Modeling Smith-Lemli-Opitz syndrome with induced pluripotent stem cells reveals a causal role for Wnt/β-catenin defects in neuronal cholesterol synthesis phenotypes

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Smith-Lemli-Opitz syndrome (SLOS) is a malformation disorder caused by mutations in DHCR7, which impair the reduction of 7-dehydrocholesterol (7DHC) to cholesterol. SLOS results in cognitive impairment, behavioral abnormalities and nervous system defects, though neither affected cell types nor impaired signaling pathways are fully understood. Whether 7DHC accumulation or cholesterol loss is primarily responsible for disease pathogenesis is also unclear. Using induced pluripotent stem cells (iPSCs) from subjects with SLOS, we identified cellular defects that lead to precocious neuronal specification within SLOS derived neural progenitors. We also demonstrated that 7DHC accumulation, not cholesterol deficiency, is critical for SLOS-associated defects. We further identified downregulation of Wnt/β-catenin signaling as a key initiator of aberrant SLOS iPSC differentiation through the direct inhibitory effects of 7DHC on the formation of an active Wnt receptor complex. Activation of canonical Wnt signaling prevented the neural phenotypes observed in SLOS iPSCs, suggesting that Wnt signaling may be a promising therapeutic target for SLOS.

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This work was supported by the intramural research programs of the Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD) and the National Human Genome Research Institute (NHGRI), a pilot award for iPSC research from the NIH Center for Regenerative Medicine, and NIH grant GM110128 (W.C.). We thank K. Perez, N. Khezri and G.F. Rodriguez for assistance with hESC and iPSC culture. We also thank C. Toth and A. Ely for assistance with mice and sequencing. We thank A. Dutra, E. Pak and the NHGRI Cytogenetics Core for assistance with hESC and iPSC karyotyping. We thank L. (Chip) Dye and the NICHD Microscopy and Imaging Core for electron microscopy assistance. We thank B.S. Mallon (National Institute of Neurological Disorders and Stroke, NIH Stem Cell Unit), S. Kuznetsov (National Institute of Dental and Craniofacial Research, NIDCR), and P.G. Robey (NIDCR, NIH Stem Cell Unit) for experimental advice and the i19 and i21 human iPSC lines. We thank G. Mostoslavksy (Boston University) for the kind gift of the STEMCCA plasmid. We thank D. Kotton (Boston University) for the kind gift of the Cre-IRES-PuroR plasmid. We thank F. Zhang (Massachusetts Institute of Technology) for the gift of the pX330-U6-Chimeric_BB-CBh-hSpCas9 plasmid. We thank J. Mills (Children's Hospital of Philadelphia) for experimental advice, the National Heart, Lung and Blood Institute iPSC Core Facility for assistance with lathosterolosis iPSC generation, and M. Rao and K. Pfeifer for critical discussions. Finally, we would like to thank the patients and guardians who participated in the NIH clinical program and contributed to this study.

Author information


  1. Program in Genomics of Differentiation, The Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health (NIH), Bethesda, Maryland, USA.

    • Kevin R Francis
    • , Amy N Ton
    • , Peter E O'Halloran
    • , Nasir Malik
    •  & Heiner Westphal
  2. Program in Developmental Endocrinology and Genetics, The Eunice Kennedy Shriver National Institute of Child Health and Human Development, NIH, Bethesda, Maryland, USA.

    • Kevin R Francis
    • , Christopher A Wassif
    • , Ian M Williams
    • , Celine V Cluzeau
    •  & Forbes D Porter
  3. Department of Chemistry, University of Illinois at Chicago, Chicago, Illinois, USA.

    • Yao Xin
    •  & Wonhwa Cho
  4. Computational and Statistical Genomics Branch, National Human Genome Research Institute, NIH, Bethesda, Maryland, USA.

    • Niraj S Trivedi
  5. Genetic Diseases Research Branch, National Human Genome Research Institute, NIH, Bethesda, Maryland, USA.

    • William J Pavan


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K.R.F. and F.D.P. designed and directed the study, analyzed data and wrote the manuscript. K.R.F., A.N.T., P.E.O'H., N.M., I.M.W. and C.V.C. performed experiments and analyzed data. H.W. contributed to study design. C.A.W. provided assistance with GC/MS and study design. N.S.T. and W.J.P. provided support and performed statistical analysis of microarray experiments. Y.X. and W.C. designed, carried out and analyzed experiments on lipid-protein interactions.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Kevin R Francis or Forbes D Porter.

Supplementary information

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    Supplementary Text and Figures

    Supplementary Figures 1–10 and Supplementary Tables 1–3

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    Supplementary Table 4

    Gene network analysis of microarray data identify highly correlated signaling pathways associated with sterol perturbation and pluripotent signaling.


  1. 1.

    Beating areas generated from the CWI 4F-2 SLOS iPS line. Areas of cardiomyocyte formation and rhythmic beating were observed during germ layer differentiation assays. Beating area corresponds to immunocytochemical staining positive for NKX2.5 (immature cardiomyocyte marker) in Supplementary Figure 1h.