Skip to main content
NCBI home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1978 Mar;75(3):1289–1293. doi: 10.1073/pnas.75.3.1289

DNA polymerase activities in differentiating mouse neuroblastoma N-18 cells.

P Bhattacharya, S Basu
PMCID: PMC411456  PMID: 274718

Abstract

The activities of two DNA polymerases (DNA nucleotidyltransferases) were characterized in mouse neuroblastoma clone N-18 on the basis of their apparent molecular weights (determined by sucrose density gradient centrifugation: polymerase-alpha, 7.5-8 S; polymerase-beta, 3-4 S) and relative inhibition by sulfhydryl-blocking agents. N-Ethylmaleimide (10 mM) and iodoacetamide (1.5 mM) inhibited DNA polymerase-alpha activity completely, whereas only 35-40% inhibition was observed for DNA polymerase-beta under similar conditions. DNA polymerase-alpha activity was reduced 50-70% in N-18 cells that had been induced to differentiate by 4 micro M bromodeoxyuridine, and the low molecular weight DNA polymerase-beta activity remain unchanged. With activated calf thymus DNA as template, only DNA polymerase-alpha activity was stimulated in the presence of added ribonucleotides and purified Escherichia coli RNA polymerase.

Full text

PDF
1289

Images in this article

1290Image
on p.1290

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. APOSHIAN H. V., KORNBERG A. Enzymatic synthesis of deoxyribonucleic acid. IX. The polymerase formed after T2 bacteriophage infection of Escherichia coli: a new enzyme. J Biol Chem. 1962 Feb;237:519–525. [PubMed] [Google Scholar]
  2. Agarwal S. S., Loeb L. A. Studies on the induction of DNA polymerase during transformation of human lymphocytes. Cancer Res. 1972 Jan;32(1):107–113. [PubMed] [Google Scholar]
  3. Amano T., Richelson E., Nirenberg M. Neurotransmitter synthesis by neuroblastoma clones (neuroblast differentiation-cell culture-choline acetyltransferase-acetylcholinesterase-tyrosine hydroxylase-axons-dendrites). Proc Natl Acad Sci U S A. 1972 Jan;69(1):258–263. doi: 10.1073/pnas.69.1.258. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Arfellini G., Prodi G., Grilli S. Removal of 5-bromo-2-deoxyuridine incorporated in DNA of regenerating rat liver. Nature. 1977 Jan 27;265(5592):377–379. doi: 10.1038/265377a0. [DOI] [PubMed] [Google Scholar]
  5. Augusti-Tocco G., Sato G. Establishment of functional clonal lines of neurons from mouse neuroblastoma. Proc Natl Acad Sci U S A. 1969 Sep;64(1):311–315. doi: 10.1073/pnas.64.1.311. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bhattacharya P., Moskal J. R., Basu S. Embryonic chicken brain and mouse neuroblastoma cells N1E-115 and N-18 contain an inhibitor of acid deoxyribonuclease. Proc Natl Acad Sci U S A. 1977 Mar;74(3):842–845. doi: 10.1073/pnas.74.3.842. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Bick M. D., Davidson R. L. Nucleotide analysis of DNA and RNA in cells with thymidine totally replaced by bromodeoxyuridine. Somatic Cell Genet. 1976 Jan;2(1):63–76. doi: 10.1007/BF01539243. [DOI] [PubMed] [Google Scholar]
  8. Bick M. D., Soffer M. Altered glucose-6-phosphate dehydrogenase in bromodeoxyuridine-substituted cells. Nature. 1976 Apr 29;260(5554):788–791. doi: 10.1038/260788a0. [DOI] [PubMed] [Google Scholar]
  9. Bischoff R., Holtzer H. Inhibition of myoblast fusion after one round of DNA synthesis in 5-bromodeoxyuridine. J Cell Biol. 1970 Jan;44(1):134–150. doi: 10.1083/jcb.44.1.134. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Bollum F. J. Mammalian DNA polymerases. Prog Nucleic Acid Res Mol Biol. 1975;15(0):109–144. doi: 10.1016/s0079-6603(08)60118-x. [DOI] [PubMed] [Google Scholar]
  11. Chang L. M., Bollum F. J. A chemical model for transcriptional initiation of DNA replication. Biochem Biophys Res Commun. 1972 Feb 16;46(3):1354–1360. doi: 10.1016/s0006-291x(72)80124-4. [DOI] [PubMed] [Google Scholar]
  12. Chang L. M., Bollum F. J. Variation of deoxyribonucleic acid polymerase activities during rat liver regeneration. J Biol Chem. 1972 Dec 25;247(24):7948–7950. [PubMed] [Google Scholar]
