Delineating the psychiatric and behavioral phenotype of recurrent 2q13 deletions and duplications

Recurrent deletions and duplications at the 2q13 locus have been associated with developmental delay (DD) and dysmorphisms. We aimed to undertake detailed clinical characterization of individuals with 2q13 copy number variations (CNVs), with a focus on behavioral and psychiatric phenotypes. Participants were recruited via the Unique chromosomal disorder support group, U.K. National Health Service Regional Genetics Centres, and the DatabasE of genomiC varIation and Phenotype in Humans using Ensembl Resources (DECIPHER) database. A review of published 2q13 patient case reports was undertaken to enable combined phenotypic analysis. We present a new case series of 2q13 CNV carriers (21 deletion, 4 duplication) and the largest ever combined analysis with data from published studies, making a total of 54 deletion and 23 duplication carriers. DD/intellectual disabilities was identified in the majority of carriers (79% deletion, 70% duplication), although in the new cases 52% had an IQ in the borderline or normal range. Despite the median age of the new cases being only 9 years, 64% had a clinical psychiatric diagnosis. Combined analysis found attention deficit hyperactivity disorder (ADHD) to be the most frequent diagnosis (48% deletion, 60% duplication), followed by autism spectrum disorders (33% deletion, 17% duplication). Aggressive (33%) and self-injurious behaviors (33%) were also identified in the new cases. CNVs at 2q13 are typically associated with DD with mildly impaired intelligence, and a high rate of childhood psychiatric diagnosesparticularly ADHD. We have further characterized the clinical phenotype related to imbalances of the 2q13 region and identified it as a region of interest for the neurobiological investigation of ADHD

MYT1L mutations cause intellectual disability and variable obesity by dysregulating gene expression and development of the neuroendocrine hypothalamus

Deletions at chromosome 2p25.3 are associated with a syndrome consisting of intellectual disability and obesity. The smallest region of overlap for deletions at 2p25.3 contains PXDN and MYT1L. MYT1L is expressed only within the brain in humans. We hypothesized that single nucleotide variants (SNVs) in MYT1L would cause a phenotype resembling deletion at 2p25.3. To examine this we sought MYT1L SNVs in exome sequencing data from 4, 296 parent-child trios. Further variants were identified through a genematcher-facilitated collaboration. We report 9 patients with MYT1L SNVs (4 loss of function and 5 missense). The phenotype of SNV carriers overlapped with that of 2p25.3 deletion carriers. To identify the transcriptomic consequences of MYT1L loss of function we used CRISPR-Cas9 to create a knockout cell line. Gene Ontology analysis in knockout cells demonstrated altered expression of genes that regulate gene expression and that are localized to the nucleus. These differentially expressed genes were enriched for OMIM disease ontology terms “mental retardation”. To study the developmental effects of MYT1L loss of function we created a zebrafish knockdown using morpholinos. Knockdown zebrafish manifested loss of oxytocin expression in the preoptic neuroendocrine area. This study demonstrates that MYT1L variants are associated with syndromic obesity in humans. The mechanism is related to dysregulated expression of neurodevelopmental genes and altered development of the neuroendocrine hypothalamus

Hypothalamic peptide expression in zebrafish knockdown for <i>MYT1L</i> orthologues.

<p>(A)Whole mount in situ hybridization demonstrating that <i>MYT1L</i> orthologs <i>myt1la</i> and <i>myt1lb</i> are expressed throughout the zebrafish central nervous system. T = telencephalon, te = tectum, hy = hypothalamus, h = hindbrain. (B)Whole mount in situ hybridization demonstrating loss of <i>myt1la</i> expression in <i>arnt2</i> mutant zebrafish, top panel shows control fish and bottom panel <i>arnt2</i> mutant fish. The embryos are heavily over-stained to show the low-level expression of <i>myt1la</i> in the ventral diencephalon. The arrow indicates the region of the neuroendocrine preoptic area where oxytocin expressing neurons are located. (C)Whole mount in situ hybridization demonstrating that knockdown of <i>myt1la</i> and <i>myt1lb</i>, alone or in combination, results in loss of oxytocin expression in the neuroendocrine preoptic area. Arrows indicate neuroendocrine preoptic area. (D)Bar chart quantifying loss of oxytocin expression in neuroendocrine preoptic area. WISH for oxytocin was quantified as follows: ~30 cells = wild type expression, 5–15 cells = reduced, 1–4 = highly reduced, 0 = no expression (E)Whole mount in situ hybridization demonstrating that knockdown of <i>myt1la/b</i> results in loss of arginine vasopressin expression in the neuroendocrine preoptic region (indicated by arrow) but not the ventral hypothalamus (indicated by arrowhead). (F) Bar chart quantifying loss of arginine vasopression expression in <i>myt1la/b</i> morphants. WISH for arginine vasopressin was quantified as follows: ~30 cells = wild type expression, 5–15 cells = reduced, 1–4 = highly reduced, 0 = no expression.</p

Gene expression profiling of <i>MYT1L</i> knockout cell line.

<p>(A)Enrichment analysis demonstrates enrichment for Gene Ontology Biological Process 2015 term gene expression (GO:0010467, adjusted p-value 0.00077, Z-score -2.34, combined score 16.77). (B)Enrichment analysis demonstrates enrichment for Gene Ontology Cellular Component 2015 terms nucleolus (GO: 0005730, adjusted p-value 0.0023, Z-score -2.21, combined score 13.43) and nucleoplasm (GO: 0005654, adjusted p-value 0.005853, Z-scre -2.08, combined score 11.4). (C)Enrichment analysis demonstrates enrichment for Reactome 2016 pathways Gene Expression_Homo Sapiens_R-HAS-74160 (adjusted p-value 2.2 x 10–7, Z-score -2.16, combined score 33) and Generic Transcription Pathway_Homo Sapiens_R-HAS-212436 (adjusted p-value 0.01586, Z-score -2.26, combined score 9.35). (D)Enrichment analysis demonstrates enrichment for OMIM disease term mental retardation (p-value 0.045, adjusted p-value 0.38, Z-score -1.32, combined score 1.27).</p

Clinical characterization of <i>MYT1L</i> variant carriers.

<p>The heatmap depicts the frequency of various clinical features in <i>MYT1L</i> variant carriers.</p

Summary of demographic features, developmental milestones, medical complications and exome sequencing of cohort.

<p>Summary of demographic features, developmental milestones, medical complications and exome sequencing of cohort.</p

Structural effects of <i>MYT1L</i> missense variants.

<p>(A)Schematic diagram of MYT1L protein, the yellow boxes indicate zinc finger domains. The missense variants are indicated by arrows. (B)Model of the 2^nd and 3^rd zinc fingers of MYT1L bound to DNA. This is based upon the structure of the 4^th and 5^th zinc fingers of MYT1 (protein data bank file 2mf8), which have high sequence similarity to the 2^nd and 3^rd zinc fingers of MYT1L. The second zinc finger is in magenta, and the third finger in green. The beige spheres represent the zinc ions, with the CCHC zinc ligands shown in cyan. Replacement of L520 by proline is expected to disrupt the structure of the protein by preventing the formation of a tight turn. H524 and G527 are a zinc ligands, so replacement will also disrupt the structure. H560 and R569 are conserved residues directly involved in DNA binding. Image created using Pymol.</p