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
. 1988 Mar;85(5):1442–1446. doi: 10.1073/pnas.85.5.1442

Activation of cAMP-dependent protein kinase is required for heterologous desensitization of adenylyl cyclase in S49 wild-type lymphoma cells.

R B Clark 1, M W Kunkel 1, J Friedman 1, T J Goka 1, J A Johnson 1
PMCID: PMC279787  PMID: 2830619

Abstract

We report here that, contrary to previously reported findings, treatment of S49 wild-type (WT) lymphoma cells with 0-50 nM epinephrine resulted in a heterologous desensitization of adenylyl cyclase (EC 4.6.1.1)--that is, epinephrine and prostaglandin E1 (PGE1) stimulations of adenylyl cyclase were reduced. Observation of this heterologous desensitization required the assay of adenylyl cyclase with submillimolar concentrations of Mg2+ and low concentrations of epinephrine. Also, whereas previously there had been no evidence for any role of cAMP-dependent protein kinase in the desensitization of the WT beta-adrenergic receptor, our data comparing the characteristics of the desensitization in WT, kin-, and cyc- lymphoma cells [where kin- and cyc- refer to variants of S49 WT cells lacking cAMP-dependent protein kinase activity (kin-) and the alpha subunit of the stimulatory guanine nucleotide-binding regulatory protein (cyc-)] now suggest that cAMP-dependent protein kinase mediates the heterologous desensitization of adenylyl cyclase. Specifically, we found that only the WT cells exhibited epinephrine-induced heterologous desensitization. The kin- and cyc- cells exhibited only homologous desensitization, and much higher concentrations of epinephrine were required to elicit the homologous desensitization in the variants relative to the heterologous desensitization of the WT. Treatment of WT and cyc- cells with dibutyryl cAMP or treatment of WT with forskolin or PGE1 caused the heterologous desensitization of adenylyl cyclase, indicating that neither receptor occupancy nor activation of adenylyl cyclase was necessary for the heterologous desensitization.