  13. Chang L. M., Brown M., Bollum F. J. Induction of DNA polymerase in mouse L cells. J Mol Biol. 1973 Feb 15;74(1):1–8. doi: 10.1016/0022-2836(73)90349-5. [DOI] [PubMed] [Google Scholar]
  14. Chiu J. F., Sung S. C. Pattern of developmental changes in two DNA polymerases of rat brain. Biochim Biophys Acta. 1972 May 29;269(3):364–369. doi: 10.1016/0005-2787(72)90123-2. [DOI] [PubMed] [Google Scholar]
  15. Claycomb W. C. Biochemical aspects of cardiac muscle differentiation. Deoxyribonucleic acid synthesis and nuclear and cytoplasmic deoxyribonucleic acid polymerase activity. J Biol Chem. 1975 May 10;250(9):3229–3235. [PubMed] [Google Scholar]
  16. Daniel J. C. Changes in type of collagen synthesized by chick fibroblasts in vitro in the presence of 5-bromodeoxyuridine. Cell Differ. 1976 Dec;5(4):247–253. doi: 10.1016/0045-6039(76)90035-x. [DOI] [PubMed] [Google Scholar]
  17. Gontcharoff M., Mazia D. Developmental consequences of introduction of bromouracil into the DNA of sea urchin embryos during early division stages. Exp Cell Res. 1967 May;46(2):315–327. doi: 10.1016/0014-4827(67)90069-9. [DOI] [PubMed] [Google Scholar]
  18. Gonzalez R., Verly W. G. Isolation of an inhibitor of DNA synthesis specific for normal and malignant mammary cells. Proc Natl Acad Sci U S A. 1976 Jul;73(7):2196–2200. doi: 10.1073/pnas.73.7.2196. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hampar B., Derge J. G., Martos L. M., Walker J. L. Persistence of a repressed Epstein-Barr virus genome in Burkitt lymphoma cells made resistant to 5-bromodeoxyuridine. Proc Natl Acad Sci U S A. 1971 Dec;68(12):3185–3189. doi: 10.1073/pnas.68.12.3185. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Henderson E. E., Strauss B. Differences in the incorporation of bromodeoxyuridine by human lymphoblastoid cell lines. Cell. 1975 Aug;5(4):381–387. doi: 10.1016/0092-8674(75)90057-4. [DOI] [PubMed] [Google Scholar]
  21. Hill B. T., Tsuboi A., Baserga R. Effect of 5-bromodeoxyuridine on chromatin transcription in confluent fibroblasts. Proc Natl Acad Sci U S A. 1974 Feb;71(2):455–459. doi: 10.1073/pnas.71.2.455. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Holmer A. M., Hesslewood I. P., Johnston I. R. The occurrence of multiple activities in the high-molecular-weight DNA polymerase fraction of mammalian tissues. A preliminary study of some of their properties. Eur J Biochem. 1974 Apr 16;43(3):487–499. doi: 10.1111/j.1432-1033.1974.tb03436.x. [DOI] [PubMed] [Google Scholar]
  23. Holmes A. M., Hesslewood I. P., Johnston I. R. Evidence that DNA polymerase-alpha of calf thymus contains a subunit of molecular weight 155000. Eur J Biochem. 1976 Feb 16;62(2):229–235. doi: 10.1111/j.1432-1033.1976.tb10152.x. [DOI] [PubMed] [Google Scholar]
  24. Holtzer H., Weintraub H., Mayne R., Mochan B. The cell cycle, cell lineages, and cell differentiation. Curr Top Dev Biol. 1972;7:229–256. doi: 10.1016/s0070-2153(08)60073-3. [DOI] [PubMed] [Google Scholar]
  25. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  26. Levitt D., Dorfman A. Control of chondrogenesis in limb-bud cell cultures by bromodeoxyuridine. Proc Natl Acad Sci U S A. 1973 Aug;70(8):2201–2205. doi: 10.1073/pnas.70.8.2201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Levitt D., Dorfman A. The irreversible inhibition of differentiation of limb-bud mesenchyme by bromodeoxyuridine. Proc Natl Acad Sci U S A. 1972 May;69(5):1253–1257. doi: 10.1073/pnas.69.5.1253. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Luk D. C., Bick M. D. Determination of 5'-bromodeoxyuridine in DNA by buoyant density. Anal Biochem. 1977 Feb;77(2):346–349. doi: 10.1016/0003-2697(77)90247-0. [DOI] [PubMed] [Google Scholar]
  29. Matsukage A., Bohn E. W., Wilson S. H. Multiple forms of DNA polymerase in mouse myeloma. Proc Natl Acad Sci U S A. 1974 Feb;71(2):578–582. doi: 10.1073/pnas.71.2.578. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Meuth M., Green H. Induction of a deoxycytidineless state in cultured mammalian cells by bromodeoxyuridine. Cell. 1974 Jun;2(2):109–112. doi: 10.1016/0092-8674(74)90099-3. [DOI] [PubMed] [Google Scholar]
