Abstract
Mutant sublines were derived of S49 mouse T-lymphoma cells that were resistant to tritiated deoxyadenosine. Twenty-five isolates that were selected in 1 µCi/ml of the nucleoside were cross-resistant to 6-thioguanine, were sensitive to HAT (hypoxanthine, aminopterin, and thymidine), and contained less than 1% of hypoxanthine phosphoribosyltransferase activity in wildtype cells. One of the mutant clones, S49-dA2, was further subjected to selection in a medium containing 2 µCi/ml tritiated deoxyadenosine and 1 µg/ml deoxycoformycin, an inhibitor of adenosine deaminase. All resistant subclones were cross-resistant to tubercidin, 6-methylmercaptopurine riboside, and arabinosyladenine. One of the subclones, S49-12, was completely devoid of adenosine kinase and was partially deficient in deoxyadenosine kinase. This subclone, however, contained wild-type levels of deoxycytidine kinase. DEAE chromatography of the wild-type cell extracts revealed two deoxyadenosine phosphorylating activities, one of which coeluted with adenosine kinase and was the enzyme missing in S49-12. The other species phosphorylated both deoxyadenosine and deoxycytidine, of which deoxycytidine was the preferred substrate.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Bradley, W. E. C., and Letovanc, D. (1982). High-frequency nonrandom mutational event at the adenine phosphoribosyltransferase (aprt) locus of site-selected CHO variants heterozygous for aprt. Somat. Cell Genet. 851.
Chan, T.-s. (1978). Deoxyguanosine toxicity on lymphoid cells as a cause for immunosuppression in purine nucleoside phosphorylase deficiency. Cell 14523.
Chan, T.-s. (1979). Purine excretion by mouse peritoneal macrophages lacking adenosine deaminase activity. Proc. Natl. Acad. Sci. USA 76925.
Chan, T.-s., Ishii, K., Long, C., and Green, H. (1973). Purine excretion by mammalian cells deficient in adenosine kinase. J. Cell. Physiol. 81315.
Chan, T.-s., Creagan, R. P., and Reardon, M. P. (1978). Adenosine kinase as a new selective marker in somatic cell genetics: Isolation of adenosine kinase-deficient mouse cell lines and human-mouse hybrid cell lines containing adenosine kinase. Somat. Cell Genet. 41.
Chen, T. R. (1977). In situ detection of mycoplasma contamination in cell cultures by fluorescent Hoechst 33258 stain. Exp. Cell Res. 104255.
Coffino, P., Baumal, R., Laskov, R., and Scharff, M. D. (1972). Cloning mouse myeloma cells and detection of variants. J. Cell. Physiol. 79429.
Cohen, A., Ullman, B., and Martin, D. W., Jr., (1979). Characterization of a mutant mouse lymphoma cell with deficient transport of purine and pyrimidine nucleosides. J. Biol. Chem. 254112.
Eves, E. M., and Farber, R. A. (1983). Expression of recessive aprt− mutations in mouse CAK cells resulting from chromosome loss and duplication. Somat. Cell Genet. 9771.
Francke, U., and Gehring, U. (1980). Chromosome assignment of a murine glucocorticoid receptor gene (Grl-1) using intraspecies somatic cell hybrids. Cell 22657.
Gower, W. R., Jr., Carr, M. C., and Ives, D. H. (1979). Deoxyguanosine kinase. Distinct molecular forms in mitochondria and cytosol. J. Biol. Chem. 2542180.
Green, H., and Chan, T.-s. (1973). Pyrimidine starvation induced by adenosine in fibroblasts and lymphoid cells: Role of adenosine deaminase. Science 182836.
Hartwick, R. A., and Brown, P. R. (1975). The performance of microparticle chemically-bonded anion-exchange resins in the analysis of nucleotides. J. Chromatogr. 112651.
Hershfield, M. S., Fetter, J. E., Small, W. C., Bagnara, A. S., Williams, S. R., Ullman, B., Martin, D. W., Jr., Wasson, D. B., and Carson, D. A. (1982). Effects of mutational loss of adenosine kinase and deoxycytidine kinase on deoxyATP accumulation and deoxyadenosine toxicity in cultured CEM human T-lymphoblastoid cell. J. Biol. Chem. 2576380.
Horibata, K., and Harris, A. W. (1970). Mouse myelomas and lymphomas in culture. Exp. Cell Res. 6061.
