Characteristics of concanavalin A‐resistant Chinese hamster ovary cells and certain revertants

Clones of Chinese hamster ovary (CHO) cells were isolated by single‐step selection for resistance to killing by Concanavalin A (ConA) and certain cellular and membrane properties were examined. The ConA‐resistant isolates were only about 2‐fold more resistant than wild type cells to the selecting lectin, but exhibited pleiotropic temperature‐sensitivity for growth, markedly altered morphology and adherence, and significant differences in susceptibility to other agents such as colchicine. Two revertants to full temperature‐resistance were isolated from different ConA‐resistant mutants. One revertant clone had reacquired wild type sensitivity to ConA while the other revertant remained ConA‐resistant. The two series of wild type, ConA‐resistant, and temperature revertant clones were analyzed for altered mobility of cell surface glycoproteins using lactoperoxidase/125I and galactose oxidase/[3H]borohydride labelling procedures. The ConA‐resistant clones showed increased mobility on polyacrylamide gels of three classes of labelled proteins, in the molecular weight ranges 225,000, 200,000, and 130,000 daltons. These changes persisted in the temperature‐revertant that remained ConA‐resistant, while two of the altered protein classes were restored to wild type mobility in the revertant that regained ConA‐sensitivity. Cell hybridization experiments indicated that the temperature‐sensitive phenotypes of different ConA‐resistant isolates are recessive and noncomplementing, implying that the same gene is affected in each case. The reversions to temperature resistance appear to be recessive suppressor mutations in different genes.

[1]  H. Ceri,et al.  Mammalian concanavalin A-resistant cells contain altered cell surface glycoproteins. , 1978, Experimental cell research.

[2]  H. Ceri,et al.  A correlation between concanavalin A resistance and specific alterations in growth and surface membrane-associated properties. , 1978, Experimental cell research.

[3]  D. Wirth,et al.  Proposal for a common oligosaccharide intermediate in the synthesis of membrane glycoproteins , 1977, Cell.

[4]  P. Stanley,et al.  Complementation between mutants of CHO cells resistant to a variety of plant lectins , 1977, Somatic cell genetics.

[5]  P. Stanley,et al.  Control of glycoprotein synthesis. Lectin-resistant mutant containing only one of two distinct N-acetylglucosaminyltransferase activities present in wild type Chinese hamster ovary cells. , 1977, The Journal of biological chemistry.

[6]  P. Robbins,et al.  Reduced synthesis of [14C]mannosyl oligosaccharide-lipid by membranes prepared from concanavalin A-resistant Chinese hamster ovary cells. , 1977, The Journal of biological chemistry.

[7]  R. Hynes,et al.  Restoration of normal morphology, adhesion and cytoskeleton in transformed cells by addition of a transformation-sensitive surface protein , 1977, Cell.

[8]  R. Hynes,et al.  Cell surface and metabolic labelling of the proteins of normal and transformed chicken cells. , 1976, Biochimica et biophysica acta.

[9]  I. Pastan,et al.  Cell surface protein partially restores morphology, adhesiveness, and contact inhibition of movement to transformed fibroblasts. , 1976, Proceedings of the National Academy of Sciences of the United States of America.

[10]  J. Wright.  Concanavalin A as a selective agent in tissue culture for temperature-sensitive hamster cell lines. , 1975, Canadian journal of microbiology.

[11]  P. Stanley,et al.  Selection and characterization of eight phenotypically distinct lines of lectin-resistant chinese hamster ovary cells , 1975, Cell.

[12]  P. Stanley,et al.  Chinese hamster ovary cells selected for resistance to the cytotoxicity of phytohemagglutinin are deficient in a UDP-N-acetylglucosamine--glycoprotein N-acetylglucosaminyltransferase activity. , 1975, Proceedings of the National Academy of Sciences of the United States of America.

[13]  S. Kornfeld,et al.  Deficient uridine diphosphate-N-acetylglucosamine:glycoprotein N-acetylglucosaminyltransferase activity in a clone of Chinese hamster ovary cells with altered surface glycoproteins. , 1975, The Journal of biological chemistry.

[14]  P. Stanley,et al.  Altered cell surface glycoproteins in phytohemagglutinin-resistant mutants of Chinese hamster ovary cells. , 1975, Biochimica et biophysica acta.

[15]  D. Critchley.  Cell surface proteins of NIL1 hamster fibroblasts labeled by a galactose oxidase, tritiated borohydride method. , 1974, Cell.

[16]  M. McBurney,et al.  Isolation and biochemical characterization of folate deficient mutants of Chinese hamster cells. , 1974, Cell.

[17]  W. Bonner,et al.  A film detection method for tritium-labelled proteins and nucleic acids in polyacrylamide gels. , 1974, European journal of biochemistry.

[18]  S. Kornfeld,et al.  Isolation of a clone of Chinese hamster ovary cells deficient in plant lectin-binding sites. , 1974, Proceedings of the National Academy of Sciences of the United States of America.

[19]  R. Hynes.  Alteration of cell-surface proteins by viral transformation and by proteolysis. , 1973, Proceedings of the National Academy of Sciences of the United States of America.

[20]  J. Wright.  EVIDENCE FOR PLEIOTROPIC CHANGES IN LINES OF CHINESE HAMSTER OVARY CELLS RESISTANT TO CONCANAVALIN A AND PHYTOHEMAGGLUTININ-P , 1973, The Journal of cell biology.

[21]  N. Sharon,et al.  Lectins: cell-agglutinating and sugar-specific proteins. , 1972, Science.

[22]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.

[23]  T. Puck,et al.  Genetics of somatic mammalian cells. IV. Properties of Chinese hamster cell mutants with respect to the requirement for proline. , 1967, Genetics.

[24]  Oliver H. Lowry,et al.  Protein measurement with the Folin phenol reagent. , 1951, The Journal of biological chemistry.

[25]  P. Stanley.  Surface Carbohydrate Alterations of Mutant Mammalian Cells Selected for Resistance to Plant Lectins , 1980 .

[26]  V. Ling,et al.  Membrane Mutants of Mammalian Cells in Culture , 1978 .

[27]  P. Stanley,et al.  Lectin receptors and lectin resistance in chinese hamster ovary cells. , 1977, Advances in experimental medicine and biology.

[28]  P. Stanley,et al.  Stable alterations at the cell membrane of Chinese hamster ovary cells resistant to the cytotoxicity of phytohemagglutinin , 1975, Somatic cell genetics.

[29]  G. Nicolson,et al.  The interactions of lectins with animal cell surfaces. , 1974, International review of cytology.

[30]  J. Till,et al.  Ouabain-resistant mutants of mouse and hamster cells in culture , 1974 .

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

[32]  L. Thompson,et al.  Isolation of mutants of cultured mammalian cells. , 1973, Methods in cell biology.

[33]  F. Studier.  Bacteriophage T7. , 1972, Science.