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. 1987 Nov 11;15(21):8621–8641. doi: 10.1093/nar/15.21.8621

Analysis of cDNA clones that code for the transmembrane forms of the mouse neural cell adhesion molecule (NCAM) and are generated by alternative RNA splicing.

M J Santoni 1, D Barthels 1, J A Barbas 1, M R Hirsch 1, M Steinmetz 1, C Goridis 1, W Wille 1
PMCID: PMC306395  PMID: 3684567

Abstract

The neural cell adhesion molecule (NCAM) exists in at least three different isoforms. In the mouse, NCAM proteins with apparent Mr's of 180,000, 140,000 and 120,000 have been distinguished. These are encoded by 4 to 5 different transcripts. Here we report the full amino acid sequence of an isoform which most likely represents NCAM-140. The N-terminal extracellular portion of the 829-residue polypeptide appears to be identical to all three NCAM proteins. The Mr of 91,276 is considerably smaller than the estimate based on SDS-gel electrophoresis. The 147 C-terminal residues are distinct from NCAM-120 and contain the putative transmembrane and cytoplasmic domains. The transcript encoding NCAM-140 contains almost 3.2 kb non-coding sequence with a canonical polyadenylation signal. While the 5' sequences of NCAM-140 hybridize with all NCAM mRNAs, the 3' probes recognize only the two larger transcripts of 7.4 and 6.7 kb. From S1 nuclease protection analyses and hybridization studies of several NCAM cDNA clones with genomic NCAM sequences one can conclude that the different NCAM transcripts are generated by alternative splicing. In addition to the two alternative splice sites in the sequence encoding the extracellular domains, a third one can be predicted approximately 320 nt downstream of the start of the NCAM-140-specific sequence portion. This finding is in agreement with the existence of an extra exon in the chicken NCAM-180. Comparison between mouse and chicken NCAM amino acid sequences revealed the highest homology in the second and fifth Ig-like domains and in the cytoplasmic parts suggesting that these regions serve highly conserved functions.

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Selected References

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  1. Barthels D., Santoni M. J., Wille W., Ruppert C., Chaix J. C., Hirsch M. R., Fontecilla-Camps J. C., Goridis C. Isolation and nucleotide sequence of mouse NCAM cDNA that codes for a Mr 79,000 polypeptide without a membrane-spanning region. EMBO J. 1987 Apr;6(4):907–914. doi: 10.1002/j.1460-2075.1987.tb04837.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Benda P., Lightbody J., Sato G., Levine L., Sweet W. Differentiated rat glial cell strain in tissue culture. Science. 1968 Jul 26;161(3839):370–371. doi: 10.1126/science.161.3839.370. [DOI] [PubMed] [Google Scholar]
  3. Cathala G., Savouret J. F., Mendez B., West B. L., Karin M., Martial J. A., Baxter J. D. A method for isolation of intact, translationally active ribonucleic acid. DNA. 1983;2(4):329–335. doi: 10.1089/dna.1983.2.329. [DOI] [PubMed] [Google Scholar]
  4. Chuong C. M., Edelman G. M. Expression of cell-adhesion molecules in embryonic induction. I. Morphogenesis of nestling feathers. J Cell Biol. 1985 Sep;101(3):1009–1026. doi: 10.1083/jcb.101.3.1009. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Corden J., Wasylyk B., Buchwalder A., Sassone-Corsi P., Kedinger C., Chambon P. Promoter sequences of eukaryotic protein-coding genes. Science. 1980 Sep 19;209(4463):1406–1414. doi: 10.1126/science.6251548. [DOI] [PubMed] [Google Scholar]
  6. Covault J., Merlie J. P., Goridis C., Sanes J. R. Molecular forms of N-CAM and its RNA in developing and denervated skeletal muscle. J Cell Biol. 1986 Mar;102(3):731–739. doi: 10.1083/jcb.102.3.731. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Cunningham B. A., Hemperly J. J., Murray B. A., Prediger E. A., Brackenbury R., Edelman G. M. Neural cell adhesion molecule: structure, immunoglobulin-like domains, cell surface modulation, and alternative RNA splicing. Science. 1987 May 15;236(4803):799–806. doi: 10.1126/science.3576199. [DOI] [PubMed] [Google Scholar]
  8. 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]
  9. Devereux J., Haeberli P., Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. doi: 10.1093/nar/12.1part1.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Dierks P., van Ooyen A., Cochran M. D., Dobkin C., Reiser J., Weissmann C. Three regions upstream from the cap site are required for efficient and accurate transcription of the rabbit beta-globin gene in mouse 3T6 cells. Cell. 1983 Mar;32(3):695–706. doi: 10.1016/0092-8674(83)90055-7. [DOI] [PubMed] [Google Scholar]
  11. Edelman G. M. Cell adhesion and the molecular processes of morphogenesis. Annu Rev Biochem. 1985;54:135–169. doi: 10.1146/annurev.bi.54.070185.001031. [DOI] [PubMed] [Google Scholar]
  12. Finne J., Finne U., Deagostini-Bazin H., Goridis C. Occurrence of alpha 2-8 linked polysialosyl units in a neural cell adhesion molecule. Biochem Biophys Res Commun. 1983 Apr 29;112(2):482–487. doi: 10.1016/0006-291x(83)91490-0. [DOI] [PubMed] [Google Scholar]
  13. Fraser S. E., Murray B. A., Chuong C. M., Edelman G. M. Alteration of the retinotectal map in Xenopus by antibodies to neural cell adhesion molecules. Proc Natl Acad Sci U S A. 1984 Jul;81(13):4222–4226. doi: 10.1073/pnas.81.13.4222. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. 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]
  15. 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]
  16. Gennarini G., Hirsch M. R., He H. T., Hirn M., Finne J., Goridis C. Differential expression of mouse neural cell-adhesion molecule (N-CAM) mRNA species during brain development and in neural cell lines. J Neurosci. 1986 Jul;6(7):1983–1990. doi: 10.1523/JNEUROSCI.06-07-01983.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Gennarini G., Rougon G., Deagostini-Bazin H., Hirn M., Goridis C. Studies on the transmembrane disposition of the neural cell adhesion molecule N-CAM. A monoclonal antibody recognizing a cytoplasmic domain and evidence for the presence of phosphoserine residues. Eur J Biochem. 1984 Jul 2;142(1):57–64. doi: 10.1111/j.1432-1033.1984.tb08250.x. [DOI] [PubMed] [Google Scholar]
  18. Goridis C., Deagostini-Bazin H., Hirn M., Hirsch M. R., Rougon G., Sadoul R., Langley O. K., Gombos G., Finne J. Neural surface antigens during nervous system development. Cold Spring Harb Symp Quant Biol. 1983;48(Pt 2):527–537. doi: 10.1101/sqb.1983.048.01.057. [DOI] [PubMed] [Google Scholar]
  19. Goridis C., Hirn M., Santoni M. J., Gennarini G., Deagostini-Bazin H., Jordan B. R., Kiefer M., Steinmetz M. Isolation of mouse N-CAM-related cDNA: detection and cloning using monoclonal antibodies. EMBO J. 1985 Mar;4(3):631–635. doi: 10.1002/j.1460-2075.1985.tb03676.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. He H. T., Barbet J., Chaix J. C., Goridis C. Phosphatidylinositol is involved in the membrane attachment of NCAM-120, the smallest component of the neural cell adhesion molecule. EMBO J. 1986 Oct;5(10):2489–2494. doi: 10.1002/j.1460-2075.1986.tb04526.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Hemperly J. J., Edelman G. M., Cunningham B. A. cDNA clones of the neural cell adhesion molecule (N-CAM) lacking a membrane-spanning region consistent with evidence for membrane attachment via a phosphatidylinositol intermediate. Proc Natl Acad Sci U S A. 1986 Dec;83(24):9822–9826. doi: 10.1073/pnas.83.24.9822. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. 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]
