Are CUL3 variants an underreported cause of congenital heart disease?

Complex heart defects (CHD) are a common malformation associated with disruption of developmental pathways. The Cullin-RING ligases (CRLs) are multi-subunit E3 ubiquitin ligases in which Cullin 3 (CUL3) serves as a scaffolding subunit. Heterozygous CUL3 variants have been associated with neurodevelopmental disorders and pseudohypoaldosteronism type IIE. We report a fetus with CHD and a de novo CUL3 variant (NM_003590.4:c.[1549_1552del];[=], p.(Ser517Profs*23)) and review CUL3 variants reported with CHD. We postulate that CUL3 variants predispose to CHD and hypothesize mechanisms of pathogenesis.

Generation of tandem alternative splice acceptor sites and CLTC haploinsufficiency: A cause of CLTC-related disorder.

Tandem splice acceptors (NAGNn AG) are a common mechanism of alternative splicing, but variants that are likely to generate or to disrupt tandem splice sites have rarely been reported as disease causing. We identify a pathogenic intron 23 CLTC variant (NM_004859.4:c.[3766-13_3766-5del];[=]) in a propositus with intellectual disability and behavioral problems. By RNAseq analysis of peripheral blood mRNA, this variant generates transcripts using cryptic proximal splice acceptors (NM_004859.4: r.3765_3766insTTCACAGAAAGGAACTAG, and NM_004859.4:r.3765_3766insAAAGGAACTAG). Given that the propositus expresses 38% the level of CLTC transcripts as unaffected controls, these variant transcripts, which encode premature termination codons, likely undergo nonsense mediated mRNA decay (NMD). This is the first functional evidence for CLTC haploinsufficiency as a cause of CLTC-related disorder and the first evidence that the generation of tandem alternative splice sites causes CLTC-related disorder. We suggest that variants creating tandem alternative splice sites are an underreported disease mechanism and that transcriptome-level analysis should be routinely pursued to define the pathogenicity of such variants.

Do PACS1 variants impeding adaptor protein binding predispose to syndromic intellectual disability?

To date, PACS1-neurodevelopmental disorder (PACS1-NDD) has been associated with recurrent variation of Arg203 and is considered diagnostic of PACS1-NDD, an autosomal dominant syndromic intellectual disability disorder. Although incompletely defined, the proposed disease mechanism for this variant is altered PACS1 affinity for its client proteins. Given this proposed mechanism, we hypothesized that PACS1 variants that interfere with binding of adaptor proteins might also give rise to syndromic intellectual disability. Herein, we report a proposita and her mother with phenotypic features overlapping PACS1-NDD and a novel PACS1 variant (NM_018026.3:c.[755C > T];[=], p.(Ser252Phe)) that impedes binding of the adaptor protein GGA3 (Golgi-associated, gamma-adaptin ear-containing, ARF-binding protein 3). We hypothesize that attenuating PACS1 binding of GGA3 also gives rise to a disorder with features overlapping those of PACS1-NDD. This observation better delineates the mechanism by which PACS1 variation predisposes to syndromic intellectual disability.

The Clinical Variant Analysis Tool: Analyzing the evidence supporting reported genomic variation in clinical practice.

PURPOSE: Genomic test results, regardless of laboratory variant classification, require clinical practitioners to judge the applicability of a variant for medical decisions. Teaching and standardizing clinical interpretation of genomic variation calls for a methodology or tool. METHODS: To generate such a tool, we distilled the Clinical Genome Resource framework of causality and the American College of Medical Genetics/Association of Molecular Pathology and Quest Diagnostic Laboratory scoring of variant deleteriousness into the Clinical Variant Analysis Tool (CVAT). Applying this to 289 clinical exome reports, we compared the performance of junior practitioners with that of experienced medical geneticists and assessed the utility of reported variants. RESULTS: CVAT enabled performance comparable to that of experienced medical geneticists. In total, 124 of 289 (42.9%) exome reports and 146 of 382 (38.2%) reported variants supported a diagnosis. Overall, 10.5% (1 pathogenic [P] or likely pathogenic [LP] variant and 39 variants of uncertain significance [VUS]) of variants were reported in genes without established disease association; 20.2% (23 P/LP and 54 VUS) were in genes without sufficient phenotypic concordance; 7.3% (15 P/LP and 13 VUS) conflicted with the known molecular disease mechanism; and 24% (91 VUS) had insufficient evidence for deleteriousness. CONCLUSION: Implementation of CVAT standardized clinical interpretation of genomic variation and emphasized the need for collaborative and transparent reporting of genomic variation.

X-linked neonatal-onset epileptic encephalopathy associated with a gain-of-function variant p.R660T in GRIA3.

The X-linked GRIA3 gene encodes the GLUA3 subunit of AMPA-type glutamate receptors. Pathogenic variants in this gene were previously reported in neurodevelopmental diseases, mostly in male patients but rarely in females. Here we report a de novo pathogenic missense variant in GRIA3 (c.1979G>C; p. R660T) identified in a 1-year-old female patient with severe epilepsy and global developmental delay. When exogenously expressed in human embryonic kidney (HEK) cells, GLUA3_R660T showed slower desensitization and deactivation kinetics compared to wildtype (wt) GLUA3 receptors. Substantial non-desensitized currents were observed with the mutant but not for wt GLUA3 with prolonged exposure to glutamate. When co-expressed with GLUA2, the decay kinetics were similarly slowed in GLUA2/A3_R660T with non-desensitized steady state currents. In cultured cerebellar granule neurons, miniature excitatory postsynaptic currents (mEPSCs) were significantly slower in R660T transfected cells than those expressing wt GLUA3. When overexpressed in hippocampal CA1 neurons by in utero electroporation, the evoked EPSCs and mEPSCs were slower in neurons expressing R660T mutant compared to those expressing wt GLUA3. Therefore our study provides functional evidence that a gain of function (GoF) variant in GRIA3 may cause epileptic encephalopathy and global developmental delay in a female subject by enhancing synaptic transmission.

De novo single exon deletion of AUTS2 in a patient with speech and language disorder: a review of disrupted AUTS2 and further evidence for its role in neurodevelopmental disorders.

The autism susceptibility candidate 2 (AUTS2) gene is suggested to play a critical role in early brain development, and its association with intellectual disability (ID), autism spectrum disorders, and other neurodevelopmental disorders (NDDs) has recently gained more attention. Genomic rearrangements and copy number variations (CNVs) involving AUTS2 have been implicated in a range of NDDs with or without congenital malformations and dysmorphic features. Here we report a 62 kb de novo deletion encompassing exon 6 of AUTS2 detected by chromosomal microarray analysis (CMA) in a 4.5 year-old female patient with severe speech and language disorder, history of tonic-clonic movements, and pes planus with eversion of the feet. This is one of the smallest de novo intragenic deletions of AUTS2 described in patients with NDDs. We reviewed previously reported small pathogenic CNVs (<300 kb) in 19 cases, and correlated their specific locations within AUTS2 as well as presence of enhancers, regulatory elements, and CpG islands with the clinical findings of these cases and our patient. Our report provides additional insight into the clinical spectrum of AUTS2 disruptions.