Mitochondrial matrix localization of a protein altered in mutants resistant to the microtubule inhibitor podophyllotoxin.
Specific antibodies to a protein designated P1 (Mr approximately equal to 63,000), which is specifically altered in mutants resistant to the microtubule inhibitor podophyllotoxin, bind to mitochondria in cells of various vertebrate and invertebrate species (Eur. J. Cell Biol. 44, 278-285 (1987); Can. J. Biochem. Cell Biol. 63, 489-502 (1985)). To investigate the relationship of this protein to mitochondria, rat liver mitochondria have been purified and immunoblot analysis with these provide evidence that the P1 protein is a major component of mitochondria. Two-dimensional gel electrophoretic analysis of mitochondrial proteins from Chinese hamster ovary (CHO) cells also show the P1 protein to be a major mitochondrial component. Subfractionation of rat liver mitochondria into various compartments indicates that the P1 protein is mainly associated with the matrix fraction. Effect of treatment of CHO cells with mitochondrial inhibitors on the synthesis of P1 protein was also investigated. Treatment with the K+ ionophores nonactin and valinomycin, which abolish mitochondrial membrane potential, inhibited synthesis of the mature forms of the P1 protein as well as a number of other mitochondrial proteins, as seen by two-dimensional gel electrophoresis of labeled polypeptides. Treatment of the podophyllotoxin-resistant mutant of CHO cells with the above inhibitors affected both the wild-type and the mutant forms of the P1 protein in a similar manner. Concomitant with the disappearance of the above proteins, new basic proteins of higher molecular masses, related to the P1 and other proteins by peptide analysis, were observed in the drug-treated cells.(ABSTRACT TRUNCATED AT 250 WORDS)
Gupta
R S
RS
Department of Biochemistry, McMaster University, Hamilton, Ontario, Canada.
Austin
R C
RC
D000818:Animals; D002460:Cell Line; D004351:Drug Resistance; D008297:Male; D008869:Microtubule-Associated Proteins / Q000032:analysis* / Q000096:biosynthesis; D008870:Microtubules / Q000187:drug effects*; D008928:Mitochondria / Q000032:analysis*; D008930:Mitochondria, Liver / Q000032:analysis; D009154:Mutation; D011034:Podophyllotoxin / Q000494:pharmacology*; D051381:Rats; D011919:Rats, Inbred Strains; D013367:Submitochondrial Particles / Q000032:analysis
D008869:Microtubule-Associated Proteins; D011034:Podophyllotoxin