Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Dystrophin expression in muscle stem cells regulates their polarity and asymmetric division

Abstract

Dystrophin is expressed in differentiated myofibers, in which it is required for sarcolemmal integrity, and loss-of-function mutations in the gene that encodes it result in Duchenne muscular dystrophy (DMD), a disease characterized by progressive and severe skeletal muscle degeneration. Here we found that dystrophin is also highly expressed in activated muscle stem cells (also known as satellite cells), in which it associates with the serine-threonine kinase Mark2 (also known as Par1b), an important regulator of cell polarity. In the absence of dystrophin, expression of Mark2 protein is downregulated, resulting in the inability to localize the cell polarity regulator Pard3 to the opposite side of the cell. Consequently, the number of asymmetric divisions is strikingly reduced in dystrophin-deficient satellite cells, which also display a loss of polarity, abnormal division patterns (including centrosome amplification), impaired mitotic spindle orientation and prolonged cell divisions. Altogether, these intrinsic defects strongly reduce the generation of myogenic progenitors that are needed for proper muscle regeneration. Therefore, we conclude that dystrophin has an essential role in the regulation of satellite cell polarity and asymmetric division. Our findings indicate that muscle wasting in DMD not only is caused by myofiber fragility, but also is exacerbated by impaired regeneration owing to intrinsic satellite cell dysfunction.

This is a preview of subscription content, access via your institution

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Dystrophin expression in satellite cells.
Figure 2: Impaired satellite stem cell asymmetric divisions and reduced generation of myogenic progenitors in the absence of dystrophin.
Figure 3: Dystrophin regulates PAR polarity protein localization.
Figure 4: PAR polarity proteins are required for muscle stem cell asymmetric divisions.
Figure 5: Dystrophin-deficient satellite cells display impaired mitotic spindle orientation and loss of apicobasal division.
Figure 6: Dystrophin-deficient satellite cells have reduced ability to generate myogenic progenitors in regenerating muscle.

Similar content being viewed by others

Accession codes

Primary accessions

Gene Expression Omnibus

References

  1. Anderson, M.S. & Kunkel, L.M. The molecular and biochemical basis of Duchenne muscular dystrophy. Trends Biochem. Sci. 17, 289–292 (1992).

    CAS  PubMed  Google Scholar 

  2. Cohn, R.D. & Campbell, K.P. Molecular basis of muscular dystrophies. Muscle Nerve 23, 1456–1471 (2000).

    CAS  PubMed  Google Scholar 

  3. Koenig, M. et al. Complete cloning of the Duchenne muscular dystrophy (DMD) cDNA and preliminary genomic organization of the DMD gene in normal and affected individuals. Cell 50, 509–517 (1987).

    CAS  PubMed  Google Scholar 

  4. Serrano, A.L. et al. Cellular and molecular mechanisms regulating fibrosis in skeletal muscle repair and disease. Curr. Top. Dev. Biol. 96, 167–201 (2011).

    CAS  PubMed  Google Scholar 

  5. Cohn, R.D. et al. Disruption of Dag1 in differentiated skeletal muscle reveals a role for dystroglycan in muscle regeneration. Cell 110, 639–648 (2002).

    CAS  PubMed  Google Scholar 

  6. Sacco, A. et al. Short telomeres and stem cell exhaustion model Duchenne muscular dystrophy in mdx/mTR mice. Cell 143, 1059–1071 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  7. Webster, C. & Blau, H.M. Accelerated age-related decline in replicative life span of Duchenne muscular dystrophy myoblasts: implications for cell and gene therapy. Somat. Cell Mol. Genet. 16, 557–565 (1990).

    CAS  PubMed  Google Scholar 

  8. Kottlors, M. & Kirschner, J. Elevated satellite cell number in Duchenne muscular dystrophy. Cell Tissue Res. 340, 541–548 (2010).

    PubMed  Google Scholar 

  9. Reimann, J., Irintchev, A. & Wernig, A. Regenerative capacity and the number of satellite cells in soleus muscles of normal and mdx mice. Neuromuscul. Disord. 10, 276–282 (2000).

