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. 1985 Sep 1;101(3):891–897. doi: 10.1083/jcb.101.3.891

The calcium-dependent myoblast adhesion that precedes cell fusion is mediated by glycoproteins

PMCID: PMC2113721  PMID: 4030897

Abstract

Presumptive myoblasts from explants of chick embryo pectoral muscle proliferate, differentiate, and fuse to form multinucleate myotubes. One event critical to multinucleate cell formation is the specific adhesion of myoblasts before union of their membranes. In the studies reported here five known inhibitors of myotube formation-- trifluoperazine, sodium butyrate, chloroquine, 1,10 phenanthroline, and tunicamycin--were tested for their effect on the Ca++-dependent myoblast adhesion step. The first four inhibitors of myotube formation do not perturb myoblast adhesion but rather block fusion of aggregated cells, which suggests that these agents perturb molecular events required for the union of the lipid bilayers. By contrast, tunicamycin exerts its effect by inhibiting the myoblast adhesion step, thereby blocking myotube formation. The effect of tunicamycin can be blocked by a protease inhibitor, however, which implies that the carbohydrate residues protect the glycoproteins from proteolytic degradation rather than participate directly in cell-cell adhesion. Whereas trypsin treatment of myoblasts in the absence of Ca++ destroys the cells' ability to exhibit Ca++-dependent adhesion, the presence of Ca++ during trypsin treatment inhibits the enzyme's effect, which suggests that myoblast adhesion is mediated by a glycoprotein(s) that has a conformation affected by Ca++. Finally, myoblast adhesion is inhibited by an antiserum raised against fusion-competent myoblasts. The effect of the antiserum is blocked by a fraction from the detergent extract of pectoral muscle that binds to immobilized wheat germ agglutinin, which again suggests that glycoproteins mediate Ca++-dependent myoblast adhesion.

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Selected References

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  1. Bar-Sagi D., Prives J. Trifluoperazine, a calmodulin antagonist, inhibits muscle cell fusion. J Cell Biol. 1983 Nov;97(5 Pt 1):1375–1380. doi: 10.1083/jcb.97.5.1375. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bertolotti R., Rutishauser U., Edelman G. M. A cell surface molecule involved in aggregation of embryonic liver cells. Proc Natl Acad Sci U S A. 1980 Aug;77(8):4831–4835. doi: 10.1073/pnas.77.8.4831. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bischoff R., Holtzer H. Mitosis and the processes of differentiation of myogenic cells in vitro. J Cell Biol. 1969 Apr;41(1):188–200. doi: 10.1083/jcb.41.1.188. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Brackenbury R., Rutishauser U., Edelman G. M. Distinct calcium-independent and calcium-dependent adhesion systems of chicken embryo cells. Proc Natl Acad Sci U S A. 1981 Jan;78(1):387–391. doi: 10.1073/pnas.78.1.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. COOPER W. G., KONIGSBERG I. R. Dynamics of myogenesis in vitro. Anat Rec. 1961 Jul;140:195–205. doi: 10.1002/ar.1091400305. [DOI] [PubMed] [Google Scholar]
  6. Cook J. H., Lilien J. The accessibility of certain proteins on embryonic chick neural retina cells to iodination and tryptic removal is altered by calcium. J Cell Sci. 1982 Jun;55:85–103. doi: 10.1242/jcs.55.1.85. [DOI] [PubMed] [Google Scholar]
  7. Cornell R. B., Nissley S. M., Horwitz A. F. Cholesterol availability modulates myoblast fusion. J Cell Biol. 1980 Sep;86(3):820–824. doi: 10.1083/jcb.86.3.820. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Couch C. B., Strittmatter W. J. Rat myoblast fusion requires metalloendoprotease activity. Cell. 1983 Jan;32(1):257–265. doi: 10.1016/0092-8674(83)90516-0. [DOI] [PubMed] [Google Scholar]
  9. Couch C. B., Strittmatter W. J. Specific blockers of myoblast fusion inhibit a soluble and not the membrane-associated metalloendoprotease in myoblasts. J Biol Chem. 1984 May 10;259(9):5396–5399. [PubMed] [Google Scholar]
  10. Damsky C. H., Richa J., Solter D., Knudsen K., Buck C. A. Identification and purification of a cell surface glycoprotein mediating intercellular adhesion in embryonic and adult tissue. Cell. 1983 Sep;34(2):455–466. doi: 10.1016/0092-8674(83)90379-3. [DOI] [PubMed] [Google Scholar]
  11. Den H., Malinzak D. A., Keating H. J., Rosenberg A. Influence of Concanavalin A, wheat germ agglutinin, and soybean agglutinin on the fusion of myoblasts in vitro. J Cell Biol. 1975 Dec;67(3):826–834. doi: 10.1083/jcb.67.3.826. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Ducibella T., Anderson E. Cell shape and membrane changes in the eight-cell mouse embryo: prerequisites for morphogenesis of the blastocyst. Dev Biol. 1975 Nov;47(1):45–58. doi: 10.1016/0012-1606(75)90262-6. [DOI] [PubMed] [Google Scholar]
  13. Fiszman M. Y., Montarras D., Wright W., Gros F. Expression of myogenic differentiation and myotube formation by chick embryo myoblasts in the presence of sodium butyrate. Exp Cell Res. 1980 Mar;126(1):31–37. doi: 10.1016/0014-4827(80)90467-x. [DOI] [PubMed] [Google Scholar]
  14. Gallin W. J., Edelman G. M., Cunningham B. A. Characterization of L-CAM, a major cell adhesion molecule from embryonic liver cells. Proc Natl Acad Sci U S A. 1983 Feb;80(4):1038–1042. doi: 10.1073/pnas.80.4.1038. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Gilfix B. M., Sanwal B. D. Inhibition of myoblast fusion by tunicamycin and pantomycin. Biochem Biophys Res Commun. 1980 Oct 16;96(3):1184–1191. doi: 10.1016/0006-291x(80)90077-7. [DOI] [PubMed] [Google Scholar]
  16. Gonzalez-Noriega A., Grubb J. H., Talkad V., Sly W. S. Chloroquine inhibits lysosomal enzyme pinocytosis and enhances lysosomal enzyme secretion by impairing receptor recycling. J Cell Biol. 1980 Jun;85(3):839–852. doi: 10.1083/jcb.85.3.839. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Grunwald G. B., Bromberg R. E., Crowley N. J., Lilien J. Enzymatic dissection of embryonic cell adhesive mechanisms. II. Developmental regulation of an endogenous adhesive system in the chick neural retina. Dev Biol. 1981 Sep;86(2):327–338. doi: 10.1016/0012-1606(81)90190-1. [DOI] [PubMed] [Google Scholar]
  18. Grunwald G. B., Geller R. L., Lilien J. Enzymatic dissection of embryonic cell adhesive mechanisms. J Cell Biol. 1980 Jun;85(3):766–776. doi: 10.1083/jcb.85.3.766. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Grunwald G. B., Pratt R. S., Lilien J. Enzymic dissection of embryonic cell adhesive mechanisms. III. Immunological identification of a component of the calcium-dependent adhesive system of embryonic chick neural retina cells. J Cell Sci. 1982 Jun;55:69–83. doi: 10.1242/jcs.55.1.69. [DOI] [PubMed] [Google Scholar]
  20. Holtzer H., Croop J., Dienstman S., Ishikawa H., Somlyo A. P. Effects of cytochaslasin B and colcemide on myogenic cultures. Proc Natl Acad Sci U S A. 1975 Feb;72(2):513–517. doi: 10.1073/pnas.72.2.513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Horwitz A. F., Wight A., Ludwig P., Cornell R. Interrelated lipid alterations and their influence on the proliferation and fusion of cultured myogenic cells. J Cell Biol. 1978 May;77(2):334–357. doi: 10.1083/jcb.77.2.334. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Hyafil F., Babinet C., Jacob F. Cell-cell interactions in early embryogenesis: a molecular approach to the role of calcium. Cell. 1981 Nov;26(3 Pt 1):447–454. doi: 10.1016/0092-8674(81)90214-2. [DOI] [PubMed] [Google Scholar]
  23. Kent C. Inhibition of myoblast fusion by lysosomotropic amines. Dev Biol. 1982 Mar;90(1):91–98. doi: 10.1016/0012-1606(82)90214-7. [DOI] [PubMed] [Google Scholar]
  24. King A. C., Hernaez-Davis L., Cuatrecasas P. Lysomotropic amines cause intracellular accumulation of receptors for epidermal growth factor. Proc Natl Acad Sci U S A. 1980 Jun;77(6):3283–3287. doi: 10.1073/pnas.77.6.3283. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Klee C. B., Crouch T. H., Richman P. G. Calmodulin. Annu Rev Biochem. 1980;49:489–515. doi: 10.1146/annurev.bi.49.070180.002421. [DOI] [PubMed] [Google Scholar]
  26. Knudsen K. A., Horwitz A. F., Buck C. A. A monoclonal antibody identifies a glycoprotein complex involved in cell-substratum adhesion. Exp Cell Res. 1985 Mar;157(1):218–226. doi: 10.1016/0014-4827(85)90164-8. [DOI] [PubMed] [Google Scholar]
  27. Knudsen K. A., Horwitz A. F. Differential inhibition of myoblast fusion. Dev Biol. 1978 Oct;66(2):294–307. doi: 10.1016/0012-1606(78)90239-7. [DOI] [PubMed] [Google Scholar]
  28. Knudsen K. A., Horwitz A. F. Tandem events in myoblast fusion. Dev Biol. 1977 Jul 15;58(2):328–338. doi: 10.1016/0012-1606(77)90095-1. [DOI] [PubMed] [Google Scholar]
  29. Knudsen K. A., Rao P. E., Damsky C. H., Buck C. A. Membrane glycoproteins involved in cell--substratum adhesion. Proc Natl Acad Sci U S A. 1981 Oct;78(10):6071–6075. doi: 10.1073/pnas.78.10.6071. [DOI] [PMC free article] [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. Magnani J. L., Thomas W. A., Steinberg M. S. Two distinct adhesion mechanisms in embryonic neural retina cells. I. A kinetic analysis. Dev Biol. 1981 Jan 15;81(1):96–105. doi: 10.1016/0012-1606(81)90351-1. [DOI] [PubMed] [Google Scholar]
  32. Nameroff M., Trotter J. A., Keller J. M., Munar E. Inhibition of cellular differentiation by phospholipase C. I. Effects of the enzyme on myogenesis and chondrogenesis in vitro. J Cell Biol. 1973 Jul;58(1):107–118. doi: 10.1083/jcb.58.1.107. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Neff N. T., Lowrey C., Decker C., Tovar A., Damsky C., Buck C., Horwitz A. F. A monoclonal antibody detaches embryonic skeletal muscle from extracellular matrices. J Cell Biol. 1982 Nov;95(2 Pt 1):654–666. doi: 10.1083/jcb.95.2.654. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Neff N., Decker C., Horwitz A. The kinetics of myoblast fusion. Exp Cell Res. 1984 Jul;153(1):25–31. doi: 10.1016/0014-4827(84)90444-0. [DOI] [PubMed] [Google Scholar]
  35. Ogou S. I., Yoshida-Noro C., Takeichi M. Calcium-dependent cell-cell adhesion molecules common to hepatocytes and teratocarcinoma stem cells. J Cell Biol. 1983 Sep;97(3):944–948. doi: 10.1083/jcb.97.3.944. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Ohkuma S., Poole B. Fluorescence probe measurement of the intralysosomal pH in living cells and the perturbation of pH by various agents. Proc Natl Acad Sci U S A. 1978 Jul;75(7):3327–3331. doi: 10.1073/pnas.75.7.3327. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Olden K., Law J., Hunter V. A., Romain R., Parent J. B. Inhibition of fusion of embryonic muscle cells in culture by tunicamycin is prevented by leupeptin. J Cell Biol. 1981 Jan;88(1):199–204. doi: 10.1083/jcb.88.1.199. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Olden K., Pratt R. M., Yamada K. M. Role of carbohydrates in protein secretion and turnover: effects of tunicamycin on the major cell surface glycoprotein of chick embryo fibroblasts. Cell. 1978 Mar;13(3):461–473. doi: 10.1016/0092-8674(78)90320-3. [DOI] [PubMed] [Google Scholar]
  39. Paterson B., Strohman R. C. Myosin synthesis in cultures of differentiating chicken embryo skeletal muscle. Dev Biol. 1972 Oct;29(2):113–138. doi: 10.1016/0012-1606(72)90050-4. [DOI] [PubMed] [Google Scholar]
  40. Peyriéras N., Hyafil F., Louvard D., Ploegh H. L., Jacob F. Uvomorulin: a nonintegral membrane protein of early mouse embryo. Proc Natl Acad Sci U S A. 1983 Oct;80(20):6274–6277. doi: 10.1073/pnas.80.20.6274. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Shainberg A., Yagil G., Yaffe D. Control of myogenesis in vitro by Ca 2 + concentration in nutritional medium. Exp Cell Res. 1969 Nov;58(1):163–167. doi: 10.1016/0014-4827(69)90127-x. [DOI] [PubMed] [Google Scholar]
  42. Shirayoshi Y., Okada T. S., Takeichi M. The calcium-dependent cell-cell adhesion system regulates inner cell mass formation and cell surface polarization in early mouse development. Cell. 1983 Dec;35(3 Pt 2):631–638. doi: 10.1016/0092-8674(83)90095-8. [DOI] [PubMed] [Google Scholar]
  43. Takeichi M., Atsumi T., Yoshida C., Uno K., Okada T. S. Selective adhesion of embryonal carcinoma cells and differentiated cells by Ca2+-dependent sites. Dev Biol. 1981 Oct 30;87(2):340–350. doi: 10.1016/0012-1606(81)90157-3. [DOI] [PubMed] [Google Scholar]
  44. Takeichi M. Functional correlation between cell adhesive properties and some cell surface proteins. J Cell Biol. 1977 Nov;75(2 Pt 1):464–474. doi: 10.1083/jcb.75.2.464. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Takeichi M., Ozaki H. S., Tokunaga K., Okada T. S. Experimental manipulation of cell surface to affect cellular recognition mechanisms. Dev Biol. 1979 May;70(1):195–205. doi: 10.1016/0012-1606(79)90016-2. [DOI] [PubMed] [Google Scholar]
  46. Thomas W. A., Steinberg M. S. Two distinct adhesion mechanisms in embryonic neural retina cells. II. An immunological analysis. Dev Biol. 1981 Jan 15;81(1):106–114. doi: 10.1016/0012-1606(81)90352-3. [DOI] [PubMed] [Google Scholar]
  47. Urushihara H., Ozaki H. S., Takeichi M. Immunological detection of cell surface components related with aggregation of Chinese hamster and chick embryonic cells. Dev Biol. 1979 May;70(1):206–216. doi: 10.1016/0012-1606(79)90017-4. [DOI] [PubMed] [Google Scholar]
  48. Yoshida-Noro C., Suzuki N., Takeichi M. Molecular nature of the calcium-dependent cell-cell adhesion system in mouse teratocarcinoma and embryonic cells studied with a monoclonal antibody. Dev Biol. 1984 Jan;101(1):19–27. doi: 10.1016/0012-1606(84)90112-x. [DOI] [PubMed] [Google Scholar]
  49. Yoshida C., Takeichi M. Teratocarcinoma cell adhesion: identification of a cell-surface protein involved in calcium-dependent cell aggregation. Cell. 1982 Feb;28(2):217–224. doi: 10.1016/0092-8674(82)90339-7. [DOI] [PubMed] [Google Scholar]
  50. van der Bosch J., Schudt C., Pette D. Influence of temperature, cholesterol, dipalmitoyllecithin and Ca2+ on the rate of muscle cell fusion. Exp Cell Res. 1973 Dec;82(2):433–438. doi: 10.1016/0014-4827(73)90362-5. [DOI] [PubMed] [Google Scholar]

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