Effects of mutational loss of adenosine kinase and deoxycytidine kinase on deoxyATP accumulation and deoxyadenosine toxicity in cultured CEM human T-lymphoblastoid cells

M. S. Hershfield, J. E. Fetter, W. C. Small, A. S. Bagnara, S. R. Williams, Buddy Ullman, D. W. Martin, D. B. Wasson, D. A. Carson

Research output: Contribution to journalArticle

90 Citations (Scopus)

Abstract

Accumulation of deoxyadenosine nucleotides derived from deoxyadenosine may contribute to the lymphocyte-specific toxicity associated with heritable deficiency of adenosine deaminase. To determine which enzymes mediate phosphorylation of deoxyadenosine in intact T-lymphocytes, which are particularly sensitive to the growth inhibitory effects of deoxyadenosine, we have isolated from the CEM human T-lymphoblastoid cell line mutants that are deficient in adenosine kinase, deoxycytidine kinase, or both activities. Loss of deoxycytidine kinase alone abolished the ability to phosphorylate deoxyguanosine and caused a defect in capacity to accumulate deoxyadenosine nucleotides at low concentrations (20 μM. Deoxycytidine kinase deficiency alone decreased growth sensitivity to deoxyadenosine by ~3-fold, while loss of adenosine kinase had little effect on this sensitivity. Loss of both activities completely eliminated deoxyadenosine phosphorylation and decreased deoxyadenosine toxicity by ~100-fold. A mutant with complete deficiency of adenosine kinase, and with a deoxycytidine kinase that has altered Michaelis constants for both deoxycytidine and deoxyadenosine, was also incapable of phosphorylating deoxyadenosine, and was as resistant. These observations imply that in CEM T-lymphoblasts the toxicity of deoxyadenosine at concentrations below ~100 μM is mediated by deoxyadenosine nucleotides and that both deoxycytidine kinase and adenosine kinase participate in this phosphorylation. Levels of deoxyadenosine phosphorylating activities were compared in extracts of CEM and its kinase-deficient mutants and in extracts of the analogous WI-L2 human B-lymphoblast lines. The differences found were not sufficient to account for the much lower rates of deoxyadenosine nucleotide accumulation by the latter, suggesting the operation of mechanisms that prevent accumulation of these nucleotides in B-cells.

Original languageEnglish (US)
Pages (from-to)6380-6386
Number of pages7
JournalJournal of Biological Chemistry
Volume257
Issue number11
StatePublished - 1982
Externally publishedYes

Fingerprint

Deoxycytidine Kinase
Adenosine Kinase
Toxicity
Nucleotides
Phosphorylation
2'-deoxyadenosine
Cells
Deoxycytidine
Deoxyguanosine
Adenosine Deaminase
T-cells
Lymphocytes
Growth

ASJC Scopus subject areas

  • Biochemistry

Cite this

Hershfield, M. S., Fetter, J. E., Small, W. C., Bagnara, A. S., Williams, S. R., Ullman, B., ... 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 cells. Journal of Biological Chemistry, 257(11), 6380-6386.

Effects of mutational loss of adenosine kinase and deoxycytidine kinase on deoxyATP accumulation and deoxyadenosine toxicity in cultured CEM human T-lymphoblastoid cells. / Hershfield, M. S.; Fetter, J. E.; Small, W. C.; Bagnara, A. S.; Williams, S. R.; Ullman, Buddy; Martin, D. W.; Wasson, D. B.; Carson, D. A.

In: Journal of Biological Chemistry, Vol. 257, No. 11, 1982, p. 6380-6386.

Research output: Contribution to journalArticle

Hershfield, MS, Fetter, JE, Small, WC, Bagnara, AS, Williams, SR, Ullman, B, Martin, DW, Wasson, DB & Carson, DA 1982, 'Effects of mutational loss of adenosine kinase and deoxycytidine kinase on deoxyATP accumulation and deoxyadenosine toxicity in cultured CEM human T-lymphoblastoid cells', Journal of Biological Chemistry, vol. 257, no. 11, pp. 6380-6386.
Hershfield, M. S. ; Fetter, J. E. ; Small, W. C. ; Bagnara, A. S. ; Williams, S. R. ; Ullman, Buddy ; Martin, D. W. ; Wasson, D. B. ; Carson, D. A. / Effects of mutational loss of adenosine kinase and deoxycytidine kinase on deoxyATP accumulation and deoxyadenosine toxicity in cultured CEM human T-lymphoblastoid cells. In: Journal of Biological Chemistry. 1982 ; Vol. 257, No. 11. pp. 6380-6386.
@article{b4c4380a8da84d7dbe7f4248f7628df0,
title = "Effects of mutational loss of adenosine kinase and deoxycytidine kinase on deoxyATP accumulation and deoxyadenosine toxicity in cultured CEM human T-lymphoblastoid cells",
abstract = "Accumulation of deoxyadenosine nucleotides derived from deoxyadenosine may contribute to the lymphocyte-specific toxicity associated with heritable deficiency of adenosine deaminase. To determine which enzymes mediate phosphorylation of deoxyadenosine in intact T-lymphocytes, which are particularly sensitive to the growth inhibitory effects of deoxyadenosine, we have isolated from the CEM human T-lymphoblastoid cell line mutants that are deficient in adenosine kinase, deoxycytidine kinase, or both activities. Loss of deoxycytidine kinase alone abolished the ability to phosphorylate deoxyguanosine and caused a defect in capacity to accumulate deoxyadenosine nucleotides at low concentrations (20 μM. Deoxycytidine kinase deficiency alone decreased growth sensitivity to deoxyadenosine by ~3-fold, while loss of adenosine kinase had little effect on this sensitivity. Loss of both activities completely eliminated deoxyadenosine phosphorylation and decreased deoxyadenosine toxicity by ~100-fold. A mutant with complete deficiency of adenosine kinase, and with a deoxycytidine kinase that has altered Michaelis constants for both deoxycytidine and deoxyadenosine, was also incapable of phosphorylating deoxyadenosine, and was as resistant. These observations imply that in CEM T-lymphoblasts the toxicity of deoxyadenosine at concentrations below ~100 μM is mediated by deoxyadenosine nucleotides and that both deoxycytidine kinase and adenosine kinase participate in this phosphorylation. Levels of deoxyadenosine phosphorylating activities were compared in extracts of CEM and its kinase-deficient mutants and in extracts of the analogous WI-L2 human B-lymphoblast lines. The differences found were not sufficient to account for the much lower rates of deoxyadenosine nucleotide accumulation by the latter, suggesting the operation of mechanisms that prevent accumulation of these nucleotides in B-cells.",
author = "Hershfield, {M. S.} and Fetter, {J. E.} and Small, {W. C.} and Bagnara, {A. S.} and Williams, {S. R.} and Buddy Ullman and Martin, {D. W.} and Wasson, {D. B.} and Carson, {D. A.}",
year = "1982",
language = "English (US)",
volume = "257",
pages = "6380--6386",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "11",