  31. Miura Y., Wilt F. H. The effects of 5-bromodeoxyuridine on yolk sac erythropoiesis in the chick embryo. J Cell Biol. 1971 Mar;48(3):523–532. doi: 10.1083/jcb.48.3.523. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Pawlowski P. J. Effect of 5-bromodeoxyuridine on the appearance of cytoplasmic poly-A containing RNA. J Cell Physiol. 1976 Sep;89(1):19–27. doi: 10.1002/jcp.1040890103. [DOI] [PubMed] [Google Scholar]
  33. Plevani P., Chang L. M. Enzymatic initiation of DNA synthesis by yeast DNA polymerases. Proc Natl Acad Sci U S A. 1977 May;74(5):1937–1941. doi: 10.1073/pnas.74.5.1937. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Prasad K. N., Hsie A. W. Morphologic differentiation of mouse neuroblastoma cells induced in vitro by dibutyryl adenosine 3':5'-cyclic monophosphate. Nat New Biol. 1971 Sep 29;233(39):141–142. doi: 10.1038/newbio233141a0. [DOI] [PubMed] [Google Scholar]
  35. Preisler H. S., Housman D., Scher W., Friend C. Effects of 5-bromo-2' -deoxyuridine on production of globin messenger RNA in dimethyl sulfoxide-stimulated Friend leukemia cells. Proc Natl Acad Sci U S A. 1973 Oct;70(10):2956–2959. doi: 10.1073/pnas.70.10.2956. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Roufa D. J. 5-bromodeoxyuridine-DNA strand symmetry and the repair of photolytic breaks in Chinese hamster cell chromosomes. Proc Natl Acad Sci U S A. 1976 Nov;73(11):3905–3909. doi: 10.1073/pnas.73.11.3905. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Schubert D., Jacob F. 5-bromodeoxyuridine-induced differentiation of a neuroblastoma. Proc Natl Acad Sci U S A. 1970 Sep;67(1):247–254. doi: 10.1073/pnas.67.1.247. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Schwartz S. A., Kirsten W. H. Distribution of 5-bromodeoxyuridine in the DNA of rat embryo cells. Proc Natl Acad Sci U S A. 1974 Sep;71(9):3570–3574. doi: 10.1073/pnas.71.9.3570. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Sedwick W. D., Shu-Fong Wang T., Korn D. "Cytoplasmic" deoxyribonucleic acid polymerase. Structure and properties of the highly purified enzyme from human KB cells. J Biol Chem. 1975 Sep 10;250(17):7045–7056. [PubMed] [Google Scholar]
  40. Smith R. G., Gallo R. C. DNA-dependent DNA polymerases I and II from normal human-blood lymphocytes. Proc Natl Acad Sci U S A. 1972 Oct;69(10):2879–2884. doi: 10.1073/pnas.69.10.2879. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Spadari S., Weissbach A. RNA-primed DNA synthesis: specific catalysis by HeLa cell DNA polymerase alpha. Proc Natl Acad Sci U S A. 1975 Feb;72(2):503–507. doi: 10.1073/pnas.72.2.503. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Tanabe K., Takahashi T. Studies on the molecular species of DNA polymerase extracted from rat ascites hepatoma cells. J Biochem. 1976 Jan;79(1):85–90. doi: 10.1093/oxfordjournals.jbchem.a131061. [DOI] [PubMed] [Google Scholar]
  43. Viola M. V., Norton P., Cole M. L. Nuclear DNA-dependent DNA polymerases in human brain. J Neurochem. 1977 Jan;28(1):121–126. doi: 10.1111/j.1471-4159.1977.tb07717.x. [DOI] [PubMed] [Google Scholar]
  44. Walther B. T., Pictet R. L., David J. D., Rutter W. J. On the mechanism of 5-bromodeoxyuridine inhibition of exocrine pancreas differentiation. J Biol Chem. 1974 Mar 25;249(6):1953–1964. [PubMed] [Google Scholar]
  45. Weissbach A., Baltimore D., Bollum F., Gallo R., Korn D. Nomenclature of eukaryotic DNA polymerases. Science. 1975 Oct 24;190(4212):401–402. doi: 10.1126/science.1179222. [DOI] [PubMed] [Google Scholar]
  46. Weissbach A. Eukaryotic DNA polymerases. Annu Rev Biochem. 1977;46:25–47. doi: 10.1146/annurev.bi.46.070177.000325. [DOI] [PubMed] [Google Scholar]
  47. YONEDA M., BOLLUM F. J. DEOXYNUCLEOTIDE-POLYMERIZING ENZYMES OF CALF THYMUS GLAND. I. LARGE SCALE PURIFICATION OF TERMINAL AND REPLICATIVE DEOXYNUCLEOTIDYL TRANSFERASES. J Biol Chem. 1965 Aug;240:3385–3391. [PubMed] [Google Scholar]
  48. de Recondo A. M., Abadiedebat J. Regenerating rat liver DNA polymerases: disimilitude or relationship between nuclear and cytoplasmic enzymes? Nucleic Acids Res. 1976 Aug;3(8):1823–1837. doi: 10.1093/nar/3.8.1823. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES