Transcriptional signatures of participant-derived neural progenitor cells and neurons implicate altered Wnt signaling in Phelan McDermid syndrome and autism

Abstract

28Background: Phelan-McDermid syndrome (PMS) is a rare genetic disorder with high risk of 29 autism spectrum disorder (ASD), intellectual disability and language delay, and is caused by 30 22q13.3 deletions or mutations in the SHANK3 gene. To date, the molecular and pathway changes 31 resulting from SHANK3 haploinsufficiency in PMS remain poorly understood. Uncovering these 32 mechanisms is critical for understanding pathobiology of PMS and, ultimately, for the 33 development of new therapeutic interventions. 34 35 Methods: We developed human induced pluripotent stem cell (hiPSC)-based models of PMS by 36reprogramming peripheral blood samples from individuals with PMS (n=7) and their unaffected 37 siblings (n=6). For each participant, up to three hiPSC clones were generated and differentiated 38 into induced neural progenitor cells (iNPCs; n=32) and induced forebrain neurons (iNeurons; 39 n=42). Genome-wide RNA-sequencing was applied to explore transcriptional differences between 40 PMS probands and unaffected siblings. 41 42 Results: Transcriptome analyses identified 391 differentially expressed genes (DEGs) in iNPCs 43 and 82 DEGs in iNeurons, when comparing cells from PMS probands and unaffected siblings 44 (FDR <5%). Genes under-expressed in PMS were implicated in Wnt signaling, embryonic 45 development and protein translation, while over-expressed genes were enriched for pre-and post-46 synaptic density genes, regulation of synaptic plasticity, and G-protein-gated potassium channel 47 activity. Gene co-expression network analysis identified two modules in iNeurons that were over-48 expressed in PMS, implicating postsynaptic signaling and GDP binding, and both modules 49 harbored a significant enrichment of genetic risk loci for developmental delay and intellectual 50 3 disability. Finally, PMS-associated genes were integrated with other ASD iPSC transcriptome 51 findings and several points of convergence were identified, indicating altered Wnt signaling, 52 extracellular matrix and glutamatergic synapses. 53 54 Limitations: Given the rarity of the condition, we could not carry out experimental validation in 55 independent biological samples. In addition, functional and morphological phenotypes caused by 56 loss of SHANK3 were not characterized here. 57 58 Conclusions: This is the largest human neural sample analyzed in PMS. Genome-wide RNA-59 sequencing in hiPSC-derived neural cells from individuals with PMS revealed both shared and 60 distinct transcriptional signatures across iNPCs and iNeurons, including many genes implicated in 61 risk for ASD, as well as specific neurobiological pathways, including the Wnt pathway. 62 63 64 Phelan-McDermid syndrome (PMS) is one of the most penetrant and more common single-locus 66 causes of ASD, accounting for ca. 1% of ASD diagnoses [1-3]. PMS is caused by heterozygous 67 22q13.3 deletions or mutations leading to haploinsufficiency of the SHANK3 gene [2, 4-6]. 68 Clinical manifestations of PMS include ASD, global developmental delay, severe to profound 69 intellectual ...