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.

  • Letter
  • Published:

ISPD loss-of-function mutations disrupt dystroglycan O-mannosylation and cause Walker-Warburg syndrome

Nature Genetics volume 44, pages 575–580 (2012)Cite this article

Subjects

Abstract

Walker-Warburg syndrome (WWS) is clinically defined as congenital muscular dystrophy that is accompanied by a variety of brain and eye malformations. It represents the most severe clinical phenotype in a spectrum of diseases associated with abnormal post-translational processing of α-dystroglycan that share a defect in laminin-binding glycan synthesis1. Although mutations in six genes have been identified as causes of WWS, only half of all individuals with the disease can currently be diagnosed on this basis2. A cell fusion complementation assay in fibroblasts from undiagnosed individuals with WWS was used to identify five new complementation groups. Further evaluation of one group by linkage analysis and targeted sequencing identified recessive mutations in the ISPD gene (encoding isoprenoid synthase domain containing). The pathogenicity of the identified ISPD mutations was shown by complementation of fibroblasts with wild-type ISPD. Finally, we show that recessive mutations in ISPD abolish the initial step in laminin-binding glycan synthesis by disrupting dystroglycan O-mannosylation. This establishes a new mechanism for WWS pathophysiology.

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

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

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

Figure 1: Status of α-dystroglycan glycosylation in dermal fibroblasts from subjects with dystroglycanopathy.
Figure 2: Cell fusion experiments reveal new genetic complementation groups.
Figure 3: Clinical presentation and α-dystroglycan glycosylation defect in ISPD-deficient subject P1 with WWS.
Figure 4: Identification and validation of ISPD as a disease-associated gene in subjects with WWS.
Figure 5: ISPD loss-of-function mutations cause α-dystroglycan O-mannosylation defect.

Similar content being viewed by others

References

  1. Vajsar, J. & Schachter, H. Walker-Warburg syndrome. Orphanet J. Rare Dis. 1, 29 (2006).

    Article  PubMed  PubMed Central  Google Scholar 

  2. Godfrey, C., Foley, A.R., Clement, E. & Muntoni, F. Dystroglycanopathies: coming into focus. Curr. Opin. Genet. Dev. 21, 278–285 (2011).

    Article  CAS  PubMed  Google Scholar 

  3. Michele, D.E. et al. Post-translational disruption of dystroglycan-ligand interactions in congenital muscular dystrophies. Nature 418, 417–422 (2002).

    Article  CAS  PubMed  Google Scholar 

  4. Cao, W. et al. Identification of α-dystroglycan as a receptor for lymphocytic choriomeningitis virus and Lassa fever virus. Science 282, 2079–2081 (1998).

    Article  CAS  PubMed  Google Scholar 

  5. Barresi, R. & Campbell, K.P. Dystroglycan: from biosynthesis to pathogenesis of human disease. J. Cell Sci. 119, 199–207 (2006).

    Article  CAS  PubMed  Google Scholar 

  6. Satz, J.S. et al. Brain and eye malformations resembling Walker-Warburg syndrome are recapitulated in mice by dystroglycan deletion in the epiblast. J. Neurosci. 28, 10567–10575 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Lommel, M. et al. Correlation of enzyme activity and clinical phenotype in POMT1-associated dystroglycanopathies. Neurology 74, 157–164 (2010).

    Article  CAS  PubMed  Google Scholar 

  8. Barresi, R. et al. LARGE can functionally bypass α-dystroglycan glycosylation defects in distinct congenital muscular dystrophies. Nat. Med. 10, 696–703 (2004).

    Article  CAS  PubMed  Google Scholar 

  9. Clarke, N.F. et al. Congenital muscular dystrophy type 1D (MDC1D) due to a large intragenic insertion/deletion, involving intron 10 of the LARGE gene. Eur. J. Hum. Genet. 19, 452–457 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Inamori, K. et al. Dystroglycan function requires xylosyl- and glucuronyltransferase activities of LARGE. Science 335, 93–96 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Masselot, M. & De Robichon-Szulmajster, H. Methionine biosynthesis in Saccharomyces cerevisiae. I. Genetical analysis of auxotrophic mutants. Mol. Gen. Genet. 139, 121–132 (1975).

