CASK pathogenic variant which expands the clinical spectrum for MICPCH syndrome in an adult patient.

The CASK gene and its product protein kinase have been associated with microcephaly with pontine and cerebellar hypoplasia (MICPCH) syndrome and various other neurodevelopmental disorders. Clinical presentation is highly variable and generally includes intellectual disability, neurological disorders, and dysmorphic features, at a minimum. We present the case of one of the oldest known currently living patients with MICPCH syndrome with additional features not previously described in the literature (midface retrusion, macroglossia, dental crowding, adolescent-onset contractures at large joints, laxity at finger joints, and prominent wrist dystonia). Progressive hypertonicity throughout the patient's life has been managed with serial botulinum toxin injections. A comprehensive multimodal care team including physiatry, physical therapy, exercise therapy, and audiology has been assisting her with hearing deficits, communication skills, and mobility. This potentially expands the phenotype of MICPCH syndrome and provides information about the management of this condition into adulthood.

The biological functions and pathological mechanisms of CASK in various diseases.

Background: As a scaffold protein, calcium/calmodulin-dependent serine protein kinase (CASK) has been extensively studied in a variety of tissues throughout the body. The Cask gene is ubiquitous in several tissues, such as the neurons, islets, heart, kidneys and sperm, and is mostly localised in the cytoplasm adjacent to the basement membrane. CASK binds to a variety of proteins through its domains to exerting its biological activity. Scope of review: Here, we discuss the role of CASK in multiple tissues throughout the body. The role of different CASK domains in regulating neuronal development, neurotransmitter release and synaptic vesicle secretion was emphasised; the regulatory mechanism of CASK on the function of pancreatic islet ß cells was analysed; the role of CASK in cardiac physiology, kidney and sperm development was discussed; and the role of CASK in different tumours was compared. Finally, we clarify the importance of the Cask gene in the body, and how deletion or mutation of the Cask gene can have adverse consequences. Major conclusions: CASK is a conserved gene with similar roles in various tissues. The function of the Cask gene in the nervous system is mainly involved in the development of the nervous system and the release of neurotransmitters. In the endocrine system, an involvement of CASK has been reported in the process of insulin vesicle transport. CASK is also involved in cardiomyocyte ion channel regulation, kidney and sperm development, and tumour proliferation. CASK is an indispensable gene for the whole body, and CASK mutations can cause foetal malformations or death at birth. In this review, we summarise the biological functions and pathological mechanisms of CASK in various systems, thereby providing a basis for further in-depth studies of CASK functions.

Momentum informed muon scattering tomography for monitoring spent nuclear fuels in dry storage cask.

Development of an effective monitoring method for spent nuclear fuel (SNF) in a dry storage cask (DSC) is important to meet the increasing demand for dry storage investigations. The DSC investigation should provide information about the quantity of stored SNF, and quality assurance of materials should be possible without opening the cask. However, traditional nondestructive examination (NDE) methods such as x-rays are difficult to deploy for DSC investigation because a typical DSC is intentionally designed to shield against radiation. To address this challenge, cosmic ray muons (CRMs) are used as an alternative NDE radiation probe because they can easily penetrate an entire DSC system; however, a wide application of muons is often hindered due to the naturally low CRM flux (~104 muons/m2/min). This paper introduces a newly proposed imaging algorithm, momentum-informed muon scattering tomography (MMST), and presents how a limitation of the current muon scattering tomography technique has been addressed by measuring muon momentum. To demonstrate its functionality, a commercial DSC with 24 pressurized light water reactor fuel assemblies (FAs) and the MMST system were designed in GEANT4. Three noticeable improvements were observed for MMST system as a DSC investigation tool: (1) a signal stabilization, (2) an enhanced capability to differentiate various materials, and (3) statistically increased precision to identify and locate missing FAs. The results show that MMST improves the investigation accuracy from 79 to 98% when one FA is missing and 51% to 88% when one-half FA is missing. The advancement of the NDE technique using CRM for DSC verification is expected to resolve long-standing problems in increasing demand for DSC inspections and nuclear security.

CASK Mediates Oxidative Stress-Induced Microglial Apoptosis-Inducing Factor-Independent Parthanatos Cell Death via Promoting PARP-1 Hyperactivation and Mitochondrial Dysfunction.

