Import and processing of cytochrome b-c1 complex subunits in isolated hepatoma ascites cells. Inhibition by Rhodamine 6G.
The import and processing of cytochrome c1 and the iron sulfur protein of the cytochrome b-c1 complex were studied in Zajdela hepatoma ascites cells. Both peptides were synthesized as larger percursor molecules which were approximately 2-3 kDa and 5-6 kDa larger than the mature forms of apocytochrome c1 and apo-iron sulfur protein, respectively. Comparison of these precursors to those reported for functionally homologous peptides in yeast and Neurospora indicate significant size changes have occurred in mammals. Rhodamine 6G, a specific vital stain for mitochondria, is a potent inhibitor of precursor processing in isolated hepatoma cells. Both precursor to cytochrome c1 and precursor to FeS accumulate in the soluble and particulate fractions obtained by digitonin treatment of tumor cells treated with Rhodamine 6G. Appearance of the mature peptides was abolished. The precursors are unstable, however, and disappear from the cytosolic and membrane fractions during a 10 min chase. Comparison of the effects of Rhodamine 6G and carbonylcyanide m-chlorophenylhydrazone on precursor processing shows that: (a) Rhodamine 6G is a more effective inhibitor of processing, (b) it has less of an inhibitory effect on cellular protein synthesis, and (c) it inhibits processing under conditions in which it appears to have little influence on coupled respiration in whole cells. The data suggest that the most likely mode of action of Rhodamine 6G is on the matrix processing step.
D000818:Animals; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000494:pharmacology; D014450:Electron Transport Complex III; D004792:Enzyme Precursors / Q000378:metabolism; D007120:Immunochemistry; D066298:In Vitro Techniques; D008114:Liver Neoplasms, Experimental / Q000201:enzymology*; D008297:Male; D009097:Multienzyme Complexes / Q000378:metabolism*; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism*; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D011808:Quinone Reductases / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D012235:Rhodamines / Q000494:pharmacology*; D014966:Xanthenes / Q000494:pharmacology*
D004792:Enzyme Precursors; D009097:Multienzyme Complexes; D012235:Rhodamines; D014966:Xanthenes; C026188:rhodamine 6G; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; D009247:NADH, NADPH Oxidoreductases; D011808:Quinone Reductases; D014450:Electron Transport Complex III