Mutants of CHO cells resistant to the protein synthesis inhibitors, cryptopleurine and tylocrebrine: genetic and biochemical evidence for common site of action of emetine, cryptopleurine, tylocrebine, and tubulosine.

Stable mutants resistant to the protein synthesis inhibitors cryptopleurine and tylocrebine can be isolated in Chinese hamster ovary (CHO) cells, in a single step. The frequency of occurrence of cryptopleurine (CryR) and tylocrebrine (TylR) resistant mutants in normal and mutagenized cell populations is similar to that observed for emetine resistant (EmtR) mutants. The CryR, TylR, and EmtR mutants exhibit strikingly similar cross-resistance to the three drugs used for selection, to tubulosine and also to two emetine derivatives cephaeline and dehydroemetine, based on assays of in vivo cytotoxicity and on assays of protein synthesis in cell-free extracts. The identity of cross-resistance patterns of the CryR, TylR, and EmtR mutants indicates that the resistance to all these compounds results from the same primary lesion, which in the case of EmtR cells has been shown to affect the 40S ribosomal subunit. This conclusion is strongly supported by the failure of EmtR, TylR, and CryR mutants to complement each other in somatic cell hybrids. Based on these results it is suggested that the above group of compounds possesses common structural determinants which are responsible for their activity. The above mutants, however, do not show any cross-resistance to other inhibitors of protein synthesis such as cycloheximide, trichodermin, anisomycin, pactamycin, and sparsomycin, either in vivo or in vitro, indicating that the site of action of these inhibitors is different from that of the emetine-like compounds.

[1]  L. Siminovitch,et al.  The isolation and preliminary characterization of somatic cell mutants resistant to the protein synthesis inhibitor—Emetine , 1976, Cell.

[2]  H. Munro,et al.  Mechanism of cycloheximide inhibition of protein synthesis in a cell-free system prepared from rat liver. , 1969, The Journal of biological chemistry.

[3]  D. Schindler,et al.  Trichodermin resistance—mutation affecting eukaryotic ribosomes , 1974, Nature.

[4]  S. Pestka.  Inhibitors of ribosome functions. , 1971, Annual review of microbiology.

[5]  A. Jiménez,et al.  Cryptopleurine Resistance: Genetic Locus for a 40S Ribosomal Component in Saccharomyces cerevisiae , 1974, Journal of bacteriology.

[6]  A. W. Murray,et al.  Inhibition of protein synthesis in Ehrlich ascites-tumour cells by the phenanthrene alkaloids tylophorine, tylocrebrine and cryptopleurine. , 1968, Biochemical and biophysical research communications.

[7]  A. Grollman,et al.  Cycloheximide resistance in yeast: a property of the 60s ribosomal subunit. , 1967, Biochemical and biophysical research communications.

[8]  A. Grollman.  Structural Basis for the Inhibition of Protein Biosynthesis: Mode of Action of Tubulosine , 1967, Science.

[9]  A. Grollman.  STRUCTURAL BASIS FOR INHIBITION OF PROTEIN SYNTHESIS BY EMETINE AND CYCLOHEXIMIDE BASED ON AN ANALOGY BETWEEN IPECAC ALKALOIDS AND GLUTARIMIDE ANTIBIOTICS* , 1966, Proceedings of the National Academy of Sciences of the United States of America.

[10]  A. Jimenez,et al.  Simultaneous ribosomal resistance to trichodermin and anisomycin in Saccharomyces cerevisiae mutants. , 1975, Biochimica et biophysica acta.

[11]  B. Abbott,et al.  Increased lifespan of leukemic mice treated with drugs related to (-)-emetine. , 1971, Chemotherapy.

[12]  C. Coltman,et al.  Phase I experience with emetine hydrochloride (NSC 33669) as an antitumor agent , 1971, Cancer.

[13]  A. Grollman,et al.  Mode of action of tylocrebrine: effects on protein and nucleic acid synthesis. , 1972, Molecular pharmacology.

[14]  Luis Carrasco,et al.  Specific inhibition of translocation by tubulosine in eukaryotic polysomes. , 1976, European journal of biochemistry.

[15]  E. Battaner,et al.  Inhibitors of protein synthesis by ribosomes of the 80-S type. , 1971, Biochimica et biophysica acta.

[16]  E. Maxwell,et al.  Peptidylpuromycin formation on mammalian polysomes. Studies on transpeptidation and translocation. , 1973, Biochemistry.

[17]  A. Grollman,et al.  Inhibitors of protein biosynthesis. V. Effects of emetine on protein and nucleic acid biosynthesis in HeLa cells. , 1967, The Journal of biological chemistry.

[18]  C. McLaughlin,et al.  Modification of Ribosomes in Cryptopleurine-Resistant Mutants of Yeast , 1973, Journal of bacteriology.

[19]  G. Klatskin,et al.  Emetine toxicity in man; studies on the nature of early toxic manifestations, their relation to the dose level, and their significance in determining safe dosage. , 1948, Annals of internal medicine.

[20]  J. Till,et al.  Pleiotropic phenotype of colchicine‐resistant CHO cells: Cross‐resistance and collateral sensitivity , 1976, Journal of cellular physiology.

[21]  Michael Sela,et al.  Selected highlights in immunological research in the last decade , 1974 .

[22]  L. Thompson,et al.  Chapter 7: Isolation of Mutants of Cultured Mammalian Cells , 1973 .

[23]  L. Siminovitch,et al.  The molecular basis of emetine resistance in chinese hamster ovary cells: Alteration in the 40S ribosomal subunit , 1977, Cell.

[24]  S. Pestka,et al.  Studies on transfer ribonucleic acid-ribosome complexes. XXI. Effect of antibiotics on peptidyl-puromycin synthesis by mammalian polyribosomes. , 1972, The Journal of biological chemistry.

[25]  D. Schindler,et al.  Chapter 2 Inhibitors of Macromolecular Synthesis in Yeast , 1975 .

[26]  B. Hardesty,et al.  The mechanism by which cycloheximide and related glutarimide antibiotics inhibit peptide synthesis on reticulocyte ribosomes. , 1971, The Journal of biological chemistry.