    Article  CAS  PubMed  Google Scholar 

  12. Stanley, P. Membrane mutants of animal cells: rapid identification of those with a primary defect in glycosylation. Mol. Cell. Biol. 5, 923–929 (1985).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Pontecorvo, G., Riddle, P.N. & Hales, A. Time and mode of fusion of human fibroblasts treated with polyethylene glycol (PEG). Nature 265, 257–258 (1977).

    Article  CAS  PubMed  Google Scholar 

  14. Kanoff, R.J. et al. Walker-Warburg syndrome: neurologic features and muscle membrane structure. Pediatr. Neurol. 18, 76–80 (1998).

    Article  CAS  PubMed  Google Scholar 

  15. Vajsar, J., Ackerley, C., Chitayat, D. & Becker, L.E. Basal lamina abnormality in the skeletal muscle of Walker-Warburg syndrome. Pediatr. Neurol. 22, 139–143 (2000).

    Article  CAS  PubMed  Google Scholar 

  16. Skorupinska-Tudek, K. et al. Contribution of the mevalonate and methylerythritol phosphate pathways to the biosynthesis of dolichols in plants. J. Biol. Chem. 283, 21024–21035 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Richard, S.B. et al. Structure of 4-diphosphocytidyl-2-C-methylerythritol synthetase involved in mevalonate-independent isoprenoid biosynthesis. Nat. Struct. Biol. 8, 641–648 (2001).

    Article  CAS  PubMed  Google Scholar 

  18. Surmacz, L. & Swiezewska, E. Polyisoprenoids—secondary metabolites or physiologically important superlipids? Biochem. Biophys. Res. Commun. 407, 627–632 (2011).

    Article  CAS  PubMed  Google Scholar 

  19. Kuzuyama, T. Mevalonate and nonmevalonate pathways for the biosynthesis of isoprene units. Biosci. Biotechnol. Biochem. 66, 1619–1627 (2002).

    Article  CAS  PubMed  Google Scholar 

  20. Miziorko, H.M. Enzymes of the mevalonate pathway of isoprenoid biosynthesis. Arch. Biochem. Biophys. 505, 131–143 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Yoshida-Moriguchi, T. et al. O-Mannosyl phosphorylation of α-dystroglycan is required for laminin binding. Science 327, 88–92 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Ervasti, J.M. & Campbell, K.P. Membrane organization of the dystrophin-glycoprotein complex. Cell 66, 1121–1131 (1991).

    Article  CAS  PubMed  Google Scholar 

  23. Duclos, F. et al. Progressive muscular dystrophy in α-sarcoglycan–deficient mice. J. Cell Biol. 142, 1461–1471 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Kunz, S., Sevilla, N., McGavern, D.B., Campbell, K.P. & Oldstone, M.B. Molecular analysis of the interaction of LCMV with its cellular receptor α-dystroglycan. J. Cell Biol. 155, 301–310 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. 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. 264, 2816–2825 (1989).

    CAS  PubMed  Google Scholar 

  26. Manya, H. et al. Demonstration of mammalian protein O-mannosyltransferase activity: coexpression of POMT1 and POMT2 required for enzymatic activity. Proc. Natl. Acad. Sci. USA 101, 500–505 (2004).

    Article  CAS  PubMed  Google Scholar 

  27. de Bernabé, D.B. et al. Loss of α-dystroglycan laminin binding in epithelium-derived cancers is caused by silencing of LARGE. J. Biol. Chem. 284, 11279–11284 (2009).

    Article  PubMed  PubMed Central  Google Scholar 

  28. Davidson, R.L. & Gerald, P.S. Improved techniques for the induction of mammalian cell hybridization by polyethylene glycol. Somatic Cell Genet. 2, 165–176 (1976).

    Article  CAS  PubMed  Google Scholar 

  29. Lee, H., Jen, J.C., Cha, Y.H., Nelson, S.F. & Baloh, R.W. Phenotypic and genetic analysis of a large family with migraine-associated vertigo. Headache 48, 1460–1467 (2008).

