New mdx mutation disrupts expression of muscle and nonmuscle isoforms of dystrophin

Abstract

The dystrophin gene encodes several tissue–specific protein isoforms that are generated by alternative splicing and by transcription from at least three separate promoters. We have characterized the mutation in a new strain of mdx mice that results in aberrant splicing of both the 14 and 4.8 kilobase dystrophin mRNAs and disrupts expression of the muscle and brain 427K and nonmuscle 70K isoforms of dystrophin. In contrast, we have determined that expression of the 70K isoform is normal in the original mdx mutant. We have cloned the unique 5′ exon of the murine 4.8 kb mRNA and have analysed the tissue distribution and aberrant splicing of this transcript in the mdx3Cv mutant. This new mdx mutant will provide an improved model system for functional studies of the dystrophin C–terminus in muscle and nonmuscle tissues.

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References

  1. 1

    Hoffman, E.P., Brown, R.H. Jr., & Kunkel, L.M. Dystrophin: the protein product of the Duchenne muscular dystrophy locus. Cell 51, 919–928 (1987).

    CAS  Article  Google Scholar 

  2. 2

    Emery, A.E.H. Duchenne Muscular Dystrophy, Oxford Monographs on Medical Genetics No. 15, Oxford Medical Publications, Oxford, 1987).

    Google Scholar 

  3. 3

    Nudel, U. et al. Duchenne muscular dystrophy gene product is not identical in muscle and brain. Nature 337, 76–78 (1988).

    Article  Google Scholar 

  4. 4

    Feener, C.A., Koenig, M. & Kunkel, L.M. Alternative splicing of human dystrophin mRNA generates isoforms at the carboxy terminus. Nature 338, 509–511 (1989).

    CAS  Article  Google Scholar 

  5. 5

    Lederfein, D. et al. A 71-kilodalton protein is a major product of the Duchenne muscular dystrophy gene in brain and other nonmuscle tissues. Proc. natn. Acad. Sci. U.S.A. 89, 5346–5350 (1992).

    CAS  Article  Google Scholar 

  6. 6

    Blake, D.J. et al. Characterization of a 4.8kb transcript from the Duchenne muscular dystrophy locus expressed in Scwannoma cells. Hum. molec. Genet. 1, 103–110 (1992).

    CAS  Article  Google Scholar 

  7. 7

    Koenig, M. et al. The molecular basis for Duchenne versus Becker muscular dystrophy: correlation of severity with type of deletion. Am. J. hum. Genet. 46, 498–506 (1989).

    Google Scholar 

  8. 8

    Gillard, E.F. et al. Molecular and phenotypic analysis of patients with deletions within the deletion-rich region of the Duchenne muscular dystrophy (DMD) gene. Am. J. hum. Genet. 45, 507–520 (1989).

    CAS  PubMed  PubMed Central  Google Scholar 

  9. 9

    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 345, 315–319 (1990).

    CAS  Article  Google Scholar 

  10. 10

    Bulfield, G., Siller, W.G., Wight, P.A. & Moore, K.J. X chromosome-linked muscular dystrophy (mdx) in the mouse. Proc. natn. Acad. Sci. U.S.A. 81, 1189–1192 (1984).

    CAS  Article  Google Scholar 

  11. 11

    Chamberlain, J.S. et al. Expression of the murine Duchenne muscular dystrophy gene in muscle and brain. Science 239, 1416–1418 (1988).

    CAS  Article  Google Scholar 

  12. 12

    Sicinski, P. et al. The molecular basis of muscular dystrophy in the mdx mouse: a point mutation. Science 244, 1578–1580 (1989).

    CAS  Article  Google Scholar 

  13. 13

    Bies, R.D. et al. Human and murine dystrophin mRNA transcripts are differentially expressed during skeletal muscle, heart, and brain development. Nucl. Acids Res. 20, 1725–1731 (1992).

