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. 1991 Dec 2;115(6):1685–1694. doi: 10.1083/jcb.115.6.1685

Dystrophin-associated proteins are greatly reduced in skeletal muscle from mdx mice

PMCID: PMC2289197  PMID: 1757468

Abstract

Dystrophin, the protein product of the human Duchenne muscular dystrophy gene, exists in skeletal muscle as a large oligomeric complex that contains four glycoproteins of 156, 50, 43, and 35 kD and a protein of 59 kD. Here, we investigated the relative abundance of each of the components of the dystrophin-glycoprotein complex in skeletal muscle from normal and mdx mice, which are missing dystrophin. Immunoblot analysis using total muscle membranes from control and mdx mice of ages 1 d to 30 wk found that all of the dystrophin-associated proteins were greatly reduced (80-90%) in mdx mouse skeletal muscle. The specificity of the loss of the dystrophin-associated glycoproteins was demonstrated by the finding that the major glycoprotein composition of skeletal muscle membranes from normal and mdx mice was identical. Furthermore, skeletal muscle membranes from the dystrophic dy/dy mouse exhibited a normal density of dystrophin and dystrophin-associated proteins. Immunofluorescence microscopy confirmed the results from the immunoblot analysis and showed a drastically reduced density of dystrophin-associated proteins in mdx muscle cryosections compared with normal and dy/dy mouse muscle. Therefore, our results demonstrate that all of the dystrophin-associated proteins are significantly reduced in mdx skeletal muscle and suggest that the loss of dystrophin-associated proteins is due to the absence of dystrophin and not due to secondary effects of muscle fiber degradation.