    CAS  PubMed  Google Scholar 

  10. Chakkalakal, J.V. et al. Early forming label-retaining muscle stem cells require p27kip1 for maintenance of the primitive state. Development 141, 1649–1659 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  11. Yamashita, K. et al. The eighth and ninth tandem spectrin-like repeats of utrophin cooperatively form a functional unit to interact with polarity-regulating kinase PAR-1b. Biochem. Biophys. Res. Commun. 391, 812–817 (2010).

    CAS  PubMed  Google Scholar 

  12. Masuda-Hirata, M. et al. Intracellular polarity protein PAR-1 regulates extracellular laminin assembly by regulating the dystroglycan complex. Genes Cells 14, 835–850 (2009).

    CAS  PubMed  Google Scholar 

  13. Neumüller, R.A. & Knoblich, J.A. Dividing cellular asymmetry: asymmetric cell division and its implications for stem cells and cancer. Genes Dev. 23, 2675–2699 (2009).

    PubMed  PubMed Central  Google Scholar 

  14. Knoblich, J.A. Asymmetric cell division: recent developments and their implications for tumor biology. Nat. Rev. Mol. Cell Biol. 11, 849–860 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  15. Goulas, S., Conder, R. & Knoblich, J.A. The Par complex and integrins direct asymmetric cell division in adult intestinal stem cells. Cell Stem Cell 11, 529–540 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  16. Troy, A. et al. Coordination of satellite cell activation and self-renewal by Par-complex–dependent asymmetric activation of p38-α/β MAPK. Cell Stem Cell 11, 541–553 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  17. Miranda, A.F. et al. Immunocytochemical study of dystrophin in muscle cultures from patients with Duchenne muscular dystrophy and unaffected control patients. Am. J. Pathol. 132, 410–416 (1988).

    CAS  PubMed  PubMed Central  Google Scholar 

  18. Huard, J., Labrecque, C., Dansereau, G., Robitaille, L. & Tremblay, J.P. Dystrophin expression in myotubes formed by the fusion of normal and dystrophic myoblasts. Muscle Nerve 14, 178–182 (1991).

    CAS  PubMed  Google Scholar 

  19. Bentzinger, C.F. et al. Fibronectin regulates Wnt7a signaling and satellite cell expansion. Cell Stem Cell 12, 75–87 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  20. Kuang, S., Kuroda, K., Le Grand, F. & Rudnicki, M.A. Asymmetric self-renewal and commitment of satellite stem cells in muscle. Cell 129, 999–1010 (2007).

    CAS  PubMed  PubMed Central  Google Scholar 

  21. Rocheteau, P., Gayraud-Morel, B., Siegl-Cachedenier, I., Blasco, M. & Tajbakhsh, S. A subpopulation of adult skeletal muscle stem cells retains all template DNA strands after cell division. Cell 148, 112–125 (2012).

    CAS  PubMed  Google Scholar 

  22. Ono, Y. et al. Slow-dividing satellite cells retain long-term self-renewal ability in adult muscle. J. Cell Sci. 125, 1309–1317 (2012).

    CAS  PubMed  Google Scholar 

  23. Nishijo, K. et al. Biomarker system for studying muscle, stem cells and cancer in vivo. FASEB J. 23, 2681–2690 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  24. Fredriksson, S. et al. Protein detection using proximity-dependent DNA ligation assays. Nat. Biotechnol. 20, 473–477 (2002).

    CAS  PubMed  Google Scholar 

  25. Hurov, J.B. et al. Immune system dysfunction and autoimmune disease in mice lacking Emk (Par-1) protein kinase. Mol. Cell. Biol. 21, 3206–3219 (2001).

    CAS  PubMed  PubMed Central  Google Scholar 

  26. Lu, M.S. & Johnston, C.A. Molecular pathways regulating mitotic spindle orientation in animal cells. Development 140, 1843–1856 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  27. Wang, G., Jiang, Q. & Zhang, C. The role of mitotic kinases in coupling the centrosome cycle with the assembly of the mitotic spindle. J. Cell Sci. 127, 4111–4122 (2014).

    CAS  PubMed  Google Scholar 

  28. Carmena, M. & Earnshaw, W.C. The cellular geography of Aurora kinases. Nat. Rev. Mol. Cell Biol. 4, 842–854 (2003).

    CAS  PubMed  Google Scholar 

  29. Fukada, S. et al. Molecular signature of quiescent satellite cells in adult skeletal muscle. Stem Cells 25, 2448–2459 (2007).

    CAS  PubMed  Google Scholar 

  30. Liu, L. et al. Chromatin modifications as determinants of muscle stem cell quiescence and chronological aging. Cell Rep. 4, 189–204 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  31. Tennyson, C.N., Klamut, H.J. & Worton, R.G. The human dystrophin gene requires 16 hours to be transcribed and is cotranscriptionally spliced. Nat. Genet. 9, 184–190 (1995).

    CAS  PubMed  Google Scholar 

  32. Lewandowski, K.T. & Piwnica-Worms, H. Phosphorylation of the E3 ubiquitin ligase RNF41 by the kinase Par-1b is required for epithelial cell polarity. J. Cell Sci. 127, 315–327 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  33. Knoblich, J.A. Mechanisms of asymmetric stem cell division. Cell 132, 583–597 (2008).

    CAS  PubMed  Google Scholar 

  34. Kwon, M. et al. Mechanisms to suppress multipolar divisions in cancer cells with extra centrosomes. Genes Dev. 22, 2189–2203 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  35. Yennek, S., Burute, M., Théry, M. & Tajbakhsh, S. Cell adhesion geometry regulates nonrandom DNA segregation and asymmetric cell fates in mouse skeletal muscle stem cells. Cell Rep. 7, 961–970 (2014).

    CAS  PubMed  Google Scholar 

  36. Marumoto, T., Zhang, D. & Saya, H. Aurora-A—a guardian of poles. Nat. Rev. Cancer 5, 42–50 (2005).

    CAS  PubMed  Google Scholar 

  37. Kollu, S., Abou-Khalil, R., Shen, C. & Brack, A.S. The spindle assembly checkpoint safeguards genomic integrity of skeletal muscle satellite cells. Stem Cell Reports 4, 1061–1074 (2015).

    PubMed  PubMed Central  Google Scholar 

  38. Galluzzi, L. et al. Molecular definitions of cell death subroutines: recommendations of the Nomenclature Committee on Cell Death 2012. Cell Death Differ. 19, 107–120 (2012).

    CAS  PubMed  Google Scholar 

  39. Steigemann, P. et al. Aurora B–mediated abscission checkpoint protects against tetraploidization. Cell 136, 473–484 (2009).

    PubMed  Google Scholar 

  40. Ross, J. et al. Defects in glycosylation impair satellite stem cell function and niche composition in the muscles of the dystrophic Largemyd mouse. Stem Cells 30, 2330–2341 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  41. Irintchev, A., Zweyer, M. & Wernig, A. Impaired functional and structural recovery after muscle injury in dystrophic mdx mice. Neuromuscul. Disord. 7, 117–125 (1997).

    CAS  PubMed  Google Scholar 

  42. Hayashiji, N. et al. G-CSF supports long-term muscle regeneration in mouse models of muscular dystrophy. Nat. Commun. 6, 6745 (2015).

    CAS  PubMed  Google Scholar 

  43. Giliberto, F., Ferreiro, V., Dalamon, V. & Szijan, I. Dystrophin deletions and cognitive impairment in Duchenne/Becker muscular dystrophy. Neurol. Res. 26, 83–87 (2004).

    PubMed  Google Scholar 

  44. De Stefano, M.E., Leone, L., Lombardi, L. & Paggi, P. Lack of dystrophin leads to the selective loss of superior cervical ganglion neurons projecting to muscular targets in genetically dystrophic mdx mice. Neurobiol. Dis. 20, 929–942 (2005).

    CAS  PubMed  Google Scholar 

  45. Morrison, S.J. & Kimble, J. Asymmetric and symmetric stem-cell divisions in development and cancer. Nature 441, 1068–1074 (2006).

    CAS  PubMed  Google Scholar 

  46. Wang, Y. et al. Dystrophin is a tumor suppressor in human cancers with myogenic programs. Nat. Genet. 46, 601–606 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  47. Long, C. et al. Prevention of muscular dystrophy in mice by CRISPR/Cas9–mediated editing of germline DNA. Science 345, 1184–1188 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  48. Wang, B., Li, J. & Xiao, X. Adeno-associated virus vector carrying human minidystrophin genes effectively ameliorates muscular dystrophy in mdx mouse model. Proc. Natl. Acad. Sci. USA 97, 13714–13719 (2000).