}

TY - JOUR

T1 - Effects of mutational loss of adenosine kinase and deoxycytidine kinase on deoxyATP accumulation and deoxyadenosine toxicity in cultured CEM human T-lymphoblastoid cells

AU - Hershfield, M. S.

AU - Fetter, J. E.

AU - Small, W. C.

AU - Bagnara, A. S.

AU - Williams, S. R.

AU - Ullman, Buddy

AU - Martin, D. W.

AU - Wasson, D. B.

AU - Carson, D. A.

PY - 1982

Y1 - 1982

N2 - Accumulation of deoxyadenosine nucleotides derived from deoxyadenosine may contribute to the lymphocyte-specific toxicity associated with heritable deficiency of adenosine deaminase. To determine which enzymes mediate phosphorylation of deoxyadenosine in intact T-lymphocytes, which are particularly sensitive to the growth inhibitory effects of deoxyadenosine, we have isolated from the CEM human T-lymphoblastoid cell line mutants that are deficient in adenosine kinase, deoxycytidine kinase, or both activities. Loss of deoxycytidine kinase alone abolished the ability to phosphorylate deoxyguanosine and caused a defect in capacity to accumulate deoxyadenosine nucleotides at low concentrations (20 μM. Deoxycytidine kinase deficiency alone decreased growth sensitivity to deoxyadenosine by ~3-fold, while loss of adenosine kinase had little effect on this sensitivity. Loss of both activities completely eliminated deoxyadenosine phosphorylation and decreased deoxyadenosine toxicity by ~100-fold. A mutant with complete deficiency of adenosine kinase, and with a deoxycytidine kinase that has altered Michaelis constants for both deoxycytidine and deoxyadenosine, was also incapable of phosphorylating deoxyadenosine, and was as resistant. These observations imply that in CEM T-lymphoblasts the toxicity of deoxyadenosine at concentrations below ~100 μM is mediated by deoxyadenosine nucleotides and that both deoxycytidine kinase and adenosine kinase participate in this phosphorylation. Levels of deoxyadenosine phosphorylating activities were compared in extracts of CEM and its kinase-deficient mutants and in extracts of the analogous WI-L2 human B-lymphoblast lines. The differences found were not sufficient to account for the much lower rates of deoxyadenosine nucleotide accumulation by the latter, suggesting the operation of mechanisms that prevent accumulation of these nucleotides in B-cells.

AB - Accumulation of deoxyadenosine nucleotides derived from deoxyadenosine may contribute to the lymphocyte-specific toxicity associated with heritable deficiency of adenosine deaminase. To determine which enzymes mediate phosphorylation of deoxyadenosine in intact T-lymphocytes, which are particularly sensitive to the growth inhibitory effects of deoxyadenosine, we have isolated from the CEM human T-lymphoblastoid cell line mutants that are deficient in adenosine kinase, deoxycytidine kinase, or both activities. Loss of deoxycytidine kinase alone abolished the ability to phosphorylate deoxyguanosine and caused a defect in capacity to accumulate deoxyadenosine nucleotides at low concentrations (20 μM. Deoxycytidine kinase deficiency alone decreased growth sensitivity to deoxyadenosine by ~3-fold, while loss of adenosine kinase had little effect on this sensitivity. Loss of both activities completely eliminated deoxyadenosine phosphorylation and decreased deoxyadenosine toxicity by ~100-fold. A mutant with complete deficiency of adenosine kinase, and with a deoxycytidine kinase that has altered Michaelis constants for both deoxycytidine and deoxyadenosine, was also incapable of phosphorylating deoxyadenosine, and was as resistant. These observations imply that in CEM T-lymphoblasts the toxicity of deoxyadenosine at concentrations below ~100 μM is mediated by deoxyadenosine nucleotides and that both deoxycytidine kinase and adenosine kinase participate in this phosphorylation. Levels of deoxyadenosine phosphorylating activities were compared in extracts of CEM and its kinase-deficient mutants and in extracts of the analogous WI-L2 human B-lymphoblast lines. The differences found were not sufficient to account for the much lower rates of deoxyadenosine nucleotide accumulation by the latter, suggesting the operation of mechanisms that prevent accumulation of these nucleotides in B-cells.

UR - http://www.scopus.com/inward/record.url?scp=0020328813&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0020328813&partnerID=8YFLogxK

M3 - Article

C2 - 6281270

AN - SCOPUS:0020328813

VL - 257

SP - 6380

EP - 6386

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 11

ER -