    Article  PubMed  Google Scholar 

  30. Lee, H. et al. Improving the efficiency of genomic loci capture using oligonucleotide arrays for high throughput resequencing. BMC Genomics 10, 646 (2009).

    Article  PubMed  PubMed Central  Google Scholar 

  31. Li, H. et al. The Sequence Alignment/Map format and SAMtools. Bioinformatics 25, 2078–2079 (2009).

    Article  PubMed  PubMed Central  Google Scholar 

  32. McKenna, A. et al. The Genome Analysis Toolkit: a MapReduce framework for analyzing next-generation DNA sequencing data. Genome Res. 20, 1297–1303 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. DePristo, M.A. et al. A framework for variation discovery and genotyping using next-generation DNA sequencing data. Nat. Genet. 43, 491–498 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We thank the Gene Transfer Vector Core (University of Iowa, supported by the US National Institutes of Health (NIH) and the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) (P30 DK 54759) for generating adenoviruses, T. Toy at the UCLA Clinical Genomics Center (see URLs) for assisting with construction of the sequencing libraries, S. Feng at the UCLA Broad Stem Cell Research Center (BSCRC) for assisting in running HiSeq2000 and B. Harry at the UCLA Clinical Genomics Center for maintaining the data analysis pipeline. Sequencing and sequence analysis were supported by the Center for Duchenne Muscular Dystrophy and the UCLA Muscular Dystrophy Core Center grant from the NIH (5P30AR05723). This study used fibroblast samples from the National Institute of Neurological Disorders and Stroke (NINDS) Cell Line Repository (see URLs) and the Miami Brain and Tissue Bank for Developmental Disorders, funded by the National Commission for Human Development (NICHD). We thank members of the Campbell laboratory and C.A. Campbell for fruitful discussions, A. Dietz, A. Crimmins, G. Morgensen, J. Eskuri, P. Guicheney and H.v. Bokhoven for technical support and C. Blaumueller for critical reading of the manuscript. This work was supported in part by a Paul D. Wellstone Muscular Dystrophy Cooperative Research Center Grant (1U54NS053672 to K.P.C., K.D.M., S.A.M. and T.W.) and an American Recovery and Reinvestment Act (ARRA) Go Grant (1 RC2 NS069521-01 to K.P.C. and T.W.). The Muscular Dystrophy Campaign Grant to F.M. is also gratefully acknowledged. S.C. and F.M. are investigators of and are supported by the European Framework Programme 7 (FP7) NMD-Chip and Bio-NMD projects. F.M. is supported by the Great Ormond Street Hospital Children's Charity. K.P.C. is an investigator of the Howard Hughes Medical Institute.

Author information

Authors and Affiliations

  1. Department of Molecular Physiology and Biophysics, University of Iowa Roy J and Lucille A Carver College of Medicine, Iowa City, Iowa, USA

    Tobias Willer, Takako Yoshida-Moriguchi, Daniel Beltran Valero de Bernabe, David Venzke & Kevin P Campbell

  2. Department of Neurology, University of Iowa Roy J and Lucille A Carver College of Medicine, Iowa City, Iowa, USA

    Tobias Willer, Takako Yoshida-Moriguchi, Daniel Beltran Valero de Bernabe, David Venzke, Katherine D Mathews & Kevin P Campbell

  3. Department of Internal Medicine, University of Iowa Roy J and Lucille A Carver College of Medicine, Iowa City, Iowa, USA

    Tobias Willer, Takako Yoshida-Moriguchi, Daniel Beltran Valero de Bernabe, David Venzke & Kevin P Campbell

  4. Howard Hughes Medical Institute, University of Iowa Roy J and Lucille A Carver College of Medicine, Iowa City, Iowa, USA

    Tobias Willer, Takako Yoshida-Moriguchi, Daniel Beltran Valero de Bernabe, David Venzke & Kevin P Campbell

  5. Department of Human Genetics, David Geffen School of Medicine, University of California, Los Angeles, California, USA

    Hane Lee & Stanley F Nelson

  6. Department of Pathology and Laboratory Medicine, David Geffen School of Medicine, University of California, Los Angeles, California, USA

    Hane Lee & Stanley F Nelson

  7. Department of Cell Chemistry, Centre for Organismal Studies, University of Heidelberg, Heidelberg, Germany

    Mark Lommel & Sabine Strahl

  8. Division of Neuroscience, Dubowitz Neuromuscular Centre, Institute of Child Health & Great Ormond Street Hospital for Children, London, UK

    Sebahattin Cirak & Francesco Muntoni

  9. Molecular Structure and Function, The Hospital for Sick Children, Toronto, Ontario, Canada

    Harry Schachter

  10. Division of Neurology, The Hospital for Sick Children, Toronto, Ontario, Canada

    Jiri Vajsar

  11. University Pierre et Marie Curie, UM 76, Institut National de la Santé et de la Recherche Médicale (INSERM) U974, Centre Nationale de la Recherche Scientifique (CNRS) Unité Mixte de Recherce (UMR) 7215, Assistance Publoique–Hôpitaux de Paris (AP-HP), Institute of Myology, Paris, France

    Thomas Voit

  12. White-Wilson Medical Center, Fort Walton Beach, Florida, USA

    Andrea S Loder

  13. Department of Pediatrics, University of Chicago, Chicago, Illinois, USA

    William B Dobyns

  14. PreventionGenetics, Marshfield, Wisconsin, USA

    Thomas L Winder

  15. Department of Pediatrics, University of Iowa Roy J and Lucille A Carver College of Medicine, Iowa City, Iowa, USA

    Katherine D Mathews

  16. Department of Pathology, University of Iowa Roy J and Lucille A Carver College of Medicine, Iowa City, Iowa, USA

    Steven A Moore

Authors
  1. Tobias Willer
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Hane Lee
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Mark Lommel
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Takako Yoshida-Moriguchi
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Daniel Beltran Valero de Bernabe
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. David Venzke
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Sebahattin Cirak
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Harry Schachter
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Jiri Vajsar
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Thomas Voit
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Francesco Muntoni
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Andrea S Loder
    View author publications

    You can also search for this author in PubMed Google Scholar

  13. William B Dobyns
    View author publications

    You can also search for this author in PubMed Google Scholar

  14. Thomas L Winder
    View author publications

    You can also search for this author in PubMed Google Scholar

  15. Sabine Strahl
    View author publications

    You can also search for this author in PubMed Google Scholar

  16. Katherine D Mathews
    View author publications

    You can also search for this author in PubMed Google Scholar

  17. Stanley F Nelson
    View author publications

    You can also search for this author in PubMed Google Scholar

  18. Steven A Moore
    View author publications

    You can also search for this author in PubMed Google Scholar

  19. Kevin P Campbell
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

T.W., H.L., S.F.N. and K.P.C. designed the research. T.W. performed the research and analyzed the data. H.L. and S.F.N. performed SNP analysis, next-generation sequencing and data filtering. M.L. and S.S. carried out POMT enzyme activity assays. T.Y.-M. performed α-dystroglycan orthophosphate cell labeling experiments. D.B.V.d.B. performed qRT-PCR expression analysis. D.V. carried out antibody affinity purification and labeling. T.L.W. carried out Sanger sequencing of known WWS-associated genes. S.A.M. performed muscle histology and clinical data interpretation. H.S., J.V., S.C., F.M., T.V., A.S.L., W.B.D. and K.D.M. provided clinical data and/or fibroblast samples from individuals with WWS. K.P.C. supervised and mentored the project. T.W. and K.P.C. wrote the initial manuscript, and all authors approved and commented on the manuscript.

Corresponding author

Correspondence to Kevin P Campbell.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–8 and Supplementary Tables 1–6 (PDF 851 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Willer, T., Lee, H., Lommel, M. et al. ISPD loss-of-function mutations disrupt dystroglycan O-mannosylation and cause Walker-Warburg syndrome. Nat Genet 44, 575–580 (2012). https://doi.org/10.1038/ng.2252

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/ng.2252

This article is cited by

Search

Advanced 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