    CAS  Article  Google Scholar 

  14. 14

    Chapman, V.M., Miller, D.M., Armstrong, D. & Caskey, C.T. Recovery of induced mutations for X chromosome-linked muscular dystrophy in mice. Proc. natn. Acad. Sci. U.S.A. 86, 1292–1296 (1989).

    CAS  Article  Google Scholar 

  15. 15

    Chamberlain, J.S., Farwell, N.J., Chamberlain, J.R., Cox, G.A. & Caskey, C.T. PCR analysis of dystrophin gene mutation and expression. J. Cell Biochem. 46, 255–259 (1991).

    CAS  Article  Google Scholar 

  16. 16

    Green, M.R. Pre-mRNA splicing. Ann. Rev. Genet. 20, 671–708 (1986).

    CAS  Article  Google Scholar 

  17. 17

    Roberts, R.G., Coffey, A.J., Bobrow, M. & Bentley, D.R. Determination of the exon structure of the distal portion of the dystrophin gene by vectorette PCR. Genomics 13, 942–950 (1992).

    CAS  Article  Google Scholar 

  18. 18

    Kozak, M. Point mutations define a sequence flanking the AUG initiator codon that modulates translation by eukaryotic ribosomes. Cell 44, 283–292 (1986).

    CAS  Article  Google Scholar 

  19. 19

    Chamberlain, J.S. et al. Analysis of Duchenne muscular dystrophy mutations in mice and humans. In Cellular and Molecular Biology of Muscle Development, UCLA Symposia on Molecular and Cellular Biology (eds Stockdale, F. & Kedes, L.) 951–962 (Alan R. Liss, New York, 1989).

    Google Scholar 

  20. 20

    Lee, C.C., Pearlman, J.A., Chamberlain, J.S. & Caskey, C.T. Expression of recombinant dystrophin and its localization to the cell membrane. Nature 349, 334–336 (1991).

    CAS  Article  Google Scholar 

  21. 21

    Danko, I., Chapman, V. & Wolff, J.A. The frequency of revertants in mdx mouse genetic models for Duchenne muscular dystrophy. Pediatr. Res. 32, 128–131 (1992).

    CAS  Article  Google Scholar 

  22. 22

    Stedman, H.H. et al. The mdx mouse diaphragm reproduces the degenerative changes of Duchenne muscular dystrophy. Nature 352, 536–539 (1991).

    CAS  Article  Google Scholar 

  23. 23

    Metherall, J.E., Collins, F.S., Pan, J., Weissman, S.M. & Forget, B.G. Beta-zero thalassemia caused by a base substitution that creates an alternative splice acceptor site in an intron. EMBO J. 5, 2551–2557 (1986).

    CAS  Article  Google Scholar 

  24. 24

    Fu, X., Colgan, J.D. & Manley, J.L. Multiple cis-acting sequence elements are required for efficient splicing of Simian Virus 40 small-t antigen pre-mRNA. Molec. cell. Biol. 8, 3582–3590 (1988).

    CAS  Article  Google Scholar 

  25. 25

    Smith, C.W.J., Porro, E.B., Patton, J.G. & Nadal-Ginard, B. Scanning from an independently specified branch point defines the 3′ splice site of mammalian introns. Nature 342, 243–247 (1989).

    CAS  Article  Google Scholar 

  26. 26

    Hoffman, E.P. et al. Characterization of dystrophin in muscle-biopsy specimens from patients with Duchenne's or Becker's muscular dystrophy. New Engl. J. Med. 318, 1363–1368 (1988).

    CAS  Article  Google Scholar 

  27. 27

    Beggs, A.H. et al. Exploring the molecular basis for variability among patients with Becker muscular dystrophy: dystrophin gene and protein studies. Am. J. hum. Genet. 49, 54–67 (1991).

