Skip to main content
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1987 Jun 1;104(6):1579–1586. doi: 10.1083/jcb.104.6.1579

Visualization of neural cell adhesion molecule by electron microscopy

PMCID: PMC2114501  PMID: 2438287

Abstract

The 130- and 160-kD polypeptide forms of the neural cell adhesion molecule (NCAM) were analyzed by electron microscopy after low angle rotary shadowing and freeze replication. Individual NCAM molecules appeared as uniformly thick rods, with a distinct bend or hinge region near their middle. Aggregates were also present, containing two to six rods in a pinwheel-like configuration without measurable overlap between rods. The 130- and 160-kD NCAM forms had lengths of 38 and 51 nm, respectively, with a difference in arm length distal to the bend, but not toward the center of the pinwheel. Although enzymatic removal of the polysialic acid moiety on NCAM did not alter the appearance of individual molecules, it did increase the average number of arms per aggregate. Monoclonal antibodies that recognize defined regions of the NCAM polypeptide were used to provide landmarks on the observed molecular figures. Two antibodies specific for cytoplasmic epitopes near the COOH terminus were clustered at the distal tip of aggregated arms. Two other antibodies that react with epitopes near the NH2 terminus and the middle of the molecule bound to sites more centrally located on the pinwheel structure. Together, these results suggest that the observed aggregates represent an association of molecules near their NH2-terminal homophilic binding site, and have led to several predictions about the nature of an NCAM-mediated cell-cell bond.

Full Text

The Full Text of this article is available as a PDF (3.1 MB).