Calcium/calmodulin-dependent serine protein kinase (CASK) is a scaffold protein and plays critical roles in neuronal synaptic formation and brain development. Previously, CASK was shown to associate with EGFR to maintain the vulval cell differentiation in C. elegans. In this study, we explored the role of CASK in CHME3 microglial cells. We found that CASK silencing protects cells from H2O2-induced cell death by attenuating PARP-1 activation, mitochondrial membrane potential loss, reactive oxygen species production, and mitochondrial fission, but it increases oxidative phosphorylation. The PARP-1 inhibitor olaparib blocks H2O2-induced cell death, suggesting the death mode of parthanatos. CASK silencing also increases AKT activation but decreases AMPK activation under H2O2 treatment. Pharmacological data further indicate that both signaling changes contribute to cell protection. Different from the canonical parthanatos pathway, we did not observe the AIF translocation from mitochondria into the nucleus, suggesting a non-canonical AIF-independent parthanatos in H2O2-treated CHME3 cells. Moreover, we found that CASK silencing upregulates the EGFR gene and protein expression and increases H2O2-induced EGFR phosphorylation in CHME3 microglia. However, EGFR activation does not contribute to cell protection caused by CASK silencing. In conclusion, CASK plays a crucial role in microglial parthanatos upon H2O2 treatment via stimulation of PARP-1 and AMPK but the inhibition of AKT. These findings suggest that CASK might be an ideal therapeutic target for CNS disorders.

Novel characterization of CASK variant c.1963 A>G (p.Asn655Asp) through whole-exome sequencing in a monochorionic diamniotic twin fetus with significant brain anomalies: A case report.

Whole-exome sequencing is an evolving technology in perinatal diagnosis which allows identification of genetic etiologies that would otherwise go undetermined. In this case report, a 38-year-old Hispanic woman, G5P3013, with a monochorionic diamniotic twin gestation with one fetus displaying significant cranial abnormalities on prenatal ultrasound and magnetic resonance imaging (MRI) of the brain is presented. Fetal anomalies included bilateral ventriculomegaly, absent cavum septum pellucidum, and absent corpus callosum. Diagnostic amniocentesis with chromosome analysis, chromosomal microarray, alpha-fetoprotein, cytomegalovirus, toxoplasmosis, and parvovirus had normal results. Whole-exome sequencing for the anomalous fetus detected a de novo mosaic variant of uncertain significance (VUS) in the calcium/calmodulin dependent serine protein kinase (CASK) gene: c.1963 A > G (p.Asn655Asp). This variant was absent in the normal twin fetus, the mother, and the father. Pathogenic CASK gene mutations are associated with three syndromes: FG syndrome 4, intellectual developmental disorder and microcephaly with pontine and cerebellar hypoplasia (MICPCH), and intellectual developmental disorder with or without nystagmus. Whole-exome sequencing identified a potential etiology for the anomalies detected. The variant likely arose de novo and was the potential cause of the identified cranial abnormalities in one fetus of this monochorionic diamniotic twin gestation. Whole-exome sequencing may provide additional diagnostic utility when standard diagnostic testing is noncontributory.

Interrogation and validation of the interactome of neuronal Munc18-interacting Mint proteins with AlphaFold2.

Munc18-interacting proteins (Mints) are multidomain adaptors that regulate neuronal membrane trafficking, signaling, and neurotransmission. Mint1 and Mint2 are highly expressed in the brain with overlapping roles in the regulation of synaptic vesicle fusion required for neurotransmitter release by interacting with the essential synaptic protein Munc18-1. Here, we have used AlphaFold2 to identify and then validate the mechanisms that underpin both the specific interactions of neuronal Mint proteins with Munc18-1 as well as their wider interactome. We found that a short acidic α-helical motif within Mint1 and Mint2 is necessary and sufficient for specific binding to Munc18-1 and binds a conserved surface on Munc18-1 domain3b. In Munc18-1/2 double knockout neurosecretory cells, mutation of the Mint-binding site reduces the ability of Munc18-1 to rescue exocytosis, and although Munc18-1 can interact with Mint and Sx1a (Syntaxin1a) proteins simultaneously in vitro, we find that they have mutually reduced affinities, suggesting an allosteric coupling between the proteins. Using AlphaFold2 to then examine the entire cellular network of putative Mint interactors provides a structural model for their assembly with a variety of known and novel regulatory and cargo proteins including ADP-ribosylation factor (ARF3/ARF4) small GTPases and the AP3 clathrin adaptor complex. Validation of Mint1 interaction with a new predicted binder TJAP1 (tight junction-associated protein 1) provides experimental support that AlphaFold2 can correctly predict interactions across such large-scale datasets. Overall, our data provide insights into the diversity of interactions mediated by the Mint family and show that Mints may help facilitate a key trigger point in SNARE (soluble N-ethylmaleimide-sensitive factor attachment receptor) complex assembly and vesicle fusion.