    CAS  PubMed  PubMed Central  Google Scholar 

  28. 28

    Koenig, M., Monaco, A.P. & Kunkel, L.M. The complete sequence of dystrophin predicts a rod-shaped cytoskeletal protein. Cell 53, 219–226 (1988).

    CAS  Article  Google Scholar 

  29. 29

    Fong, P.Y., Turner, P.R., Denetclaw, W.F. & Steinhardt, R.A. Increased activity of calcium leak channels in myotubes of Duchenne human and mdx mouse origin. Science 250, 673–676 (1990).

    CAS  Article  Google Scholar 

  30. 30

    Ohlendieck, K. & Campbell, K.P. Dystrophin-associated proteins are greatly reduced in skeletal muscle from mdx mice. J. cell Biol. 115, 1685–1694 (1991).

    CAS  Article  Google Scholar 

  31. 31

    Ibraghimov-Beskrovnaya, O. et al. Primary structure of dystrophin-associated glycoproteins linking dystrophin to the extracellular matrix. Nature 355, 696–702 (1992).

    CAS  Article  Google Scholar 

  32. 32

    Hoffman, E.P. et al. Is the carboxyl-terminus of dystrophin required for membrane association? A novel, severe case of Duchenne muscular dystrophy. Ann. Neurol. 30, 605–610 (1991).

    CAS  Article  Google Scholar 

  33. 33

    Roberts, R.G., Bobrow, M. & Bentley, D.R. Point mutations in the dystrophin gene. Proc. natn. Acad. Sci. U.S.A. 89, 2331–2335 (1992).

    CAS  Article  Google Scholar 

  34. 34

    Towbin, J.A. et al. Characterization of patients with glycerol kinase deficiency utilizing cDNA probes for the Duchenne muscular dystrophy locus. Hum. Genet. 83, 122–126 (1989).

    CAS  Article  Google Scholar 

  35. 35

    Ochman, H., Medhora, M.M., Garza, D. & Hartl, D.L. Amplification of flanking sequences by inverse PCR. in PCR Protocols. A Guide to Methods and Applications (eds Innis, et al.) 219–227 (Academic, San Diego, 1990).

    Google Scholar 

  36. 36

    Chirgwin, J.M., Przybyla, A.E., MacDonald, R.J. & Rutter, W.J. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry 18, 5294–5299 (1979).

    CAS  Article  Google Scholar 

  37. 37

    Frohman, M.A., Dush, M.K. & Martin, G.R. Rapid production of full-length cDNAs from rare transcripts: amplification using a single gene-specific oligonucleotide primer. Proc. natn. Acad. Sci. U.S.A. 85, 8998–9002 (1988).

    CAS  Article  Google Scholar 

  38. 38

    Jaynes, J.B., Chamberlain, J.S., Buskin, J.N., Johnson, J.E. & Hauschka, S.D. Transcriptional regulation of the muscle creatine kinase gene and regulated expression in transfected mouse myoblasts. Molec. cell. Biol. 6, 2855–2864 (1986).

    CAS  Article  Google Scholar 

  39. 39

    Burrow, K.L. et al. Dystrophin expression and somatic reversion in prednisone-treated and untreated Duchenne dystrophy. CIDD Study Group. Neurology 41, 661–666 (1991).

    CAS  Article  Google Scholar 

  40. 40

    Kaatz, K.W., Bazzett, T.J. & Albin, R.L. A new, simple myelin stain. Brain Res. Bull. 29, 697–698 (1992).

    CAS  Article  Google Scholar 

  41. 41

    Maconochie, M.K., Brown, S.D. & Greenfield, A.J. Sequence analysis of two exons from the murine dystrophin locus. Mam. Genome. 2, 64–68 (1992).

    CAS  Article  Google Scholar 

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Cox, G., Phelps, S., Chapman, V. et al. New mdx mutation disrupts expression of muscle and nonmuscle isoforms of dystrophin. Nat Genet 4, 87–93 (1993). https://doi.org/10.1038/ng0593-87

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