  23. Hoffman S., Chuong C. M., Edelman G. M. Evolutionary conservation of key structures and binding functions of neural cell adhesion molecules. Proc Natl Acad Sci U S A. 1984 Nov;81(21):6881–6885. doi: 10.1073/pnas.81.21.6881. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Hoffman S., Edelman G. M. Kinetics of homophilic binding by embryonic and adult forms of the neural cell adhesion molecule. Proc Natl Acad Sci U S A. 1983 Sep;80(18):5762–5766. doi: 10.1073/pnas.80.18.5762. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. 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]
  26. Keilhauer G., Faissner A., Schachner M. Differential inhibition of neurone-neurone, neurone-astrocyte and astrocyte-astrocyte adhesion by L1, L2 and N-CAM antibodies. Nature. 1985 Aug 22;316(6030):728–730. doi: 10.1038/316728a0. [DOI] [PubMed] [Google Scholar]
  27. Kyte J., Doolittle R. F. A simple method for displaying the hydropathic character of a protein. J Mol Biol. 1982 May 5;157(1):105–132. doi: 10.1016/0022-2836(82)90515-0. [DOI] [PubMed] [Google Scholar]
  28. Lawler J., Hynes R. O. The structure of human thrombospondin, an adhesive glycoprotein with multiple calcium-binding sites and homologies with several different proteins. J Cell Biol. 1986 Nov;103(5):1635–1648. doi: 10.1083/jcb.103.5.1635. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Lyles J. M., Linnemann D., Bock E. Biosynthesis of the D2-cell adhesion molecule: post-translational modifications, intracellular transport, and developmental changes. J Cell Biol. 1984 Dec;99(6):2082–2091. doi: 10.1083/jcb.99.6.2082. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Mostov K. E., Friedlander M., Blobel G. The receptor for transepithelial transport of IgA and IgM contains multiple immunoglobulin-like domains. Nature. 1984 Mar 1;308(5954):37–43. doi: 10.1038/308037a0. [DOI] [PubMed] [Google Scholar]
  31. Murray B. A., Hemperly J. J., Gallin W. J., MacGregor J. S., Edelman G. M., Cunningham B. A. Isolation of cDNA clones for the chicken neural cell adhesion molecule (N-CAM). Proc Natl Acad Sci U S A. 1984 Sep;81(17):5584–5588. doi: 10.1073/pnas.81.17.5584. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Murray B. A., Hemperly J. J., Prediger E. A., Edelman G. M., Cunningham B. A. Alternatively spliced mRNAs code for different polypeptide chains of the chicken neural cell adhesion molecule (N-CAM). J Cell Biol. 1986 Jan;102(1):189–193. doi: 10.1083/jcb.102.1.189. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Murray B. A., Owens G. C., Prediger E. A., Crossin K. L., Cunningham B. A., Edelman G. M. Cell surface modulation of the neural cell adhesion molecule resulting from alternative mRNA splicing in a tissue-specific developmental sequence. J Cell Biol. 1986 Oct;103(4):1431–1439. doi: 10.1083/jcb.103.4.1431. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Nguyen C., Mattei M. G., Mattei J. F., Santoni M. J., Goridis C., Jordan B. R. Localization of the human NCAM gene to band q23 of chromosome 11: the third gene coding for a cell interaction molecule mapped to the distal portion of the long arm of chromosome 11. J Cell Biol. 1986 Mar;102(3):711–715. doi: 10.1083/jcb.102.3.711. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Owens G. C., Edelman G. M., Cunningham B. A. Organization of the neural cell adhesion molecule (N-CAM) gene: alternative exon usage as the basis for different membrane-associated domains. Proc Natl Acad Sci U S A. 1987 Jan;84(1):294–298. doi: 10.1073/pnas.84.1.294. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. 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]
  37. Ruppert C., Goldowitz D., Wille W. Proto-oncogene c-myc is expressed in cerebellar neurons at different developmental stages. EMBO J. 1986 Aug;5(8):1897–1901. doi: 10.1002/j.1460-2075.1986.tb04442.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Ruppert C., Wille W. Proto-oncogene c-fos is highly induced by disruption of neonatal but not of mature brain tissue. Brain Res. 1987 Apr;388(1):51–56. doi: 10.1016/0169-328x(87)90020-9. [DOI] [PubMed] [Google Scholar]
  39. Rutishauser U., Grumet M., Edelman G. M. Neural cell adhesion molecule mediates initial interactions between spinal cord neurons and muscle cells in culture. J Cell Biol. 1983 Jul;97(1):145–152. doi: 10.1083/jcb.97.1.145. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Sadoul R., Hirn M., Deagostini-Bazin H., Rougon G., Goridis C. Adult and embryonic mouse neural cell adhesion molecules have different binding properties. 1983 Jul 28-Aug 3Nature. 304(5924):347–349. doi: 10.1038/304347a0. [DOI] [PubMed] [Google Scholar]
  41. 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]
  42. Silver J., Rutishauser U. Guidance of optic axons in vivo by a preformed adhesive pathway on neuroepithelial endfeet. Dev Biol. 1984 Dec;106(2):485–499. doi: 10.1016/0012-1606(84)90248-3. [DOI] [PubMed] [Google Scholar]
  43. Sorkin B. C., Hoffman S., Edelman G. M., Cunningham B. A. Sulfation and phosphorylation of the neural cell adhesion molecule, N-CAM. Science. 1984 Sep 28;225(4669):1476–1478. doi: 10.1126/science.6474186. [DOI] [PubMed] [Google Scholar]
  44. 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]
  45. 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]
  46. Wilbur W. J., Lipman D. J. Rapid similarity searches of nucleic acid and protein data banks. Proc Natl Acad Sci U S A. 1983 Feb;80(3):726–730. doi: 10.1073/pnas.80.3.726. [DOI] [PMC free article] [PubMed] [Google Scholar]

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