Long noncoding RNAs (lncRNAs) have been shown to act as important cell biological regulators including cell fate decisions but are often ignored in human genetics. Combining differential lncRNA expression during neuronal lineage induction with copy number variation morbidity maps of a cohort of children with autism spectrum disorder/intellectual disability versus healthy controls revealed focal genomic mutations affecting several lncRNA candidate loci. Here we find that a t(5:12) chromosomal translocation in a family manifesting neurodevelopmental symptoms disrupts specifically lnc-NR2F1. We further show that lnc-NR2F1 is an evolutionarily conserved lncRNA functionally enhances induced neuronal cell maturation and directly occupies and regulates transcription of neuronal genes including autism-associated genes. Thus, integrating human genetics and functional testing in neuronal lineage induction is a promising approach for discovering candidate lncRNAs involved in neurodevelopmental diseases.

5Long noncoding RNAs (lncRNAs) have been shown to act as important cell biological regulators including cell fate decisions but are often ignored in human genetics. Combining differential lncRNA expression during neuronal lineage induction with copy number variation morbidity maps of a cohort of children with autism spectrum disorder/intellectual disability versus healthy controls revealed focal genomic mutations 5 0 affecting several lncRNA candidate loci. Here we find that a t(5:12) chromosomal translocation in a family manifesting neurodevelopmental symptoms disrupts specifically lnc-NR2F1. We further show that lnc-NR2F1 is an evolutionarily conserved lncRNA functionally enhances induced neuronal cell maturation and directly occupies and regulates transcription of neuronal genes including autism-associated genes. Thus, 5 5integrating human genetics and functional testing in neuronal lineage induction is a promising approach for discovering candidate lncRNAs involved in neurodevelopmental diseases. (129 words)3 Eukaryotic genomes are extensively transcribed to produce long non-coding 6 0 RNAs (lncRNAs) in a temporally and spatially regulated manner 1 . Until recently, lncRNAs were often dismissed as lacking functional relevance. However, lncRNAs are emerging as critical regulators of diverse biological processes and have been increasingly associated with a wide range of diseases, based primarily on dysregulated expression 2 .LncRNAs represent a new layer of complexity in the molecular architecture of the 6 5 genome, and strategies to validate disease relevant lncRNAs are much needed. Highthroughput analyses have shown that lncRNAs are widely expressed in the brain and may contribute to complex neurodevelopmental processes 2-9 . However, few studies have examined the role of lncRNAs in brain development mostly due to technical difficulties.Direct lineage conversion by the transcription factors Brn2, Ascl1 and Myt1l (termed 7 0 BAM factors in combination) into induced neuronal (iN) cells, recapitulates significant events controlling neurogenesis programs 10-12 , and therefore, it is a facile and informative system to study the role of lncRNAs in the establishment of neuronal identity.The noncoding genome has emerged as a major source for human diversity and disease origins. Given that less than 2% of the genome encodes protein-coding genes, the 7 5 majority of the genomic landscape is largely encompassed by non-coding elements.Efforts to identify genetic variation linked to human disease through genome-wide association studies revealed a significant majority affecting the non-coding landscape.Based on their expression and diversity in the mammalian brain, we postulate neuronal lncRNAs may be recurrently affected by mutations that disrupt normal brain function. 0Neurodevelopmental disorders manifest as a spectrum of phenotypes particularly early in life 13 . Recent studies suggest that this diversity is the result of different combinations of mutations in multiple genes, often impacting key pathways such as synap...

Dendritic spine morphology and dendritic arborization are key determinants of neuronal connectivity and play critical roles in learning, memory and behavior function. Recently, defects of ZBTB20, a BTB and zinc finger domain containing transcriptional repressor, have been implicated in a wide range of neurodevelopmental disorders, including intellectual disability and autism. Here we show distinct effects of expression of two major isoforms, long and short, of ZBTB20, and its neurodevelopmental disorder-linked variants, on dendritic architecture of cultured rat cortical pyramidal neurons. The N-terminal of ZBTB20 showed a role in regulating dendritic spine morphology. Two ZBTB20 single nucleotide variants, located at the N-terminal and central regions of the protein and potentially conferring autism risk, altered dendritic spine morphology. In contrast, a single nucleotide variant identified in patients with intellectual disability and located at the C-terminus of ZBTB20 affected dendritic arborization and dendritic length but had no effect on dendritic spine morphology. Furthermore, truncation of the extreme C-terminus of ZBTB20 caused spine and dendritic morphological changes that were similar but distinct from those caused by the C-terminal variant. Taken together, our study suggests ZBTB20’s role in dendritic and synaptic structure and provide possible mechanisms of its effect in neurodevelopmental disorders.

Deletions and mutations involving the Retinoic Acid Induced 1 (RAI1) gene at 17p11.2 cause Smith-Magenis syndrome (SMS). Here we report a patient with autism as the main clinical presentation, with some SMS-like features and a rare de novo RAI1 gene mutation, c.3440G > A (p.R1147Q). We functionally characterized the RAI1 p.R1147Q mutant protein. The mutation, located near the nuclear localization signal, had no effect on the subcellular localization of the mutant protein. However, similar to previously reported RAI1 missense mutations in SMS patients, the RAI1 p.R1147Q mutant protein showed a significant deficiency in activating in vivo transcription of a reporter gene driven by a BDNF (brain-derived neurotrophic factor) intronic enhancer. In addition, expression of other genes associated with neurobehavioral abnormalities and/or neurodevelopmental disorders were found to be altered in this patient. These results suggest a likely contribution of RAI1, either alone or in combination of other factors, to social behavior and reinforce the RAI1 gene as a candidate gene in patients with autistic manifestations or social behavioral abnormalities.

Chromosomal imbalances are a major cause of intellectual disability (ID) and multiple congenital anomalies. We have clinically and molecularly characterized two patients with chromosome translocations and ID. Using whole genome array CGH analysis, we identified a microdeletion involving 4q21.3, unrelated to the translocations in both patients. We confirmed the 4q21.3 microdeletions using fluorescence in situ hybridization and quantitative genomic PCR. The corresponding deletion boundaries in the patients were further mapped and compared to previously reported 4q21 deletions and the associated clinical features. We determined a common region of deletion overlap that appears unique to ID, short stature, hypotonia, and dysmorphic facial features.