Selected References

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

  1. Cole G. J., Loewy A., Glaser L. Neuronal cell-cell adhesion depends on interactions of N-CAM with heparin-like molecules. Nature. 1986 Apr 3;320(6061):445–447. doi: 10.1038/320445a0. [DOI] [PubMed] [Google Scholar]
  2. Crossin K. L., Edelman G. M., Cunningham B. A. Mapping of three carbohydrate attachment sites in embryonic and adult forms of the neural cell adhesion molecule. J Cell Biol. 1984 Nov;99(5):1848–1855. doi: 10.1083/jcb.99.5.1848. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Cunningham B. A., Hoffman S., Rutishauser U., Hemperly J. J., Edelman G. M. Molecular topography of the neural cell adhesion molecule N-CAM: surface orientation and location of sialic acid-rich and binding regions. Proc Natl Acad Sci U S A. 1983 May;80(10):3116–3120. doi: 10.1073/pnas.80.10.3116. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Edelman G. M., Hoffman S., Chuong C. M., Thiery J. P., Brackenbury R., Gallin W. J., Grumet M., Greenberg M. E., Hemperly J. J., Cohen C. Structure and modulation of neural cell adhesion molecules in early and late embryogenesis. Cold Spring Harb Symp Quant Biol. 1983;48(Pt 2):515–526. doi: 10.1101/sqb.1983.048.01.056. [DOI] [PubMed] [Google Scholar]
  5. Edelman G. M. Modulation of cell adhesion during induction, histogenesis, and perinatal development of the nervous system. Annu Rev Neurosci. 1984;7:339–377. doi: 10.1146/annurev.ne.07.030184.002011. [DOI] [PubMed] [Google Scholar]
  6. Elder J. H., Alexander S. endo-beta-N-acetylglucosaminidase F: endoglycosidase from Flavobacterium meningosepticum that cleaves both high-mannose and complex glycoproteins. Proc Natl Acad Sci U S A. 1982 Aug;79(15):4540–4544. doi: 10.1073/pnas.79.15.4540. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Frelinger A. L., 3rd, Rutishauser U. Topography of N-CAM structural and functional determinants. II. Placement of monoclonal antibody epitopes. J Cell Biol. 1986 Nov;103(5):1729–1737. doi: 10.1083/jcb.103.5.1729. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Gennarini G., Hirn M., Deagostini-Bazin H., Goridis C. Studies on the transmembrane disposition of the neural cell adhesion molecule N-CAM. The use of liposome-inserted radioiodinated N-CAM to study its transbilayer orientation. Eur J Biochem. 1984 Jul 2;142(1):65–73. doi: 10.1111/j.1432-1033.1984.tb08251.x. [DOI] [PubMed] [Google Scholar]
  9. Glenney J. R., Jr, Glenney P., Weber K. Mapping the fodrin molecule with monoclonal antibodies. A general approach for rod-like multidomain proteins. J Mol Biol. 1983 Jun 25;167(2):275–293. doi: 10.1016/s0022-2836(83)80336-2. [DOI] [PubMed] [Google Scholar]
  10. Hall A. K., Rutishauser U. Phylogeny of a neural cell adhesion molecule. Dev Biol. 1985 Jul;110(1):39–46. doi: 10.1016/0012-1606(85)90061-2. [DOI] [PubMed] [Google Scholar]
  11. Hemperly J. J., Murray B. A., Edelman G. M., Cunningham B. A. Sequence of a cDNA clone encoding the polysialic acid-rich and cytoplasmic domains of the neural cell adhesion molecule N-CAM. Proc Natl Acad Sci U S A. 1986 May;83(9):3037–3041. doi: 10.1073/pnas.83.9.3037. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Heuser J. E. Procedure for freeze-drying molecules adsorbed to mica flakes. J Mol Biol. 1983 Sep 5;169(1):155–195. doi: 10.1016/s0022-2836(83)80179-x. [DOI] [PubMed] [Google Scholar]
  13. Hoffman S., Sorkin B. C., White P. C., Brackenbury R., Mailhammer R., Rutishauser U., Cunningham B. A., Edelman G. M. Chemical characterization of a neural cell adhesion molecule purified from embryonic brain membranes. J Biol Chem. 1982 Jul 10;257(13):7720–7729. [PubMed] [Google Scholar]
  14. Laurie G. W., Bing J. T., Kleinman H. K., Hassell J. R., Aumailley M., Martin G. R., Feldmann R. J. Localization of binding sites for laminin, heparan sulfate proteoglycan and fibronectin on basement membrane (type IV) collagen. J Mol Biol. 1986 May 5;189(1):205–216. doi: 10.1016/0022-2836(86)90391-8. [DOI] [PubMed] [Google Scholar]
  15. Noble M., Albrechtsen M., Møller C., Lyles J., Bock E., Goridis C., Watanabe M., Rutishauser U. Glial cells express N-CAM/D2-CAM-like polypeptides in vitro. Nature. 1985 Aug 22;316(6030):725–728. doi: 10.1038/316725a0. [DOI] [PubMed] [Google Scholar]
  16. Oakley B. R., Kirsch D. R., Morris N. R. A simplified ultrasensitive silver stain for detecting proteins in polyacrylamide gels. Anal Biochem. 1980 Jul 1;105(2):361–363. doi: 10.1016/0003-2697(80)90470-4. [DOI] [PubMed] [Google Scholar]
  17. Rougon G., Marshak D. R. Structural and immunological characterization of the amino-terminal domain of mammalian neural cell adhesion molecules. J Biol Chem. 1986 Mar 5;261(7):3396–3401. [PubMed] [Google Scholar]
  18. Rutishauser U. Developmental biology of a neural cell adhesion molecule. Nature. 1984 Aug 16;310(5978):549–554. doi: 10.1038/310549a0. [DOI] [PubMed] [Google Scholar]
  19. Rutishauser U., Hoffman S., Edelman G. M. Binding properties of a cell adhesion molecule from neural tissue. Proc Natl Acad Sci U S A. 1982 Jan;79(2):685–689. doi: 10.1073/pnas.79.2.685. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Rutishauser U., Watanabe M., Silver J., Troy F. A., Vimr E. R. Specific alteration of NCAM-mediated cell adhesion by an endoneuraminidase. J Cell Biol. 1985 Nov;101(5 Pt 1):1842–1849. doi: 10.1083/jcb.101.5.1842. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Slayter H. S. High-resolution metal replication of macromolecules. Ultramicroscopy. 1976 Sep-Oct;1(4):341–357. doi: 10.1016/0304-3991(76)90050-4. [DOI] [PubMed] [Google Scholar]
  22. Thanos S., Bonhoeffer F., Rutishauser U. Fiber-fiber interaction and tectal cues influence the development of the chicken retinotectal projection. Proc Natl Acad Sci U S A. 1984 Mar;81(6):1906–1910. doi: 10.1073/pnas.81.6.1906. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Tyler J. M., Branton D. Rotary shadowing of extended molecules dried from glycerol. J Ultrastruct Res. 1980 May;71(2):95–102. doi: 10.1016/s0022-5320(80)90098-2. [DOI] [PubMed] [Google Scholar]
  24. Ungewickell E., Branton D. Assembly units of clathrin coats. Nature. 1981 Jan 29;289(5796):420–422. doi: 10.1038/289420a0. [DOI] [PubMed] [Google Scholar]
  25. Vimr E. R., McCoy R. D., Vollger H. F., Wilkison N. C., Troy F. A. Use of prokaryotic-derived probes to identify poly(sialic acid) in neonatal neuronal membranes. Proc Natl Acad Sci U S A. 1984 Apr;81(7):1971–1975. doi: 10.1073/pnas.81.7.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Watanabe M., Frelinger A. L., 3rd, Rutishauser U. Topography of N-CAM structural and functional determinants. I. Classification of monoclonal antibody epitopes. J Cell Biol. 1986 Nov;103(5):1721–1727. doi: 10.1083/jcb.103.5.1721. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES