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Intellectual disability is a common and highly heterogeneous disorder etiologically. In a multiplex consanguineous family, we applied autozygosity mapping and exome sequencing and identified a novel homozygous truncating mutation in PUS3 that fully segregates with the intellectual disability phenotype. Consistent with the known role of Pus3 in isomerizing uracil to pseudouridine at positions 38 and 39 in tRNA, we found a significant reduction in this post-transcriptional modification of tRNA in patient cells. Our finding adds to a growing list of intellectual disability disorders that are caused by perturbation of various tRNA modifications, which highlights the sensitivity of the brain to these highly conserved processes.

Background At least 50% of patients with suspected Mendelian disorders remain undiagnosed after whole-exome sequencing (WES), and the extent to which non-coding variants that are not captured by WES contribute to this fraction is unclear. Whole transcriptome sequencing is a promising supplement to WES, although empirical data on the contribution of RNA analysis to the diagnosis of Mendelian diseases on a large scale are scarce. Results Here, we describe our experience with transcript-deleterious variants (TDVs) based on a cohort of 5647 families with suspected Mendelian diseases. We first interrogate all families for which the respective Mendelian phenotype could be mapped to a single locus to obtain an unbiased estimate of the contribution of TDVs at 18.9%. We examine the entire cohort and find that TDVs account for 15% of all “solved” cases. We compare the results of RT-PCR to in silico prediction. Definitive results from RT-PCR are obtained from blood-derived RNA for the overwhelming majority of variants (84.1%), and only a small minority (2.6%) fail analysis on all available RNA sources (blood-, skin fibroblast-, and urine renal epithelial cells-derived), which has important implications for the clinical application of RNA-seq. We also show that RNA analysis can establish the diagnosis in 13.5% of 155 patients who had received “negative” clinical WES reports. Finally, our data suggest a role for TDVs in modulating penetrance even in otherwise highly penetrant Mendelian disorders. Conclusions Our results provide much needed empirical data for the impending implementation of diagnostic RNA-seq in conjunction with genome sequencing.

Pseudouridylation is the most common post-transcriptional modification, wherein uridine is isomerized into 5-ribosyluracil (pseudouridine, Ψ). The resulting increase in base stacking and creation of additional hydrogen bonds are thought to enhance RNA stability. Pseudouridine synthases are encoded in humans by 13 genes, two of which are linked to Mendelian diseases: PUS1 and PUS3. Very recently, PUS7 mutations were reported to cause intellectual disability with growth retardation. We describe two families in which two different homozygous PUS7 mutations (missense and frameshift deletion) segregate with a phenotype comprising intellectual disability and progressive microcephaly. Short stature and hearing loss were variable in these patients. Functional characterization of the two mutations confirmed that both result in decreased levels of Ψ13 in tRNAs. Furthermore, the missense variant of the S. cerevisiae ortholog failed to complement the growth defect of S. cerevisiae pus7Δ trm8Δ mutants. Our results confirm that PUS7 is a bona fide Mendelian disease gene and expand the list of human diseases caused by impaired pseudouridylation.

Background Pathogenic variants in SPTBN4 have been linked to autosomal recessive “neurodevelopmental disorder with hypotonia, neuropathy, and deafness” (MIM# 617519) known as NEDHND. The disorder is highlighted with neuropathy, muscle weakness, and infrequent appearance of seizures in the affected individuals. This study aims to investigate the natural history of the disease, present genetic and clinical appearance of the syndrome in a highly consanguineous population, Saudi Arabia, and finally provide an overview of the reported cases, their clinical features, and disease-causing variants. Methods The study started with a search through neurology clinics and local databases and utilized genetic testing records after diagnosing a patient with NEDHND at our hospital (King Faisal Specialist Hospital and Research Centre, KFSHRC). Based on the search we have identified additional patients (in total, n  = 10) with the disease and performed genetic testing using whole exome sequencing and confirmatory Sanger sequencing. We performed RT-PCR on RNA extracted from lymphoblastoid cell line from a patient who found to have an aberrant splicing variant. Finally, we comprehensively reviewed current literature and available data related to the disease. Results We present natural history of SPTBN4-associated neurodevelopmental disorder with hypotonia, neuropathy, and deafness in addition to four Saudi families with ten affected individuals who share clinical features of NEDHND. We report three known mutations and one novel nonsense variant, highlight atypical clinical features related to cerebellar involvement, confirm the pathogenicity of a splicing variant by RT-PCR, and review the findings of previously reported patients. Conclusion Our study defines the clinical phenotype of a cohort of NEDHND in detail including the evolution of patients’ clinical features, compares them to previously reported cases, and utilizes the existing data on the disease to direct development of a better prevention plan by means of genetic and preimplantation counseling. Our study may help and enable future clinical trials focusing on NEDHND in our country.

Background Variable expressivity is a well-known phenomenon in which patients with mutations in one gene display varying degrees of clinical severity, potentially displaying only subsets of the clinical manifestations associated with the multisystem disorder linked to the gene. This remains an incompletely understood phenomenon with proposed mechanisms ranging from allele-specific to stochastic. Results We report three consanguineous families in which an isolated ocular phenotype is linked to a novel 3′ UTR mutation in SLC4A4, a gene known to be mutated in a syndromic form of intellectual disability with renal and ocular involvement. Although SLC4A4 is normally devoid of AU-rich elements (AREs), a 3′ UTR motif that mediates post-transcriptional control of a subset of genes, the mutation we describe creates a functional ARE. We observe a marked reduction in the transcript level of SLC4A4 in patient cells. Experimental confirmation of the ARE-creating mutation is shown using a post-transcriptional reporter system that reveals consistent reduction in the mRNA-half life and reporter activity. Moreover, the neo-ARE binds and responds to the zinc finger protein ZFP36/TTP, an ARE-mRNA decay-promoting protein. Conclusions This novel mutational mechanism for a Mendelian disease expands the potential mechanisms that underlie variable phenotypic expressivity in humans to also include 3′ UTR mutations with tissue-specific pathology.

