Skip to main content
NCBI home page
Biochemical Journal logoLink to Biochemical Journal
. 1997 Mar 15;322(Pt 3):801–808. doi: 10.1042/bj3220801

Identification and characterization of a novel A-kinase-anchoring protein (AKAP120) from rabbit gastric parietal cells.

D T Dransfield 1, J L Yeh 1, A J Bradford 1, J R Goldenring 1
PMCID: PMC1218258  PMID: 9148752

Abstract

The type-II cAMP-dependent protein kinase (A-Kinase) partitions primarily into the particulate fraction in gastric parietal cells. Localization of this kinase to particular subcellular domains is mediated through the binding of the regulatory subunit (RII) dimer to A-Kinase-anchoring proteins (AKAPs). Using a [32P]RII overlay assay, we have screened a rabbit gastric parietal cell cDNA library and have isolated a single RII-binding protein clone. Sequence analysis revealed an open reading frame coding for 1022 amino acids (AKAP120). Recombinant fragments of the full-length clone were prepared and the RII-binding region mapped to an area between amino acids 489 and 549. This area contained a putative alpha-helical RII-binding region between amino acids 503 and 516. Incubation of [32P]RII with a synthetic peptide of AKAP120-(489-522) completely inhibited the binding of [32P]RII to the recombinant AKAP120 fragments that demonstrated RII binding. In vitro RII-binding affinity studies indicated a high-affinity interaction between AKAP120 and RII with a Kapp between 50 and 120 nM for the three recombinant fragments that bound [32P]RII. RNase-protection analysis revealed that AKAP120 is a widely distributed protein, with the highest levels of mRNA observed in gastric fundus. The presence of this novel high-affinity AKAP in gastric parietal cells suggests that it may regulate RII subcellular sequestration in this cell type.

Full Text

The Full Text of this article is available as a PDF (711.3 KB).