Full text

PDF
1442

Selected References

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

  1. Barber R. Discrimination between intact cell desensitization and agonist affinity changes. Mol Cell Endocrinol. 1986 Aug;46(3):263–270. doi: 10.1016/0303-7207(86)90008-0. [DOI] [PubMed] [Google Scholar]
  2. Barovsky K., Pedone C., Brooker G. Forskolin-stimulated cyclic AMP accumulation mediates protein synthesis-dependent refractoriness in C6-2B rat glioma cells. J Cyclic Nucleotide Protein Phosphor Res. 1983;9(3):181–189. [PubMed] [Google Scholar]
  3. Benovic J. L., Strasser R. H., Caron M. G., Lefkowitz R. J. Beta-adrenergic receptor kinase: identification of a novel protein kinase that phosphorylates the agonist-occupied form of the receptor. Proc Natl Acad Sci U S A. 1986 May;83(9):2797–2801. doi: 10.1073/pnas.83.9.2797. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Clark R. B. Desensitization of hormonal stimuli coupled to regulation of cyclic AMP levels. Adv Cyclic Nucleotide Protein Phosphorylation Res. 1986;20:151–209. [PubMed] [Google Scholar]
  5. Clark R. B., Friedman J., Johnson J. A., Kunkel M. W. Beta-adrenergic receptor desensitization of wild-type but not cyc lymphoma cells unmasked by submillimolar Mg2+. FASEB J. 1987 Oct;1(4):289–297. doi: 10.1096/fasebj.1.4.2820824. [DOI] [PubMed] [Google Scholar]
  6. Clark R. B., Friedman J., Prashad N., Ruoho A. E. Epinephrine-induced sequestration of the beta-adrenergic receptor in cultured S49 WT and cyc- lymphoma cells. J Cyclic Nucleotide Protein Phosphor Res. 1985;10(1):97–119. [PubMed] [Google Scholar]
  7. Clark R. B., Goka T. J., Green D. A., Barber R., Butcher R. W. Differences in the forskolin activation of adenylate cyclases in wild-type and variant lymphoma cells. Mol Pharmacol. 1982 Nov;22(3):609–613. [PubMed] [Google Scholar]
  8. Erdos J. J., Maguire M. E. Hormone-sensitive magnesium transport in murine S49 lymphoma cells: characterization and specificity for magnesium. J Physiol. 1983 Apr;337:351–371. doi: 10.1113/jphysiol.1983.sp014628. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Green D. A., Clark R. B. Adenylate cyclase coupling proteins are not essential for agonist-specific desensitization of lymphoma cells. J Biol Chem. 1981 Mar 10;256(5):2105–2108. [PubMed] [Google Scholar]
  10. Green D. A., Friedman J., Clark R. B. Epinephrine desensitization of adenylate cyclase from cyc- and S49 cultured lymphoma cells. J Cyclic Nucleotide Res. 1981;7(3):161–172. [PubMed] [Google Scholar]
  11. Grubbs R. D., Collins S. D., Maguire M. E. Differential compartmentation of magnesium and calcium in murine S49 lymphoma cells. J Biol Chem. 1984 Oct 10;259(19):12184–12192. [PubMed] [Google Scholar]
  12. Hertel C., Perkins J. P. Receptor-specific mechanisms of desensitization of beta-adrenergic receptor function. Mol Cell Endocrinol. 1984 Oct;37(3):245–256. doi: 10.1016/0303-7207(84)90094-7. [DOI] [PubMed] [Google Scholar]
  13. Hoyer D., Reynolds E. E., Molinoff P. B. Agonist-induced changes in the properties of beta-adrenergic receptors on intact S49 lymphoma cells. Time-dependent changes in the affinity of the receptor for agonists. Mol Pharmacol. 1984 Mar;25(2):209–218. [PubMed] [Google Scholar]
  14. Iyengar R., Birnbaumer L. Hormone receptor modulates the regulatory component of adenylyl cyclase by reducing its requirement for Mg2+ and enhancing its extent of activation by guanine nucleotides. Proc Natl Acad Sci U S A. 1982 Sep;79(17):5179–5183. doi: 10.1073/pnas.79.17.5179. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Johnson J. A., Goka T. J., Clark R. B. Phorbol ester-induced augmentation and inhibition of epinephrine-stimulated adenylate cyclase in S49 lymphoma cells. J Cyclic Nucleotide Protein Phosphor Res. 1986;11(3):199–215. [PubMed] [Google Scholar]
  16. Mahan L. C., Koachman A. M., Insel P. A. Genetic analysis of beta-adrenergic receptor internalization and down-regulation. Proc Natl Acad Sci U S A. 1985 Jan;82(1):129–133. doi: 10.1073/pnas.82.1.129. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Rashidbaigi A., Ruoho A. E., Green D. A., Clark R. B. Photoaffinity labeling of the beta-adrenergic receptor from cultured lymphoma cells with [125I]iodoazidobenzylpindolol: loss of the label with desensitization. Proc Natl Acad Sci U S A. 1983 May;80(10):2849–2853. doi: 10.1073/pnas.80.10.2849. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Roskoski R., Jr Assays of protein kinase. Methods Enzymol. 1983;99:3–6. doi: 10.1016/0076-6879(83)99034-1. [DOI] [PubMed] [Google Scholar]
  19. Salomon Y., Londos C., Rodbell M. A highly sensitive adenylate cyclase assay. Anal Biochem. 1974 Apr;58(2):541–548. doi: 10.1016/0003-2697(74)90222-x. [DOI] [PubMed] [Google Scholar]
  20. Sibley D. R., Benovic J. L., Caron M. G., Lefkowitz R. J. Regulation of transmembrane signaling by receptor phosphorylation. Cell. 1987 Mar 27;48(6):913–922. doi: 10.1016/0092-8674(87)90700-8. [DOI] [PubMed] [Google Scholar]
  21. Sternweis P. C., Gilman A. G. Reconstitution of catecholamine-sensitive adenylate cyclase. Reconstitution of the uncoupled variant of the S40 lymphoma cell. J Biol Chem. 1979 May 10;254(9):3333–3340. [PubMed] [Google Scholar]
  22. Strasser R. H., Sibley D. R., Lefkowitz R. J. A novel catecholamine-activated adenosine cyclic 3',5'-phosphate independent pathway for beta-adrenergic receptor phosphorylation in wild-type and mutant S49 lymphoma cells: mechanism of homologous desensitization of adenylate cyclase. Biochemistry. 1986 Mar 25;25(6):1371–1377. doi: 10.1021/bi00354a027. [DOI] [PubMed] [Google Scholar]
  23. Su Y. F., Cubeddu L., Perkins J. P. Regulation of adenosine 3':5'-monophosphate content of human astrocytoma cells: desensitization to catecholamines and prostaglandins. J Cyclic Nucleotide Res. 1976 Jul-Aug;2(4):257–270. [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