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Selected References

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  1. Alloisio N., Morlé L., Bachir D., Guetarni D., Colonna P., Delaunay J. Red cell membrane sialoglycoprotein beta in homozygous and heterozygous 4.1(-) hereditary elliptocytosis. Biochim Biophys Acta. 1985 Jun 11;816(1):57–62. doi: 10.1016/0005-2736(85)90392-x. [DOI] [PubMed] [Google Scholar]
  2. Anderson J. E., Kao L., Bressler B. H., Gruenstein E. Analysis of dystrophin in fast- and slow-twitch skeletal muscles from mdx and dy2J mice at different ages. Muscle Nerve. 1990 Jan;13(1):6–11. doi: 10.1002/mus.880130103. [DOI] [PubMed] [Google Scholar]
  3. Arahata K., Ishiura S., Ishiguro T., Tsukahara T., Suhara Y., Eguchi C., Ishihara T., Nonaka I., Ozawa E., Sugita H. Immunostaining of skeletal and cardiac muscle surface membrane with antibody against Duchenne muscular dystrophy peptide. Nature. 1988 Jun 30;333(6176):861–863. doi: 10.1038/333861a0. [DOI] [PubMed] [Google Scholar]
  4. Bieber F. R., Hoffman E. P. Duchenne and Becker muscular dystrophies: genetics, prenatal diagnosis, and future prospects. Clin Perinatol. 1990 Dec;17(4):845–865. [PubMed] [Google Scholar]
  5. Bonilla E., Samitt C. E., Miranda A. F., Hays A. P., Salviati G., DiMauro S., Kunkel L. M., Hoffman E. P., Rowland L. P. Duchenne muscular dystrophy: deficiency of dystrophin at the muscle cell surface. Cell. 1988 Aug 12;54(4):447–452. doi: 10.1016/0092-8674(88)90065-7. [DOI] [PubMed] [Google Scholar]
  6. Bray G. M., Banker B. Q. An ultrastructural study of degeneration and necrosis of muscle in the dystrophic mouse. Acta Neuropathol. 1970;15(1):34–44. doi: 10.1007/BF00690687. [DOI] [PubMed] [Google Scholar]
  7. Buckle V. J., Guenet J. L., Simon-Chazottes D., Love D. R., Davies K. E. Localisation of a dystrophin-related autosomal gene to 6q24 in man, and to mouse chromosome 10 in the region of the dystrophia muscularis (dy) locus. Hum Genet. 1990 Aug;85(3):324–326. doi: 10.1007/BF00206755. [DOI] [PubMed] [Google Scholar]
  8. Bulfield G., Siller W. G., Wight P. A., Moore K. J. X chromosome-linked muscular dystrophy (mdx) in the mouse. Proc Natl Acad Sci U S A. 1984 Feb;81(4):1189–1192. doi: 10.1073/pnas.81.4.1189. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Campbell K. P., Kahl S. D. Association of dystrophin and an integral membrane glycoprotein. Nature. 1989 Mar 16;338(6212):259–262. doi: 10.1038/338259a0. [DOI] [PubMed] [Google Scholar]
  10. Campbell K. P., MacLennan D. H., Jorgensen A. O. Staining of the Ca2+-binding proteins, calsequestrin, calmodulin, troponin C, and S-100, with the cationic carbocyanine dye "Stains-all". J Biol Chem. 1983 Sep 25;258(18):11267–11273. [PubMed] [Google Scholar]
  11. Carpenter S., Karpati G. Duchenne muscular dystrophy: plasma membrane loss initiates muscle cell necrosis unless it is repaired. Brain. 1979 Mar;102(1):147–161. doi: 10.1093/brain/102.1.147. [DOI] [PubMed] [Google Scholar]
  12. Cohen P. T., Burchell A., Cohen P. The molecular basis of skeletal muscle phosphorylase kinase deficiency. Eur J Biochem. 1976 Jul 1;66(2):347–356. doi: 10.1111/j.1432-1033.1976.tb10524.x. [DOI] [PubMed] [Google Scholar]
  13. Delaporte C., Dautreaux B., Rouche A., Fardeau M. Changes in surface morphology and basal lamina of cultured muscle cells from Duchenne muscular dystrophy patients. J Neurol Sci. 1990 Jan;95(1):77–88. doi: 10.1016/0022-510x(90)90118-7. [DOI] [PubMed] [Google Scholar]
  14. Ervasti J. M., Campbell K. P. Membrane organization of the dystrophin-glycoprotein complex. Cell. 1991 Sep 20;66(6):1121–1131. doi: 10.1016/0092-8674(91)90035-w. [DOI] [PubMed] [Google Scholar]
  15. Ervasti J. M., Kahl S. D., Campbell K. P. Purification of dystrophin from skeletal muscle. J Biol Chem. 1991 May 15;266(14):9161–9165. [PubMed] [Google Scholar]
  16. Ervasti J. M., Ohlendieck K., Kahl S. D., Gaver M. G., Campbell K. P. Deficiency of a glycoprotein component of the dystrophin complex in dystrophic muscle. Nature. 1990 May 24;345(6273):315–319. doi: 10.1038/345315a0. [DOI] [PubMed] [Google Scholar]
  17. Hoffman E. P., Brown R. H., Jr, Kunkel L. M. Dystrophin: the protein product of the Duchenne muscular dystrophy locus. Cell. 1987 Dec 24;51(6):919–928. doi: 10.1016/0092-8674(87)90579-4. [DOI] [PubMed] [Google Scholar]
  18. Hoffman E. P., Fischbeck K. H., Brown R. H., Johnson M., Medori R., Loike J. D., Harris J. B., Waterston R., Brooke M., Specht L. Characterization of dystrophin in muscle-biopsy specimens from patients with Duchenne's or Becker's muscular dystrophy. N Engl J Med. 1988 May 26;318(21):1363–1368. doi: 10.1056/NEJM198805263182104. [DOI] [PubMed] [Google Scholar]
  19. Jorgensen A. O., Arnold W., Shen A. C., Yuan S. H., Gaver M., Campbell K. P. Identification of novel proteins unique to either transverse tubules (TS28) or the sarcolemma (SL50) in rabbit skeletal muscle. J Cell Biol. 1990 Apr;110(4):1173–1185. doi: 10.1083/jcb.110.4.1173. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Khurana T. S., Hoffman E. P., Kunkel L. M. Identification of a chromosome 6-encoded dystrophin-related protein. J Biol Chem. 1990 Oct 5;265(28):16717–16720. [PubMed] [Google Scholar]
  21. Knudson C. M., Chaudhari N., Sharp A. H., Powell J. A., Beam K. G., Campbell K. P. Specific absence of the alpha 1 subunit of the dihydropyridine receptor in mice with muscular dysgenesis. J Biol Chem. 1989 Jan 25;264(3):1345–1348. [PubMed] [Google Scholar]
