Skip to main content
NCBI home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1981 Jun 1;89(3):449–455. doi: 10.1083/jcb.89.3.449

Function of a calmodulin in postsynaptic densities. III. Calmodulin- binding proteins of the postsynaptic density

PMCID: PMC2111790  PMID: 6265467

Abstract

A method has been developed for binding calmodulin, radioiodinated by the lactoperoxidase method, to denaturing gels and has been used to attempt to identify the calmodulin-binding proteins of cerebral cortex postsynaptic densities (PSDs). Calmodulin primarily bound to the major 51,000 Mr protein in a saturatable manner; secondarily bound to the 60,000 Mr region, 140,000 Mr region, and 230,000 Mr protein; and bound in lesser amounts to a number of other proteins. The major 51,000 Mr calmodulin-binding protein is one of unknown identity. Binding of iodinated calmodulin to these proteins was blocked by EDTA, EGTA, chlorpromazine, and preincubation with unlabeled calmodulin. Calmodulin iodinated by the chloramine-T method, which inactivates calmodulin did not bind to the PSD but bound nonspecifically to histone. Calmodulin did not bind to proteins from a variety of sources for which calmodulin interactions have not been found. Except for three proteins, all of the proteins of synaptic membranes that bind calmodulin could be accounted for by proteins of the PSD which are a part of the synaptic membrane fraction. The major 51,000 M, protein and the corresponding iodinated calmodulin binding were greatly reduced in cerebellar PSDs and this difference between cerebral cortex and cerebellar PSDs is discussed in light of the possible function of calmodulin in synaptic excitatory responses.

Full Text

The Full Text of this article is available as a PDF (825.1 KB).

Selected References

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

  1. Adair W. S., Jurivich D., Goodenough U. W. Localization of cellular antigens in sodium dodecyl sulfate-polyacrylamide gels. J Cell Biol. 1978 Oct;79(1):281–285. doi: 10.1083/jcb.79.1.281. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Amphlett G. W., Vanaman T. C., Perry S. V. Effect of the troponin C-like protein from bovine brain (brain modulator protein) on the Mg2+-stimulated ATPase of skeletal muscle actinomyosin. FEBS Lett. 1976 Dec 15;72(1):163–168. doi: 10.1016/0014-5793(76)80836-8. [DOI] [PubMed] [Google Scholar]
  3. Argov Z., Yaari Y. The action of chlorpromazine at an isolated cholinergic synapse. Brain Res. 1979 Mar 23;164:227–236. doi: 10.1016/0006-8993(79)90018-0. [DOI] [PubMed] [Google Scholar]
  4. Ariano M. A., Appleman M. M. Biochemical characterization of postsynaptically localized cyclic nucleotide phosphodiesterase. Brain Res. 1979 Nov 16;177(2):301–309. doi: 10.1016/0006-8993(79)90781-9. [DOI] [PubMed] [Google Scholar]
  5. Baudry M., Lynch G. Regulation of glutamate receptors by cations. Nature. 1979 Dec 13;282(5740):748–750. doi: 10.1038/282748a0. [DOI] [PubMed] [Google Scholar]
  6. Bigelis R., Burridge K. The immunological detection of yeast nonsense termination fragments on sodium dodecylsulfate-polyacrylamide gels. Biochem Biophys Res Commun. 1978 May 15;82(1):322–327. doi: 10.1016/0006-291x(78)90612-5. [DOI] [PubMed] [Google Scholar]
  7. Brostrom C. O., Huang Y. C., Breckenridge B. M., Wolff D. J. Identification of a calcium-binding protein as a calcium-dependent regulator of brain adenylate cyclase. Proc Natl Acad Sci U S A. 1975 Jan;72(1):64–68. doi: 10.1073/pnas.72.1.64. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Burridge K. Changes in cellular glycoproteins after transformation: identification of specific glycoproteins and antigens in sodium dodecyl sulfate gels. Proc Natl Acad Sci U S A. 1976 Dec;73(12):4457–4461. doi: 10.1073/pnas.73.12.4457. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Carlin R. K., Grab D. J., Cohen R. S., Siekevitz P. Isolation and characterization of postsynaptic densities from various brain regions: enrichment of different types of postsynaptic densities. J Cell Biol. 1980 Sep;86(3):831–845. doi: 10.1083/jcb.86.3.831. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Carlin R. K., Grab D. J., Siekevitz P. The binding of radio-iodinated calmodulin to proteins on denaturing gels. Ann N Y Acad Sci. 1980;356:73–74. doi: 10.1111/j.1749-6632.1980.tb29600.x. [DOI] [PubMed] [Google Scholar]
  11. Cheung W. Y., Bradham L. S., Lynch T. J., Lin Y. M., Tallant E. A. Protein activator of cyclic 3':5'-nucleotide phosphodiesterase of bovine or rat brain also activates its adenylate cyclase. Biochem Biophys Res Commun. 1975 Oct 6;66(3):1055–1062. doi: 10.1016/0006-291x(75)90747-0. [DOI] [PubMed] [Google Scholar]
  12. Childers S. R., Siegel F. L. Isolation and purification of calcium-binding protein from electroplax of Electrophorus electricus. Biochim Biophys Acta. 1975 Sep 9;405(1):99–108. doi: 10.1016/0005-2795(75)90319-0. [DOI] [PubMed] [Google Scholar]
  13. Cohen R. S., Blomberg F., Berzins K., Siekevitz P. The structure of postsynaptic densities isolated from dog cerebral cortex. I. Overall morphology and protein composition. J Cell Biol. 1977 Jul;74(1):181–203. doi: 10.1083/jcb.74.1.181. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Dunwiddie T. V., Lynch G. The relationship between extracellular calcium concentrations and the induction of hippocampal long-term potentiation. Brain Res. 1979 Jun 15;169(1):103–110. doi: 10.1016/0006-8993(79)90377-9. [DOI] [PubMed] [Google Scholar]
  15. Feit H., Kelly P., Cotman C. W. Identification of a protein related to tubulin in the postsynaptic density. Proc Natl Acad Sci U S A. 1977 Mar;74(3):1047–1051. doi: 10.1073/pnas.74.3.1047. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Glenney J. R., Jr, Weber K. Calmodulin-binding proteins of the microfilaments present in isolated brush borders and microvilli of intestinal epithelial cells. J Biol Chem. 1980 Nov 25;255(22):10551–10554. [PubMed] [Google Scholar]
  17. Grab D. J., Berzins K., Cohen R. S., Siekevitz P. Presence of calmodulin in postsynaptic densities isolated from canine cerebral cortex. J Biol Chem. 1979 Sep 10;254(17):8690–8696. [PubMed] [Google Scholar]
  18. Grab D. J., Carlin R. K., Siekevitz P. Function of a calmodulin in postsynaptic densities. II. Presence of a calmodulin-activatable protein kinase activity. J Cell Biol. 1981 Jun;89(3):440–448. doi: 10.1083/jcb.89.3.440. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Grab D. J., Carlin R. K., Siekevitz P. Function of calmodulin in postsynaptic densities. I. Presence of a calmodulin-activatable cyclic nucleotide phosphodiesterase activity. J Cell Biol. 1981 Jun;89(3):433–439. doi: 10.1083/jcb.89.3.433. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Grab D. J., Carlin R. K., Siekevitz P. The presence and functions of calmodulin in the postsynaptic density. Ann N Y Acad Sci. 1980;356:55–72. doi: 10.1111/j.1749-6632.1980.tb29599.x. [DOI] [PubMed] [Google Scholar]
  21. Graf E., Filoteo A. G., Penniston J. T. Preparation of 125I-calmodulin with retention of full biological activity: its binding to human erythrocyte ghosts. Arch Biochem Biophys. 1980 Sep;203(2):719–726. doi: 10.1016/0003-9861(80)90231-3. [DOI] [PubMed] [Google Scholar]