Selected References

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

  1. Carr D. W., Hausken Z. E., Fraser I. D., Stofko-Hahn R. E., Scott J. D. Association of the type II cAMP-dependent protein kinase with a human thyroid RII-anchoring protein. Cloning and characterization of the RII-binding domain. J Biol Chem. 1992 Jul 5;267(19):13376–13382. [PubMed] [Google Scholar]
  2. Carr D. W., Stofko-Hahn R. E., Fraser I. D., Bishop S. M., Acott T. S., Brennan R. G., Scott J. D. Interaction of the regulatory subunit (RII) of cAMP-dependent protein kinase with RII-anchoring proteins occurs through an amphipathic helix binding motif. J Biol Chem. 1991 Aug 5;266(22):14188–14192. [PubMed] [Google Scholar]
  3. Carr D. W., Stofko-Hahn R. E., Fraser I. D., Cone R. D., Scott J. D. Localization of the cAMP-dependent protein kinase to the postsynaptic densities by A-kinase anchoring proteins. Characterization of AKAP 79. J Biol Chem. 1992 Aug 25;267(24):16816–16823. [PubMed] [Google Scholar]
  4. Chew C. S., Brown M. R. Histamine increases phosphorylation of 27- and 40-kDa parietal cell proteins. Am J Physiol. 1987 Dec;253(6 Pt 1):G823–G829. doi: 10.1152/ajpgi.1987.253.6.G823. [DOI] [PubMed] [Google Scholar]
  5. Chew C. S. Parietal cell protein kinases. Selective activation of type I cAMP-dependent protein kinase by histamine. J Biol Chem. 1985 Jun 25;260(12):7540–7550. [PubMed] [Google Scholar]
  6. Coghlan V. M., Perrino B. A., Howard M., Langeberg L. K., Hicks J. B., Gallatin W. M., Scott J. D. Association of protein kinase A and protein phosphatase 2B with a common anchoring protein. Science. 1995 Jan 6;267(5194):108–111. doi: 10.1126/science.7528941. [DOI] [PubMed] [Google Scholar]
  7. Dransfield D. T., Bradford A. J., Goldenring J. R. Distribution of A-kinase anchoring proteins in parietal cells. Biochim Biophys Acta. 1995 Nov 30;1269(3):215–220. doi: 10.1016/0167-4889(95)00121-2. [DOI] [PubMed] [Google Scholar]
  8. Glantz S. B., Li Y., Rubin C. S. Characterization of distinct tethering and intracellular targeting domains in AKAP75, a protein that links cAMP-dependent protein kinase II beta to the cytoskeleton. J Biol Chem. 1993 Jun 15;268(17):12796–12804. [PubMed] [Google Scholar]
  9. Goldenring J. R., Asher V. A., Barreuther M. F., Lewis J. J., Lohmann S. M., Walter U., Modlin I. M. Dephosphorylation of cAMP-dependent protein kinase regulatory subunit in stimulated parietal cells. Am J Physiol. 1992 Apr;262(4 Pt 1):G763–G773. doi: 10.1152/ajpgi.1992.262.4.G763. [DOI] [PubMed] [Google Scholar]
  10. Hirsch A. H., Glantz S. B., Li Y., You Y., Rubin C. S. Cloning and expression of an intron-less gene for AKAP 75, an anchor protein for the regulatory subunit of cAMP-dependent protein kinase II beta. J Biol Chem. 1992 Feb 5;267(4):2131–2134. [PubMed] [Google Scholar]
  11. Klauck T. M., Faux M. C., Labudda K., Langeberg L. K., Jaken S., Scott J. D. Coordination of three signaling enzymes by AKAP79, a mammalian scaffold protein. Science. 1996 Mar 15;271(5255):1589–1592. doi: 10.1126/science.271.5255.1589. [DOI] [PubMed] [Google Scholar]
  12. Lin R. Y., Moss S. B., Rubin C. S. Characterization of S-AKAP84, a novel developmentally regulated A kinase anchor protein of male germ cells. J Biol Chem. 1995 Nov 17;270(46):27804–27811. doi: 10.1074/jbc.270.46.27804. [DOI] [PubMed] [Google Scholar]
  13. McKnight S. L. Molecular zippers in gene regulation. Sci Am. 1991 Apr;264(4):54–64. doi: 10.1038/scientificamerican0491-54. [DOI] [PubMed] [Google Scholar]
  14. O'Shea E. K., Rutkowski R., Kim P. S. Evidence that the leucine zipper is a coiled coil. Science. 1989 Jan 27;243(4890):538–542. doi: 10.1126/science.2911757. [DOI] [PubMed] [Google Scholar]
  15. Oddsdottir M., Goldenring J. R., Adrian T. E., Zdon M. J., Zucker K. A., Modlin I. M. Identification and characterization of a cytosolic 30 kDa histamine stimulated phosphoprotein in parietal cell cytosol. Biochem Biophys Res Commun. 1988 Jul 29;154(2):489–496. doi: 10.1016/0006-291x(88)90166-0. [DOI] [PubMed] [Google Scholar]
  16. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Scott J. D., McCartney S. Localization of A-kinase through anchoring proteins. Mol Endocrinol. 1994 Jan;8(1):5–11. doi: 10.1210/mend.8.1.8152430. [DOI] [PubMed] [Google Scholar]
  18. Scott J. D., Stofko R. E., McDonald J. R., Comer J. D., Vitalis E. A., Mangili J. A. Type II regulatory subunit dimerization determines the subcellular localization of the cAMP-dependent protein kinase. J Biol Chem. 1990 Dec 15;265(35):21561–21566. [PubMed] [Google Scholar]
  19. Urushidani T., Hanzel D. K., Forte J. G. Characterization of an 80-kDa phosphoprotein involved in parietal cell stimulation. Am J Physiol. 1989 Jun;256(6 Pt 1):G1070–G1081. doi: 10.1152/ajpgi.1989.256.6.G1070. [DOI] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES