scholarly journals EVIDENCE FOR PLEIOTROPIC CHANGES IN LINES OF CHINESE HAMSTER OVARY CELLS RESISTANT TO CONCANAVALIN A AND PHYTOHEMAGGLUTININ-P

1973 ◽  
Vol 56 (3) ◽  
pp. 666-675 ◽  
Author(s):  
J. A. Wright
Keyword(s):  
Concanavalin A ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Single Step ◽  
Cross Resistance ◽  
Wild Type ◽  
Con A ◽  
Intact Cells ◽  
Ovary Cells ◽  
Resistant Lines

Lines of Chinese hamster ovary cells resistant to the lectins concanavalin A (Con A) and phytohemagglutinin-P (PHA-P) have been isolated and characterized. Lines were isolated by a stepwise, a single-step, or a cycling single-step procedure, from both mutagen-treated and untreated cultures. The resistant lines showed a higher efficiency of colony formation in the presence of the appropriate lectin than did the wild-type parental line. The cell lines resistant to Con A did not exhibit any detectable cross resistance to PHA-P, nor did the PHA-resistant cells exhibit cross resistance to Con A. The toxicity of Con A from the wild-type and Con A-resistant lines was reduced in the presence of methyl α-D-glucopyranoside; this effect was not seen with the PHA-resistant line. Using 125I-labeled Con A, it was found that Con A was bound preferentially to the surface of intact cells, and that the amount of labeled Con A bound to intact cells was similar for the wild-type and lectin-resistant lines. The Con A-resistant lines were found to be more susceptible to the toxic effects of a number of different compounds, including cyclic AMP and its dibutyryl derivative, sodium butyrate, high concentrations of glucose, phenethyl alcohol, phenol, ouabain, and testosterone. It appears that, in these lines, acquisition of resistance to Con A gave rise to pleiotropic effects which were detected by changes in the sensitivity of the cells to a variety of agents.

Maytansine-resistant mutants of Chinese hamster ovary cells with an alteration in α-tubulin

1985 ◽  
Vol 63 (6) ◽  
pp. 503-510 ◽  
Author(s):  
Matthew J. Schibler ◽  
Fernando R. Cabral
Keyword(s):  
Chinese Hamster Ovary ◽  
Chinese Hamster Ovary Cells ◽  
Vinca Alkaloid ◽  
Chinese Hamster ◽  
Single Step ◽  
Cross Resistance ◽  
Two Dimensional ◽  
Wild Type ◽  
Electrophoretic Variants ◽  
Ovary Cells

Mutant clones of Chinese hamster ovary cells resistant to killing by the Vinca alkaloid maytansine have been isolated using a single-step procedure. These mutants are threefold more resistant to killing by the drug than the wild-type parent. The majority of the clones (30 of 34) probably contain alterations in membrane permeability based on their cross-resistance to an unrelated drug, puromycin. Two of the four puromycin-sensitive clones were found to contain "extra" spots which migrated close to α-tubulin on two-dimensional gels. The "extra" spots were shown to be electrophoretic variants of α-tubulin with an identical two-dimensional tryptic peptide map to that of the wild-type α-tubulin. The α-tubulin mutants were cross-resistant to other microtubule disrupting drugs such as griseofulvin, vinblastine, and colcemid, but were more sensitive to the microtubule-stabilizing agent taxol than the wild-type parental cells. Mutant – wild-type hybrids were found to be resistant to levels of maytansine intermediate between the lethal doses for mutant and wild-type cells. A possible explanation for the drug resistance of these mutants is discussed.

Formycin B-resistant mutants of Chinese hamster ovary cells: novel genetic and biochemical phenotype affecting adenosine kinase

1983 ◽  
Vol 3 (8) ◽  
pp. 1468-1477
Author(s):  
K D Mehta ◽  
R S Gupta
Keyword(s):  
Chinese Hamster Ovary ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Complex Form ◽  
Single Step ◽  
Adenosine Kinase ◽  
Cross Resistance ◽  
Ovary Cells ◽  
Cell Extracts

Stable mutants which are approximately three- and eightfold resistant to the pyrazolopyrimidine nucleosides formycin A and formycin B (FomR) have been selected in a single step from mutagenized Chinese hamster ovary cells. In cell extracts, the two FomR mutants which were examined were both found to contain no measurable activity of the enzyme adenosine kinase (AK). However, cross-resistance studies with other adenosine analogs such as toyocamycin and tubercidin show that these mutants are distinct from toyocamycin or tubercidin resistant (Toyr) mutants which also contain no measurable AK activity in cell extracts. Studies on the uptake and incorporation of [3H]adenosine and [3H]tubercidin by various mutants and parental cell lines show that unlike the Toyr mutants, which are severely deficient in the phosphorylation of these compounds, the FomR mutants possess nearly normal capacity to phosphorylate these compounds and incorporate them into cellular macromolecules. These results suggest that the FomR mutants contain normal levels of AK activity in vivo. In cell hybrids formed between FomR X FomS cells and FomR X Toyr cells, the formycin-resistant phenotype of both of the FomR mutants behaved codominantly. However, the extracts from these hybrid cells contained either congruent to 50% (FomR X FomS) or no measurable (FomR X Toyr) AK activity, indicating that the lesion in these mutants neither suppresses the wild-type AK activity nor complements the AK deficiency of the Toyr mutants. The presence of AK activity in the FomR mutants in vivo, but not in their cell extracts, along with the codominant behavior of the mutants in hybrids, indicates that the lesions in the FomR mutant are of a novel nature. It is suggested that the genetic lesion in these mutants affects AK activity indirectly and that it may involve an essential cellular function which exists in a complex form with AK. Some implications of these results regarding the mechanism of action of formycin B are discussed.

scholarly journals Formycin B-resistant mutants of Chinese hamster ovary cells: novel genetic and biochemical phenotype affecting adenosine kinase.

1983 ◽  
Vol 3 (8) ◽  
pp. 1468-1477 ◽  
Author(s):  
K D Mehta ◽  
R S Gupta
Keyword(s):  
Chinese Hamster Ovary ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Complex Form ◽  
Single Step ◽  
Adenosine Kinase ◽  
Cross Resistance ◽  
Ovary Cells ◽  
Cell Extracts

Stable mutants which are approximately three- and eightfold resistant to the pyrazolopyrimidine nucleosides formycin A and formycin B (FomR) have been selected in a single step from mutagenized Chinese hamster ovary cells. In cell extracts, the two FomR mutants which were examined were both found to contain no measurable activity of the enzyme adenosine kinase (AK). However, cross-resistance studies with other adenosine analogs such as toyocamycin and tubercidin show that these mutants are distinct from toyocamycin or tubercidin resistant (Toyr) mutants which also contain no measurable AK activity in cell extracts. Studies on the uptake and incorporation of [3H]adenosine and [3H]tubercidin by various mutants and parental cell lines show that unlike the Toyr mutants, which are severely deficient in the phosphorylation of these compounds, the FomR mutants possess nearly normal capacity to phosphorylate these compounds and incorporate them into cellular macromolecules. These results suggest that the FomR mutants contain normal levels of AK activity in vivo. In cell hybrids formed between FomR X FomS cells and FomR X Toyr cells, the formycin-resistant phenotype of both of the FomR mutants behaved codominantly. However, the extracts from these hybrid cells contained either congruent to 50% (FomR X FomS) or no measurable (FomR X Toyr) AK activity, indicating that the lesion in these mutants neither suppresses the wild-type AK activity nor complements the AK deficiency of the Toyr mutants. The presence of AK activity in the FomR mutants in vivo, but not in their cell extracts, along with the codominant behavior of the mutants in hybrids, indicates that the lesions in the FomR mutant are of a novel nature. It is suggested that the genetic lesion in these mutants affects AK activity indirectly and that it may involve an essential cellular function which exists in a complex form with AK. Some implications of these results regarding the mechanism of action of formycin B are discussed.

scholarly journals Cytoplasmic inheritance of oligomycin resistance in Chinese hamster ovary cells.

1980 ◽  
Vol 86 (3) ◽  
pp. 723-729 ◽  
Author(s):  
G A Breen ◽  
I E Scheffler
Keyword(s):  
Chinese Hamster Ovary ◽  
Doubling Time ◽  
Chinese Hamster Ovary Cells ◽  
Mitochondrial Protein ◽  
Chinese Hamster ◽  
Fold Increase ◽  
Cross Resistance ◽  
Mitochondrial Atpase ◽  
Wild Type ◽  
Ovary Cells

Oligomycin-resistant clones were isolated from Chinese hamster ovary cells by treatment of cells with ethidium bromide, followed by mutagenesis with ethylmethane sulfonate and selection in oligomycin. One clone (Olir 8.1) was chosen for further study. Olir 8.1 cells grow with doubling time similar to that of wild-type cells, whether grown in the presence or absence of drug (doubling time of 13-14 h). In plating efficiency experiments, Olir 8.1 cells are approximately 100-fold more resistant to oligomycin than are wild-type cells. There is approximately a 32-fold increase in the resistance to inhibition by oligomycin of the mitochondrial ATPase from Olir 8.1 cells. The electron transport chain is functional in Olir 8.1 cells. Oligomycin resistance is stable in the absence of selective pressure. There is little or no cross-resistance of Olir 8.1 cells to venturicidin and dicyclohexylcarbodiimide, other inhibitors of the mitochondrial ATPase, or to chloramphenicol, an inhibitor of mitochondrial protein synthesis. Oligomycin resistance is dominant in hybrids between Olir 8.1 cells and wild-type cells. Fusions of enucleated Olir 8.1 cells with sensitive cells and characterization of the resulting "cybrid" clones indicates that oligomycin resistance in Olir 8.1 cells is cytoplasmically inherited.

Role of concanavalin A as an endocytic modulator in Chinese hamster ovary cells

1981 ◽  
Vol 50 (1) ◽  
pp. 135-147
Author(s):  
B. Storrie ◽  
K.M. Maurey
Keyword(s):  
Concanavalin A ◽  
Chinese Hamster Ovary ◽  
Cho Cells ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Con A ◽  
Ovary Cells ◽  
Fibroblast Line ◽  
High Concentrations ◽  
Secondary Lysosomes

The effect of the lectin, concanavalin A (Con A), on pinocytic uptake and pinosome-lysosome fusion in Chinese hamster ovary (CHO) cells, a fibroblast line, was investigated. The glycosylated protein, horseradish peroxidase (HRPase), and the non-glycosylated protein, 125I-labelled bovine serum albumin ([125I]BSA), was used as endocytic tracers. Con A at high concentrations (greater than or equal to 50 micrograms/ml) promoted the uptake of HRPase and inhibited the degradation of ingested HRPase. Con A inhibited the degradation of HRPase whether the two were added simultaneously or at different times to the cultures. Fusion of HRPase-positive pinosomes with secondary lysosomes was observed by electron microscopy in Con A-treated CHO cells. Con A at 200 micrograms/ml had no effect on either the uptake or degradation of [125I]BSA. Together these observations strongly suggest that the effects of high Con A concentrations on the uptake and degradation of HRPase are a consequence of direct complex formation between lectin and glycoprotein. Con A does not appear to have a general modulating effect on the dynamics of endocytic membrane in CHO cells.

Malignancy-related characteristics of wild type and drug-resistant chinese hamster ovary cells

Pathology ◽  
1993 ◽  
Vol 25 (3) ◽  
pp. 268-276 ◽  
Author(s):  
Wanda B. Mackinnon ◽  
Marlen Dyne ◽  
Rebecca Hancock ◽  
Carolyn E. Mountford ◽  
Adrienne J. Grant ◽  
...  
Keyword(s):  
Chinese Hamster Ovary ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Drug Resistant ◽  
Wild Type ◽  
Ovary Cells

Complementation of a methotrexate uptake defect in Chinese hamster ovary cells by DNA-mediated gene transfer

1989 ◽  
Vol 9 (4) ◽  
pp. 1754-1758
Author(s):  
T M Underhill ◽  
W F Flintoff
Keyword(s):  
Gene Transfer ◽  
Dna Sequences ◽  
Chinese Hamster Ovary ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Ovary Cell ◽  
Wild Type ◽  
Ovary Cells ◽  
Ovary Cell Line ◽  
Human Specific

A methotrexate-resistant Chinese hamster ovary cell line deficient in methotrexate uptake has been complemented to methotrexate sensitivity by transfection with DNA isolated from either wild-type Chinese hamster ovary or human G2 cells. Primary and secondary transfectants regained the ability to take up methotrexate in a manner similar to that of wild-type cells, and in the case of those transfected with human DNA, to contain human-specific DNA sequences. The complementation by DNA-mediated gene transfer of this methotrexate-resistant phenotype provides a basis for the cloning of a gene involved in methotrexate uptake.

Mutation at autosomal loci of Chinese hamster ovary cells: involvement of a high-frequency event silencing two linked alleles

1983 ◽  
Vol 3 (7) ◽  
pp. 1172-1181
Author(s):  
W E Bradley
Keyword(s):  
Cell Lines ◽  
Chinese Hamster Ovary ◽  
High Frequency ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Low Frequency ◽  
Class Ii ◽  
Class I ◽  
Wild Type ◽  
Ovary Cells

Two classes of cell lines heterozygous at the galactokinase (glk) locus have been isolated from Chinese hamster ovary cells. Class I, selected by plating nonmutagenized wild-type cells at low density in medium containing 2-deoxygalactose at a partially selective concentration, underwent subsequent mutation to the glk-/- genotype at a low frequency (approximately 10(-6) per cell), which was increased by mutagenesis. Class II heterozygotes, isolated by sib selection from mutagenized wild-type cells, had a higher spontaneous frequency of mutation to the homozygous state (approximately 10(-4) per cell), which was not affected by mutagenesis. About half of the glk-/- mutants derived from a class II heterozygote, but not the heterozygote itself, were functionally hemizygous at the syntenic thymidine kinase (tk) locus. Similarly, a tk+/- heterozygote with characteristics analogous to the class II glk+/- cell lines underwent high-frequency mutation to tk-/-, and most of these mutants, but not the tk+/- heterozygote, were functionally hemizygous at the glk locus. A model is proposed, similar to that for the mutational events at the adenine phosphoribosyl transferase locus (W. E. C. Bradley and D. Letovanec, Somatic Cell Genet. 8:51-66, 1982), of two different events, high and low frequency, being responsible for mutation at either of the linked loci tk and glk. The low-frequency event may be a point mutation, but the high-frequency event, in many instances, involves coordinated inactivation of a portion of a chromosome carrying the two linked alleles. Class II heterozygotes would be generated as a result of a low-frequency event at one allele, and class I heterozygotes would be generated by a high-frequency event. Supporting this model was the demonstration that all class I glk+/- lines examined were functionally hemizygous at tk.

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