  22. Koenig M., Kunkel L. M. Detailed analysis of the repeat domain of dystrophin reveals four potential hinge segments that may confer flexibility. J Biol Chem. 1990 Mar 15;265(8):4560–4566. [PubMed] [Google Scholar]
  23. Koenig M., Monaco A. P., Kunkel L. M. The complete sequence of dystrophin predicts a rod-shaped cytoskeletal protein. Cell. 1988 Apr 22;53(2):219–228. doi: 10.1016/0092-8674(88)90383-2. [DOI] [PubMed] [Google Scholar]
  24. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  25. Lee C. C., Pearlman J. A., Chamberlain J. S., Caskey C. T. Expression of recombinant dystrophin and its localization to the cell membrane. Nature. 1991 Jan 24;349(6307):334–336. doi: 10.1038/349334a0. [DOI] [PubMed] [Google Scholar]
  26. Love D. R., Hill D. F., Dickson G., Spurr N. K., Byth B. C., Marsden R. F., Walsh F. S., Edwards Y. H., Davies K. E. An autosomal transcript in skeletal muscle with homology to dystrophin. Nature. 1989 May 4;339(6219):55–58. doi: 10.1038/339055a0. [DOI] [PubMed] [Google Scholar]
  27. Lux S. E., Tse W. T., Menninger J. C., John K. M., Harris P., Shalev O., Chilcote R. R., Marchesi S. L., Watkins P. C., Bennett V. Hereditary spherocytosis associated with deletion of human erythrocyte ankyrin gene on chromosome 8. Nature. 1990 Jun 21;345(6277):736–739. doi: 10.1038/345736a0. [DOI] [PubMed] [Google Scholar]
  28. Menke A., Jockusch H. Decreased osmotic stability of dystrophin-less muscle cells from the mdx mouse. Nature. 1991 Jan 3;349(6304):69–71. doi: 10.1038/349069a0. [DOI] [PubMed] [Google Scholar]
  29. Mokri B., Engel A. G. Duchenne dystrophy: electron microscopic findings pointing to a basic or early abnormality in the plasma membrane of the muscle fiber. Neurology. 1975 Dec;25(12):1111–1120. doi: 10.1212/wnl.25.12.1111. [DOI] [PubMed] [Google Scholar]
  30. Ohlendieck K., Campbell K. P. Dystrophin constitutes 5% of membrane cytoskeleton in skeletal muscle. FEBS Lett. 1991 Jun 3;283(2):230–234. doi: 10.1016/0014-5793(91)80595-t. [DOI] [PubMed] [Google Scholar]
  31. Ohlendieck K., Ervasti J. M., Matsumura K., Kahl S. D., Leveille C. J., Campbell K. P. Dystrophin-related protein is localized to neuromuscular junctions of adult skeletal muscle. Neuron. 1991 Sep;7(3):499–508. doi: 10.1016/0896-6273(91)90301-f. [DOI] [PubMed] [Google Scholar]
  32. Ohlendieck K., Ervasti J. M., Snook J. B., Campbell K. P. Dystrophin-glycoprotein complex is highly enriched in isolated skeletal muscle sarcolemma. J Cell Biol. 1991 Jan;112(1):135–148. doi: 10.1083/jcb.112.1.135. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Partridge T. A., Morgan J. E., Coulton G. R., Hoffman E. P., Kunkel L. M. Conversion of mdx myofibres from dystrophin-negative to -positive by injection of normal myoblasts. Nature. 1989 Jan 12;337(6203):176–179. doi: 10.1038/337176a0. [DOI] [PubMed] [Google Scholar]
  34. Peterson G. L. A simplification of the protein assay method of Lowry et al. which is more generally applicable. Anal Biochem. 1977 Dec;83(2):346–356. doi: 10.1016/0003-2697(77)90043-4. [DOI] [PubMed] [Google Scholar]
  35. Sharp A. H., Campbell K. P. Characterization of the 1,4-dihydropyridine receptor using subunit-specific polyclonal antibodies. Evidence for a 32,000-Da subunit. J Biol Chem. 1989 Feb 15;264(5):2816–2825. [PubMed] [Google Scholar]
  36. Sicinski P., Geng Y., Ryder-Cook A. S., Barnard E. A., Darlison M. G., Barnard P. J. The molecular basis of muscular dystrophy in the mdx mouse: a point mutation. Science. 1989 Jun 30;244(4912):1578–1580. doi: 10.1126/science.2662404. [DOI] [PubMed] [Google Scholar]
  37. Torres L. F., Duchen L. W. The mutant mdx: inherited myopathy in the mouse. Morphological studies of nerves, muscles and end-plates. Brain. 1987 Apr;110(Pt 2):269–299. doi: 10.1093/brain/110.2.269. [DOI] [PubMed] [Google Scholar]
  38. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Urayama O., Shutt H., Sweadner K. J. Identification of three isozyme proteins of the catalytic subunit of the Na,K-ATPase in rat brain. J Biol Chem. 1989 May 15;264(14):8271–8280. [PubMed] [Google Scholar]
  40. Watkins S. C., Hoffman E. P., Slayter H. S., Kunkel L. M. Immunoelectron microscopic localization of dystrophin in myofibres. Nature. 1988 Jun 30;333(6176):863–866. doi: 10.1038/333863a0. [DOI] [PubMed] [Google Scholar]
  41. Weller B., Karpati G., Carpenter S. Dystrophin-deficient mdx muscle fibers are preferentially vulnerable to necrosis induced by experimental lengthening contractions. J Neurol Sci. 1990 Dec;100(1-2):9–13. doi: 10.1016/0022-510x(90)90005-8. [DOI] [PubMed] [Google Scholar]
  42. White R. A., Birkenmeier C. S., Lux S. E., Barker J. E. Ankyrin and the hemolytic anemia mutation, nb, map to mouse chromosome 8: presence of the nb allele is associated with a truncated erythrocyte ankyrin. Proc Natl Acad Sci U S A. 1990 Apr;87(8):3117–3121. doi: 10.1073/pnas.87.8.3117. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Zubrzycka-Gaarn E. E., Bulman D. E., Karpati G., Burghes A. H., Belfall B., Klamut H. J., Talbot J., Hodges R. S., Ray P. N., Worton R. G. The Duchenne muscular dystrophy gene product is localized in sarcolemma of human skeletal muscle. Nature. 1988 Jun 2;333(6172):466–469. doi: 10.1038/333466a0. [DOI] [PubMed] [Google Scholar]

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