Ishii, K., and Green, H. (1973). Lethality of adenosine for cultured mammalian cells by interference with pyrimidine biosynthesis. J. Cell Sci. 13429.
Klobutcher, L. A., Nichols, E. A., Kucherlapati, R. S., and Ruddle, F. H. (1976). Assignment of the gene for human adenosine kinase to chromosome 10 using a somatic cell hybrid clone panel. Cytogenet. Cell Genet. 15129.
Kredich, N. M., and Martin, D. W., Jr. (1977). Role of S-adenosylhomocysteine in adenosine-mediated toxicity in cultured mouse T lymphoma cells. Cell 12931.
Kusano, T., Long, C., and Green, H. (1971). A new reduced human-mouse somatic cell hybrid containing the human gene for adenine phosphoribosyltransferase. Proc. Natl. Acad. Sci. USA 6882.
Littlefield, J. (1964). Selection of hybrids from matings of fibroblasts in vitro and their presumed recombinants. Science 145709.
Long, C., Chan, T., Levytska, V., Kusano, T., and Green, H. (1973). Absence of demonstrable linkage of human genes for enzymes of the purine and pyrimidine salvage pathways in human-mouse somatic cell hybrids. Biochem. Genet. 9283.
Medrano, L., and Green, H. (1974). A uridine kinase-deficient mutant of 3T3 and a selective method for cells containing the enzyme. Cell 123.
Meuth, M., and Arrand, J. E. (1982). Alterations of gene structure in ethyl methane sulfonateinduced mutants of mammalian cells. Mol. Cell. Biol. 21459.
Meuth, M., and Green, H. (1974). Alteration leading to increased ribonucleotide reductase in cells selected for resistance to deoxynucleosides. Cell 3367.
Momparler, R. L., Chu, M., and Fischer, G. A. (1968). Studies on a new mechanism of resistance of L5178Y murine leukemia cells to cytosine arabinoside. Biochim. Biophys. Acta 161481.
Morrow, J. (1983). Eukaryotic Cell Genetics Academic Press, New York, pp. 21–23.
Siaw, M. F. E., Mitchell, B. S., Koller, C. A., Coleman, M. S., and Hutton, J. J. (1980). ATP depeletion as a consequence of adenosine deaminase inhibition in man. Proc. Natl. Acad. Sci. USA 776157.
Thompson, L. H., and Baker, R. M. (1973). Isolation of mutants of cultured mammalian cells. Methods Cell Biol. 6209.
Ullman, B., Cohen, A., and Martin, D. W., Jr. (1976). Characterization of a cell culture model for the study of adenosine deaminase and purine nucleoside phosphorylase-deficient immunologic disease. Cell 9205.
Ullman, B., Gudas, L. J., Cohen, A., and Martin, D. W., Jr. (1978). Deoxyadenosine metabolism and cytotoxicity in cultured mouse T lymphoma cells: A model for immunodeficiency disease. Cell 14365.
Ullman, B., Levinson, B. B., Hershfield, M. S., and Martin, D. W., Jr. (1981). A biochemical genetic study of the role of specific nucleoside kinases in deoxyadenosine phosphorylation by cultured human cells. J. Biol. Chem. 256848.
Urlaub, G., and Chasin, L. A. (1980). Isolation of Chinese hamster cell mutants deficient in dihydrofolate reductase activity. Proc. Natl. Acad. Sci. USA 774216.
Verhoef, V., Sarup, J., and Friedland, A. (1981). Identification of the mechanism of activation of 9-β-D-arabinosyladenine in human lymphoid cells using mutants deficient in nucleoside kinase. Cancer Res. 414478.
Yandell, D. W., Dryja, T. P., and Little, J. B. (1986). Somatic mutations at a heterozygous autosomal locus in human cells occur more frequently by allele loss than by intragenic structural alterations. Somat. Cell Mol. Genet. 12255.
Author information
Authors and Affiliations
Additional information
This research was supported in part by NIH Grants GM26522 and GM27589.
Rights and permissions
About this article
Cite this article
Jagannadha Sastry, K., Huang, C. & Chan, Ts. Adenosine kinase deficiency in tritiated deoxyadenosine-resistant mouse S49 lymphoma cell lines. Biochem Genet 25, 765–777 (1987). https://doi.org/10.1007/BF00502597
Received:
Revised:
Issue Date:
DOI: https://doi.org/10.1007/BF00502597