  22. HUNTER W. M., GREENWOOD F. C. Preparation of iodine-131 labelled human growth hormone of high specific activity. Nature. 1962 May 5;194:495–496. doi: 10.1038/194495a0. [DOI] [PubMed] [Google Scholar]
  23. Hotson J. R., Prince D. A. A calcium-activated hyperpolarization follows repetitive firing in hippocampal neurons. J Neurophysiol. 1980 Feb;43(2):409–419. doi: 10.1152/jn.1980.43.2.409. [DOI] [PubMed] [Google Scholar]
  24. Kelly P. T., Cotman C. W. Synaptic proteins. Characterization of tubulin and actin and identification of a distinct postsynaptic density polypeptide. J Cell Biol. 1978 Oct;79(1):173–183. doi: 10.1083/jcb.79.1.173. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Klee C. B., Crouch T. H., Krinks M. H. Subunit structure and catalytic properties of bovine brain Ca2+-dependent cyclic nucleotide phosphodiesterase. Biochemistry. 1979 Feb 20;18(4):722–729. doi: 10.1021/bi00571a026. [DOI] [PubMed] [Google Scholar]
  26. Krnjević K., Lamour Y., MacDonald J. F., Nistri A. Depression of monosynaptic excitatory postsynaptic potentials by Mn2+ and Co2+ in cat spinal cord. Neuroscience. 1979;4(9):1331–1339. doi: 10.1016/0306-4522(79)90160-x. [DOI] [PubMed] [Google Scholar]
  27. Kuo C. H., Ichida S., Matsuda T., Kakiuchi S., Yoshida H. Regulation of ATP-dependent Ca-uptake of synaptic plasma membranes by Ca-dependent modulator protein. Life Sci. 1979 Jul 16;25(3):235–239. doi: 10.1016/0024-3205(79)90290-x. [DOI] [PubMed] [Google Scholar]
  28. 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]
  29. LaPorte D. C., Storm D. R. Detection of calcium-dependent regulatory protein binding components using 125I-labeled calcium-dependent regulatory protein. J Biol Chem. 1978 May 25;253(10):3374–3377. [PubMed] [Google Scholar]
  30. Levin R. M., Weiss B. Specificity of the binding of trifluoperazine to the calcium-dependent activator of phosphodiesterase and to a series of other calcium-binding proteins. Biochim Biophys Acta. 1978 May 3;540(2):197–204. doi: 10.1016/0304-4165(78)90132-0. [DOI] [PubMed] [Google Scholar]
  31. Lin C. T., Dedman J. R., Brinkley B. R., Means A. R. Localization of calmodulin in rat cerebellum by immunoelectron microscopy. J Cell Biol. 1980 May;85(2):473–480. doi: 10.1083/jcb.85.2.473. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Manthey A. A. The effect of calcium on the desensitization of membrane receptors at the neuromuscular junction. J Gen Physiol. 1966 May;49(5):963–976. doi: 10.1085/jgp.49.5.963. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Marcum J. M., Dedman J. R., Brinkley B. R., Means A. R. Control of microtubule assembly-disassembly by calcium-dependent regulator protein. Proc Natl Acad Sci U S A. 1978 Aug;75(8):3771–3775. doi: 10.1073/pnas.75.8.3771. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Matus A. I., Walters B. B., Mughal S. Immunohistochemical demonstration of tubulin associated with microtubules and synaptic junctions in mammalian brain. J Neurocytol. 1975 Dec;4(6):733–744. doi: 10.1007/BF01181633. [DOI] [PubMed] [Google Scholar]
  35. Nastuk W. L., Parsons R. L. Factors in the inactivation of postjunctional membrane receptors of frog skeletal muscle. J Gen Physiol. 1970 Aug;56(2):218–249. doi: 10.1085/jgp.56.2.218. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Richman P. G., Klee C. B. Interaction of 125I-labeled Ca2+-dependent regulator protein with cyclic nucleotide phosphodiesterase and its inhibitory protein. J Biol Chem. 1978 Sep 25;253(18):6323–6326. [PubMed] [Google Scholar]
  37. Rostas J. A., Kelly P. T., Pesin R. H., Cotman C. W. Protein and glycoprotein composition of synaptic junctions prepared from discrete synaptic regions and different species. Brain Res. 1979 May 18;168(1):151–167. doi: 10.1016/0006-8993(79)90133-1. [DOI] [PubMed] [Google Scholar]
  38. Sobue K., Ichida S., Yoshida H., Yamazaki R., Kakiuchi S. Occurrence of a Ca2+- and modulator protein-activatable ATPase in the synaptic plasma membranes of brain. FEBS Lett. 1979 Mar 1;99(1):199–202. doi: 10.1016/0014-5793(79)80278-1. [DOI] [PubMed] [Google Scholar]
  39. Wallace R. W., Tallant E. A., Cheung W. Y. High levels of a heat-labile calmodulin-binding protein (CaM-BP80) in bovine neostriatum. Biochemistry. 1980 Apr 29;19(9):1831–1837. doi: 10.1021/bi00550a016. [DOI] [PubMed] [Google Scholar]
  40. Walsh M., Stevens F. C. Chemical modification studies on the Ca2+-dependent protein modulator of cyclic nucleotide phosphodiesterase. Biochemistry. 1977 Jun 14;16(12):2742–2749. doi: 10.1021/bi00631a024. [DOI] [PubMed] [Google Scholar]
  41. Watterson D. M., Harrelson W. G., Jr, Keller P. M., Sharief F., Vanaman T. C. Structural similarities between the Ca2+-dependent regulatory proteins of 3':5'-cyclic nucleotide phosphodiesterase and actomyosin ATPase. J Biol Chem. 1976 Aug 10;251(15):4501–4513. [PubMed] [Google Scholar]
  42. Westrum L. E., Gray E. G. Microtubules associated with postsynaptic 'thickenings'. J Neurocytol. 1977 Oct;6(5):505–518. doi: 10.1007/BF01205216. [DOI] [PubMed] [Google Scholar]
  43. Wong P. Y., Cheung W. Y. Calmodulin stimulates human platelet phospholipase A2. Biochem Biophys Res Commun. 1979 Sep 27;90(2):473–480. doi: 10.1016/0006-291x(79)91259-2. [DOI] [PubMed] [Google Scholar]
  44. Wood J. G., Wallace R. W., Whitaker J. N., Cheung W. Y. Immunocytochemical localization of calmodulin and a heat-labile calmodulin-binding protein (CaM-BP80) in basal ganglia of mouse brain. J Cell Biol. 1980 Jan;84(1):66–76. doi: 10.1083/jcb.84.1.66. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

RESOURCES