Purpose Ciliopathies are highly heterogeneous clinical disorders of the primary cilium. We aim to characterize a large cohort of ciliopathies phenotypically and molecularly. Methods Detailed phenotypic and genomic analysis of patients with ciliopathies, and functional characterization of novel candidate genes. Results In this study, we describe 125 families with ciliopathies and show that deleterious variants in previously reported genes, including cryptic splicing variants, account for 87% of cases. Additionally, we further support a number of previously reported candidate genes ( BBIP1 , MAPKBP1 , PDE6D , and WDPCP ), and propose nine novel candidate genes ( CCDC67 , CCDC96 , CCDC172 , CEP295 , FAM166B , LRRC34 , TMEM17 , TTC6 , and TTC23 ), three of which ( LRRC34 , TTC6 , and TTC23 ) are supported by functional assays that we performed on available patient-derived fibroblasts. From a phenotypic perspective, we expand the phenomenon of allelism that characterizes ciliopathies by describing novel associations including WDR19 -related Stargardt disease and SCLT1 - and CEP164 -related Bardet–Biedl syndrome. Conclusion In this cohort of phenotypically and molecularly characterized ciliopathies, we draw important lessons that inform the clinical management and the diagnostics of this class of disorders as well as their basic biology.

Purpose Congenital microcephaly (CM) is an important birth defect with long term neurological sequelae. We aimed to perform detailed phenotypic and genomic analysis of patients with Mendelian forms of CM. Methods Clinical phenotyping, targeted or exome sequencing, and autozygome analysis. Results We describe 150 patients (104 families) with 56 Mendelian forms of CM. Our data show little overlap with the genetic causes of postnatal microcephaly. We also show that a broad definition of primary microcephaly —as an autosomal recessive form of nonsyndromic CM with severe postnatal deceleration of occipitofrontal circumference—is highly sensitive but has a limited specificity. In addition, we expand the overlap between primary microcephaly and microcephalic primordial dwarfism both clinically (short stature in >52% of patients with primary microcephaly) and molecularly (e.g., we report the first instance of CEP135 -related microcephalic primordial dwarfism). We expand the allelic and locus heterogeneity of CM by reporting 37 novel likely disease-causing variants in 27 disease genes, confirming the candidacy of ANKLE2, YARS , FRMD4A , and THG1L , and proposing the candidacy of BPTF , MAP1B , CCNH , and PPFIBP1 . Conclusion Our study refines the phenotype of CM, expands its genetics heterogeneity, and informs the workup of children born with this developmental brain defect.

Nuclear pore complex (NPC) is a fundamental component of the nuclear envelope and is key to the nucleocytoplasmic transport. Mutations in several NUP genes that encode individual components of NPC known as nucleoporins have been identified in recent years among patients with static encephalopathies characterized by developmental delay and microcephaly. We describe a multiplex consanguineous family in which four affected members presented with severe neonatal hypotonia, profound global developmental delay, progressive microcephaly and early death. Autozygome and linkage analysis revealed that this phenotype is linked to a founder disease haplotype (chr9:127,113,732-135,288,807) in which whole exome sequencing revealed the presence of a novel homozygous missense variant in NUP214. Functional analysis of patient-derived fibroblasts recapitulated the dysmorphic phenotype of nuclei that was previously described in NUP214 knockdown cells. In addition, the typical rim staining of NUP214 is largely displaced, further supporting the deleterious effect of the variant. Our data expand the list of NUP genes that are mutated in encephalopathy disorders in humans.

Excitatory neuronal homeostasis is crucial for neuronal survival, circuit function, and plasticity. Disruptions in this form of homeostasis are believed to underpin a variety of neuronal conditions including intellectual disability, epilepsy, and autism. However, the underlying genetic and molecular mechanisms maintaining this homeostasis remain poorly understood. Biallelic recurrent loss of function mutations in , an evolutionarily conserved X amino acid novel open reading frame, underlie Temtamy syndrome (TS)-a neurodevelopmental disorder characterized by epilepsy, dysgenesis of the corpus callosum, and severe intellectual disability. Through multiple lines of inquiry, we establish that C12ORF57/GRCC10 plays an unexpected central role in synaptic homeostatic downscaling in response to elevated activity, uncovering a novel mechanism for neuronal excitatory homeostasis. To probe these mechanisms, we developed a new knockout (KO) mouse model of the gene's murine ortholog, as well as cellular and assays. KO mice exhibit the characteristic phenotypic features seen in human TS patients, including increased epileptiform activity. Corresponding with the enhanced seizure susceptibility, hippocampal neurons in these mice exhibited significantly increased AMPA receptor expression levels and higher amplitude of miniature excitatory postsynaptic currents (mEPSCs). We further found that GRCC10/C12ORF57 modulates the activity of calcium/calmodulin dependent kinase 4 (CAMK4) and thereby regulates the expression of CREB and ARC. Our study suggests through this novel mechanism, deletion of Grcc10 disrupts the characteristic synaptic AMPA receptor downscaling that accompanies increased activity in glutamatergic neurons.