Complete Disruption of Autism-Susceptibility Genes by Gene Editing Predominantly Reduces Functional Connectivity of Isogenic Human Neurons

Deneault, Éric; White, Stuart C.; Rodrigues, Deivid C.; Ross, P. Joel; Faheem, Muhammad; Zaslavsky, Kirill; Wang, Zhuozhi; Alexandrova, Roumiana; Pellecchia, Giovanna; Wei, Wei; Piekna, Alina; Kaur, Gaganjot; Howe, Jennifer; Kwan, Vickie; Thiruvahindrapuram, Bhooma; Walker, Susan; Lionel, Anath C.; Pasceri, Peter; Merico, Daniele; Yuen, Ryan K.C.; Singh, Karun K.; Ellis, James; Scherer, Stephen W.

doi:10.1016/j.stemcr.2019.01.008

Cited by 42 publications

(93 citation statements)

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“…Indeed, these Drosophila and C. elegans results likely reflect a behavioral outcome of circuit-level hyperexcitability recently discovered in several human iPSC derived neuronal culture models of a number of monogenic ASD risk factors. 68,139 The results also provide a plausible explanation for inconsistent reports of impaired habituation in humans, which variably employ diverse response metrics most often without genetic stratification of patient populations. 69 While these shared phenotypes are exciting, our results also reveal a remarkable diversity and modularity in phenotypic disruptions, and thus clearly indicate that single phenotype in vivo functional validation and characterization efforts will be insufficient to capture the complex multi-facetted phenotypic disruptions that stem from inactivating mutations in ASD-associated genes.…”

Section: Discussionmentioning

confidence: 78%

“…Rapid advances in gene discovery and orthology prediction have altered the gene lists used during the course of this project, a challenge facing similar recent systematic investigations of ASD-associated gene function. 67,68 Despite these shifts, our list still covers 82% (14/17) of the most strongly associated genes from Satterstrom et al and 87% (13/15) of the SFARI Gene high confidence category listed genes (Table S1) with a viable ortholog deletion or severe missense allele available. This led to a mix of currently defined high-and mid-confidence ASDassociated genes, giving us a unique opportunity to study putative ASD-associated genes of completely unknown function alongside well established genes with known roles in neurodevelopment and sensory processing (See Table S1 for a complete listing of characterized strains, orthology relationships, and ASD-association confidence).…”

Section: Asd-associated Genes Are Highly Conserved To C Elegansmentioning

confidence: 99%

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Systematic phenomics analysis of ASD-associated genes reveals shared functions and parallel networks underlying reversible impairments in habituation learning

McDiarmid

Belmadani

Liang

et al. 2019

Preprint

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A major challenge facing the genetics of Autism Spectrum Disorders (ASD) is the large and growing number of candidate risk genes and gene variants of unknown functional significance.Here, we used Caenorhabditis elegans to systematically functionally characterize ASDassociated genes in vivo. Using our custom machine vision system we quantified 26 phenotypes spanning morphology, locomotion, tactile sensitivity, and habituation learning in 87 strains each carrying a mutation in an ortholog of an ASD-associated gene. We identified hundreds of novel genotype-phenotype relationships ranging from severe developmental delays and uncoordinated movement to subtle deficits in sensory and learning behaviors. We clustered genes by similarity in phenomic profiles and used epistasis analysis to discover parallel networks centered on CHD8•chd-7 and NLGN3•nlg-1 that underlie mechanosensory hyper-responsivity and impaired habituation learning. We then leveraged our data for in vivo functional assays to gauge missense variant effect. Expression of wild-type NLG-1 in nlg-1 mutant C. elegans rescued their sensory and learning impairments. Testing the rescuing ability of all conserved ASD-associated neuroligin variants revealed varied partial loss-of-function despite proper subcellular localization. Finally, we used CRISPR-Cas9 auxin inducible degradation to determine that phenotypic abnormalities caused by developmental loss of NLG-1 can be reversed by adult expression. This work charts the phenotypic landscape of ASDassociated genes, offers novel in vivo variant functional assays, and potential therapeutic targets for ASD.

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Section: Discussionmentioning

confidence: 78%

Section: Asd-associated Genes Are Highly Conserved To C Elegansmentioning

confidence: 99%

Systematic phenomics analysis of ASD-associated genes reveals shared functions and parallel networks underlying reversible impairments in habituation learning

McDiarmid

Belmadani

Liang

et al. 2019

Preprint

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“…and ASTN2 (Deneault et al, 2018). Another study detected reduced neuronal firing in iPSCderived neurons of eight individuals with idiopathic ASD (Marchetto et al, 2017) and detected a correlation of reduced network complexity to behavioral and cognitive phenotype of the donors (Amatya et al, 2019).…”

Section: Cask-related Disorders Have Emerged As An Important Geneticmentioning

confidence: 94%

Presynaptic dysfunction inCASK-related neurodevelopmental disorders

Becker

Mastropasqua

Reising

et al. 2019

Preprint

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HighlightsModelling of CASK-related disorders using iPSC-derived human neuronal cells CASK mutations cause dysregulation of its protein interactor partners Reduced CASK levels primarily affect inhibitory presynapse development In vitro GABAergic phenotype predicts in vivo neurotransmitter levels Summary (150 words)CASK-related disorders are a genetically defined group of neurodevelopmental syndromes.There is limited information about the effects of CASK mutations in human neurons. Therefore, we sought to delineate CASK mutation consequences and neuronal level effects using induced pluripotent stem cell-derived neurons from two mutation carriers; one male diagnosed with ASD and a female with MICPCH. We show a reduction of the CASK protein in maturing neurons from the mutation carriers, which leads to significant downregulation of gene sets involved in presynaptic development and CASK protein interactors. Furthermore, CASKdeficient neurons showed decreased inhibitory presynapse size as indicated by VGAT staining, which may alter the excitatory-inhibitory (E/I) balance in developing neural circuitries. Using in vivo magnetic resonance spectroscopy quantification of GABA in the male mutation carrier, we further highlight the possibility to validate in vitro cellular data in brain. Our data shows that future pharmacological and clinical studies on targeting presynapses and E/I imbalance could lead to specific treatments for CASK-related disorders.

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“…We also included genes that are direct targets of FMRP [64]. For the gene sets from other iPSC transcriptome studies, we curated previously described differentially expressed genes caused by: i) shRNA KD of SHANK3 in hiPSC-derived neurons [65]; ii) CRISPR/Cas9 heterozygous KO of CHD8 in hiPSC-derived NPCs and neurons [66]; iii) shRNA KD of TCF4 and EHMT1 in hiPSC-derived NPCs [67]; iv) shRNA KD of MBD5 and SATB2 in human neural stem cells [68]; v) shRNA KD of NRXN1 in human neural stem cells [69]; and vi) CRISPR/Cas9 heterozygous and homozygous KO of ten different ASD-related genes in iPSCs and iPSC-derived neurons [70]. Full gene lists are provided in Supplemental Table 4 .…”

Section: Methodsmentioning

confidence: 99%

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

Breen

Browne

Hoffman

et al. 2019

Preprint

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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 ...

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Complete Disruption of Autism-Susceptibility Genes by Gene Editing Predominantly Reduces Functional Connectivity of Isogenic Human Neurons

Cited by 42 publications

References 0 publications

Systematic phenomics analysis of ASD-associated genes reveals shared functions and parallel networks underlying reversible impairments in habituation learning

Systematic phenomics analysis of ASD-associated genes reveals shared functions and parallel networks underlying reversible impairments in habituation learning

Presynaptic dysfunction inCASK-related neurodevelopmental disorders

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

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