; D014176:Protein Biosynthesis / Q000187:drug effects; D013997:Time Factors
D009363:Neoplasm Proteins; D003513:Cycloheximide