    CAS  PubMed  Google Scholar 

  49. Wang, J. et al. Dilated cardiomyopathy and atrioventricular conduction blocks induced by heart-specific inactivation of mitochondrial DNA gene expression. Nat. Genet. 21, 133–137 (1999).

    CAS  PubMed  Google Scholar 

  50. Tajbakhsh, S. et al. Gene targeting the myf-5 locus with nlacZ reveals expression of this myogenic factor in mature skeletal muscle fibers as well as early embryonic muscle. Dev. Dyn. 206, 291–300 (1996).

    CAS  PubMed  Google Scholar 

  51. Tallquist, M.D., Weismann, K.E., Hellström, M. & Soriano, P. Early myotome specification regulates PDGFA expression and axial skeleton development. Development 127, 5059–5070 (2000).

    CAS  PubMed  Google Scholar 

  52. Srinivas, S. et al. Cre reporter strains produced by targeted insertion of EYFP and ECFP into the ROSA26 locus. BMC Dev. Biol. 1, 4 (2001).

    CAS  PubMed  PubMed Central  Google Scholar 

  53. von Maltzahn, J., Jones, A.E., Parks, R.J. & Rudnicki, M.A. Pax7 is critical for the normal function of satellite cells in adult skeletal muscle. Proc. Natl. Acad. Sci. USA 110, 16474–16479 (2013).

    CAS  PubMed  Google Scholar 

  54. Pasut, A., Oleynik, P. & Rudnicki, M.A. Isolation of muscle stem cells by fluorescence-activated cell sorting cytometry. Methods Mol. Biol. 798, 53–64 (2012).

    CAS  PubMed  Google Scholar 

  55. Pasut, A., Jones, A.E. & Rudnicki, M.A. Isolation and culture of individual myofibers and their satellite cells from adult skeletal muscle. J. Vis. Exp. 73, 50074 (2013).

    Google Scholar 

  56. Briguet, A., Courdier-Fruh, I., Foster, M., Meier, T. & Magyar, J.P. Histological parameters for the quantitative assessment of muscular dystrophy in the mdx mouse. Neuromuscul. Disord. 14, 675–682 (2004).

    PubMed  Google Scholar 

  57. Lee, C.-Y. et al. Drosophila Aurora A kinase inhibits neuroblast self-renewal by regulating aPKC/Numb cortical polarity and spindle orientation. Genes Dev. 20, 3464–3474 (2006).

    CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We thank J. Dilworth and L. Megeney for careful reading of the manuscript. We also thank J. Ritchie for animal husbandry, and J. Fernandes and P. Oleynik of the flow cytometry facility of the StemCore laboratories for technical assistance. N.A.D. is supported by a Postdoctoral Fellowship from the Canadian Institutes of Health Research (CIHR); Y.X.W. is supported by fellowships from the Queen Elizabeth II Graduate Scholarships in Science and Technology and the CIHR; J.v.M. was supported by a grant from the Deutsche Forschungsgemeinschaft; C.F.B. was supported by a grant from the Swiss National Science Foundation; C.E.B. is supported by a Postdoctoral Fellowship from the Ontario Institute for Regenerative Medicine; and M.A.R. holds the Canada Research Chair in Molecular Genetics. These studies were carried out with support from grants to M.A.R. from the US National Institutes for Health (grant no. RO1AR044031), the CIHR (grant no. MOP-12080 and MOP-81288), the E-Rare-2 program from the CIHR and Muscular Dystrophy Canada (grant no. ERA-132935), the Muscular Dystrophy Association, the Stem Cell Network and the Ministry of Research and Innovation (MRI), Government of Ontario (grant no. ORF-RE05-084).

Author information

Authors and Affiliations

Authors

Contributions

N.A.D. and Y.X.W. designed and carried out experiments, analyzed results and wrote the manuscript. J.v.M. designed and conducted experiments and analyzed results. A.P., C.F.B. and C.E.B. conducted experiments. M.A.R. designed experiments, analyzed results, wrote the manuscript and provided financial support.

Corresponding author

Correspondence to Michael A Rudnicki.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Source data

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Dumont, N., Wang, Y., von Maltzahn, J. et al. Dystrophin expression in muscle stem cells regulates their polarity and asymmetric division. Nat Med 21, 1455–1463 (2015). https://doi.org/10.1038/nm.3990

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nm.3990

This article is cited by

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing