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SHANK3 and IGF1 restore synaptic deficits in neurons from 22q13 deletion syndrome patients

Abstract

Phelan–McDermid syndrome (PMDS) is a complex neurodevelopmental disorder characterized by global developmental delay, severely impaired speech, intellectual disability, and an increased risk of autism spectrum disorders (ASDs)1. PMDS is caused by heterozygous deletions of chromosome 22q13.3. Among the genes in the deleted region is SHANK3, which encodes a protein in the postsynaptic density (PSD)2,3. Rare mutations in SHANK3 have been associated with idiopathic ASDs4,5,6,7, non-syndromic intellectual disability8, and schizophrenia9. Although SHANK3 is considered to be the most likely candidate gene for the neurological abnormalities in PMDS patients10, the cellular and molecular phenotypes associated with this syndrome in human neurons are unknown. We generated induced pluripotent stem (iPS) cells from individuals with PMDS and autism and used them to produce functional neurons. We show that PMDS neurons have reduced SHANK3 expression and major defects in excitatory, but not inhibitory, synaptic transmission. Excitatory synaptic transmission in PMDS neurons can be corrected by restoring SHANK3 expression or by treating neurons with insulin-like growth factor 1 (IGF1). IGF1 treatment promotes formation of mature excitatory synapses that lack SHANK3 but contain PSD95 and N-methyl-d-aspartate (NMDA) receptors with fast deactivation kinetics. Our findings provide direct evidence for a disruption in the ratio of cellular excitation and inhibition in PMDS neurons, and point to a molecular pathway that can be recruited to restore it.

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Figure 1: CaMKIIα-GFP labels functional iPS cell-derived forebrain neurons that express SHANK3.
Figure 2: PMDS neurons display impaired excitatory synaptic transmission.
Figure 3: PMDS neurons show reduced expression of glutamate receptors and decreased number of synapses.
Figure 4: Reduced SHANK3 expression contributes to synaptic defects in PMDS neurons.
Figure 5: IGF1 treatment restores excitatory synaptic transmission in PMDS neurons.

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Acknowledgements

We are grateful to participants and their families for their support; M. Adam for assistance with recruitment; to X. Jia, A. Cherry, C. Bangs, P. Jones, and J. Williams for assistance with tissue culture; P. Liao for help with multiplex ligation-dependent probe amplification (MLPA); M. Fabian for astrocyte preparations; H.N. Nguyen for consultations on the neural differentiation protocol and spectral karyotyping (SKY); V. Vu and G. Lin for help with data analysis; T. Sudhof, T. Boeckers, A. Grabruker, C. Garner and C. Sala for antibodies; R. Xavier for SHANK3 complementary DNA; R. Reijo-Pera and members of the Dolmetsch laboratory for commenting on the manuscript; E. Nigh for editing the manuscript. We also thank the Stanford Neuroscience Microscopy Service (supported by National Institutes of Health (NIH) NS069375). Support for this study came from the California Institute for Regenerative Medicine CIRM, the Autism Science Foundation and the Phelan-McDermid Syndrome Foundation (to A.S.), the Swiss National Science Foundation (to T.P.), the Japan Society for the Promotion of Research Abroad and American Heart Association (to M.Y.), the National Institute of Mental Health (NIMH) grant R33MH087898 (to J.F.H.); NIH Pioneer Award (5DP1OD3889), CIRM (grant RT2-01906) and Simons Foundation (to R.E.D.). We are also grateful for funding from the JDH research fund, N. Juaw, B. and F. Horowitz, M. McCafferey, B. and J. Packard, P. Kwan and K. Wang, and the Flora foundation.

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Authors and Affiliations

Authors

Contributions

A.S. and R.E.D. designed experiments and wrote the manuscript; A.S. performed iPS cell maintenance, neural differentiation, electrophysiology, cloning and immunocytochemistry; O.S. maintained and characterized iPS cells, performed western blot and qRT–PCR; M.Y. generated and characterized iPS cells; T.P. performed multiplex single-cell qRT–PCR; V.S. performed teratoma assay; R.S. and A.K. performed qRT–PCR and data analysis; W.F., J.A.B. and J.F.H. recruited and characterized patients and performed the MLPA assay.

Corresponding author

Correspondence to Ricardo E. Dolmetsch.

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The authors declare no competing financial interests.

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This file contains Supplementary Figures 1-16 Supplementary Tables 1-5 and additional references. (PDF 9072 kb)

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Shcheglovitov, A., Shcheglovitova, O., Yazawa, M. et al. SHANK3 and IGF1 restore synaptic deficits in neurons from 22q13 deletion syndrome patients. Nature 503, 267–271 (2013). https://doi.org/10.1038/nature12618

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