Diverse Clinical Phenotypes of CASK-Related Disorders and Multiple Functional Domains of CASK Protein.

CASK-related disorders are a form of rare X-linked neurological diseases and most of the patients are females. They are characterized by several symptoms, including microcephaly with pontine and cerebellar hypoplasia (MICPCH), epilepsy, congenital nystagmus, and neurodevelopmental disorders. Whole-genome sequencing has identified various mutations, including nonsense and missense mutations, from patients with CASK-related disorders, revealing correlations between specific mutations and clinical phenotypes. Notably, missense mutations associated with epilepsy and intellectual disability were found throughout the whole region of the CASK protein, while missense mutations related to microcephaly and MICPCH were restricted in certain domains. To investigate the pathophysiology of CASK-related disorders, research groups have employed diverse methods, including the generation of CASK knockout mice and the supplementation of CASK to rescue the phenotypes. These approaches have yielded valuable insights into the identification of functional domains of the CASK protein associated with a specific phenotype. Additionally, recent advancements in the AI-based prediction of protein structure, such as AlphaFold2, and the application of genome-editing techniques to generate CASK mutant mice carrying missense mutations from patients with CASK-related disorders, allow us to understand the pathophysiology of CASK-related disorders in more depth and to develop novel therapeutic methods for the fundamental treatment of CASK-related disorders.

Presynaptic Cytomatrix Proteins.

The Cytomatrix Assembled at the active Zone (CAZ) of a presynaptic terminal displays electron-dense appearance and defines the center of the synaptic vesicle release. The protein constituents of CAZ are multiple-domain scaffolds that interact extensively with each other and also with an ensemble of synaptic vesicle proteins to ensure docking, fusion, and recycling. Reflecting the central roles of the active zone in synaptic transmission, CAZ proteins are highly conserved throughout evolution. As the nervous system increases complexity and diversity in types of neurons and synapses, CAZ proteins expand in the number of gene and protein isoforms and interacting partners. This chapter summarizes the discovery of the core CAZ proteins and current knowledge of their functions.

KIRREL3-related disorders: a case report confirming the radiological features and expanding the clinical spectrum to a less severe phenotype.

BACKGROUND: Neurodevelopmental disorders have a multifactorial etiology, since biological, genetic, psychosocial and environmental risk factors are involved. Recent studies have been linking neurodevelopmental disorders and intellectual disability with a variety of genes, some of which encoding neuronal cell-adhesion molecules. Among these, KIRREL3 is known to play a role in CNS development, and his variants have recently been related to intellectual disability, autism spectrum disorder, childhood apraxia of speech, cerebellar hypoplasia and mild dysmorphic features. CASE PRESENTATION: In this study, we describe a young Caucasian boy with mild intellectual disability, cerebellar anomalies (cerebellar hypoplasia and mega cisterna magna) and minor dysmorphic features associated to a novel KIRREL3 variant. CONCLUSIONS: Aim of the present case report is to expand the clinical spectrum of KIRREL3-related diseases towards a milder phenotype than what is already described in the literature. We speculate that the interaction between KIRREL3 and CASK might play a major role in promoting cognitive and cerebellar development, contributing to a variety of clinical manifestations.

mRNA location and translation rate determine protein targeting to dual destinations.

Numerous proteins are targeted to two or multiple subcellular destinations where they exert distinct functional consequences. The balance between such differential targeting is thought to be determined post-translationally, relying on protein sorting mechanisms. Here, we show that mRNA location and translation rate can also determine protein targeting by modulating protein binding to specific interacting partners. Peripheral localization of the NET1 mRNA and fast translation lead to higher cytosolic retention of the NET1 protein by promoting its binding to the membrane-associated scaffold protein CASK. By contrast, perinuclear mRNA location and/or slower translation rate favor nuclear targeting by promoting binding to importins. This mRNA location-dependent mechanism is modulated by physiological stimuli and profoundly impacts NET1 function in cell motility. These results reveal that the location of protein synthesis and the rate of translation elongation act in coordination as a "partner-selection" mechanism that robustly influences protein distribution and function.

Calcium/Calmodulin-Dependent Serine Protein Kinase (CASK) Gene Polymorphisms in Pigeons.

Calcium/calmodulin-dependent serine protein kinase (CASK) is an multidomain protein involved in tissue development and cell signalling. In skeletal muscle, it is involved in the development of neuromuscular junctions. The participation of a pigeon in racing is a great physical effort that causes many changes in the skeletal muscles. Thus, the purpose of the study was to detect the nucleotide sequence variability in the calcium/calmodulin-dependent serine kinase (CASK) gene in domestic pigeons (Columba livia domestica) and assess the potential impact of DNA polymorphisms on the flight performance of pigeons. The research included a total of 517 individuals. DNA was extracted from the blood. A DNA fragment from nucleotides 8689 to 9049 of the CASK (NW_004973256.1 sequence) of six unrelated pigeons were sequenced. One of the detected polymorphic sites (g.8893G > A), located a very close to the start codon, was selected for genotyping in all individuals. The association studies included a total of 311 young homing pigeons that participated in racing competitions. The homing pigeons showed higher frequencies of the AA genotype than non-homing ones (p < 0.05). In rock pigeons only the GG genotype was found. Further research could confirm the functionality of the CASK g.8893G > A SNP in shaping the racing phenotype of pigeons, and the AA genotype could be useful as a selection criterion in pigeon breeding.

RNA in situ conformation sequencing reveals novel long-range RNA structures with impact on splicing.

Over recent years, long-range RNA structure has emerged as a factor that is fundamental to alternative splicing regulation. An increasing number of human disorders are now being associated with splicing defects; hence it is essential to develop methods that assess long-range RNA structure experimentally. RNA in situ conformation sequencing (RIC-seq) is a method that recapitulates RNA structure within physiological RNA-protein complexes. In this work, we juxtapose pairs of conserved complementary regions (PCCRs) that were predicted in silico with the results of RIC-seq experiments conducted in seven human cell lines. We show statistically that RIC-seq support of PCCRs correlates with their properties, such as equilibrium free energy, presence of compensatory substitutions, and occurrence of A-to-I RNA editing sites and forked eCLIP peaks. Exons enclosed in PCCRs that are supported by RIC-seq tend to have weaker splice sites and lower inclusion rates, which is indicative of post-transcriptional splicing regulation mediated by RNA structure. Based on these findings, we prioritize PCCRs according to their RIC-seq support and show, using antisense nucleotides and minigene mutagenesis, that PCCRs in two disease-associated human genes, PHF20L1 and CASK, and also PCCRs in their murine orthologs, impact alternative splicing. In sum, we demonstrate how RIC-seq experiments can be used to discover functional long-range RNA structures, and particularly those that regulate alternative splicing.

Structural Analysis Implicates CASK-Liprin-α2 Interaction in Cerebellar Granular Cell Death in MICPCH Syndrome.

Microcephaly with pontine and cerebellar hypoplasia (MICPCH) syndrome is a neurodevelopmental disorder caused by the deficiency of the X-chromosomal gene CASK. However, the molecular mechanisms by which CASK deficiency causes cerebellar hypoplasia in this syndrome remain elusive. In this study, we used CASK knockout (KO) mice as models for MICPCH syndrome and investigated the effect of CASK mutants. Female CASK heterozygote KO mice replicate the progressive cerebellar hypoplasia observed in MICPCH syndrome. CASK KO cultured cerebellar granule (CG) cells show progressive cell death that can be rescued by co-infection with lentivirus expressing wild-type CASK. Rescue experiments with CASK deletion mutants identify that the CaMK, PDZ, and SH3, but not L27 and guanylate kinase domains of CASK are required for the survival of CG cells. We identify missense mutations in the CaMK domain of CASK derived from human patients that fail to rescue the cell death of cultured CASK KO CG cells. Machine learning-based structural analysis using AlphaFold 2.2 predicts that these mutations disrupt the structure of the binding interface with Liprin-α2. These results suggest that the interaction with Liprin-α2 via the CaMK domain of CASK may be involved in the pathophysiology of cerebellar hypoplasia in MICPCH syndrome.

Cask wine: Describing drinking patterns associated with Australia's cheapest alcohol.

INTRODUCTION: In Australia, cask wine is the cheapest alcoholic beverage available, offering the lowest price per standard drink. Despite this, there is little research on the contextual correlates of cask wine consumption. Therefore, the current study aims to describe how cask wine consumption has changed over the last decade. Then, through comparisons between cask and bottled wine, how prices, typical drinking locations, and patterns of consumption differ between the beverages. METHODS: Cross-sectional data was drawn from two sources. Four waves of the National Drug Strategy Household Survey were used (2010, 2013, 2016 and 2019) in order to examine consumption trends over time. The International Alcohol Control study (2013) in Australia was additionally used to explore pricing and consumption trends in greater detail. RESULTS: Cask wine was considerably cheaper than other forms of wine at $0.54 per standard drink (95% confidence interval [CI] 0.45-0.62, p < 0.05). Consumption trends associated with cask wine differed from that of bottled wine, being consumed almost entirely at home and in significantly greater quantity (standard drinks per day 7.8, 95% CI 6.25-9.26, p < 0.05). Among the heaviest drinkers, 13% (95% CI 7.2-18.8, p < 0.05) consumed cask wine as their main drink, compared to 5% (95% CI 3.76-6.24, p < 0.05) consuming bottled wine. CONCLUSIONS: Cask wine drinkers are disproportionately more likely to consume higher amounts of alcohol, paying less per drink doing so compared to bottled wine drinkers. As all cask wine purchases were under $1.30, a minimum unit price may largely affect cask wine purchases, applying to a far smaller proportion of bottled wine.

A Threshold Helium Leakage Detection Switch with Ultra Low Power Operation.

Detecting helium leakage is important in many applications, such as in dry cask nuclear waste storage systems. This work develops a helium detection system based on the relative permittivity (dielectric constant) difference between air and helium. This difference changes the status of an electrostatic microelectromechanical system (MEMS) switch. The switch is a capacitive-based device and requires a very negligible amount of power. Exciting the switch's electrical resonance enhances the MEMS switch sensitivity to detect low helium concentration. This work simulates two different MEMS switch configurations: a cantilever-based MEMS modeled as a single-degree-freedom model and a clamped-clamped beam MEMS molded using the COMSOL Multiphysics finite-element software. While both configurations demonstrate the switch's simple operation concept, the clamped-clamped beam was selected for detailed parametric characterization due to its comprehensive modeling approach. The beam detects at least 5% helium concentration levels when excited at 3.8 MHz, near electrical resonance. The switch performance decreases at lower excitation frequencies or increases the circuit resistance. The MEMS sensor detection level was relatively immune to beam thickness and parasitic capacitance changes. However, higher parasitic capacitance increases the switch's susceptibility to errors, fluctuations, and uncertainties.

A Missense Variant in CASKIN1's Proline-Rich Region Segregates with Psychosis in a Three-Generation Family.

The polygenic nature of schizophrenia (SCZ) implicates many variants in disease development. Rare variants of high penetrance have been shown to contribute to the disease prevalence. Whole-exome sequencing of a large three-generation family with SCZ and bipolar disorder identified a single segregating novel, rare, non-synonymous variant in the gene CASKIN1. The variant D1204N is absent from all databases, and CASKIN1 has a gnomAD missense score Z = 1.79 and pLI = 1, indicating its strong intolerance to variation. We find that introducing variants in the proline-rich region where the D1204N resides results in significant cellular changes in iPSC-derived neurons, consistent with CASKIN1's known functions. We observe significant transcriptomic changes in 368 genes (padj < 0.05) involved in neuronal differentiation and nervous system development. We also observed nominally significant changes in the frequency of action potentials during differentiation, where the speed at which the edited and unedited cells reach the same level of activity differs. Our results suggest that CASKIN1 is an excellent gene candidate for psychosis development with high penetrance in this family.

Mosaicism of a Truncating Variant of CASK Causes Congenital Heart Disease and Neurodevelopmental Disorder.

Introduction: Calcium/calmodulin-dependent serine protein kinase (CASK) gene mutations cause microcephaly with pontine and cerebellar hypoplasia (MICPCH) and X-linked intellectual disability. Congenital heart disease (CHD) is a rare complication reported in only 4 male patients with full loss-of-function mutations. Here, we report the first male patient with mosaicism of a truncating variant of CASK complicated by CHD. Case Presentation: The patient is a 6-year-old male with MICPCH, ventricular septal defect, and developmental delay. He achieved rolling over but can not speak meaningful words. We identified a somatic mosaic variant of CASK: c.[725=/G>A], p.(W242*) and high mosaic ratios of 90% and 84% for mutant alleles in peripheral blood lymphocytes and skin fibroblasts, respectively. His developmental delay was severe but milder than that of previously reported CHD patients. Discussion: Truncating CASK variants may be associated with CHD, even in a mosaic state, and even a low normal allele ratio could lengthen survivorship.

Case Report: Identification of a novel CASK missense variant in a Chinese family with MICPCH.

Mental retardation and microcephaly with pontine and cerebellar hypoplasia (MICPCH) is a rare genetic disorder that results in varying levels of pontocerebellar hypoplasia, microcephaly, and severe intellectual disabilities. Prior genetic analyses have identified the CASK gene as a driver of MICPCH. Herein, we analyzed a Chinese family with MICPCH. The index patient was an 8-year-old male. He and his 3-year-old brother suffered from microcephaly, pontocerebellar hypoplasia, serious mental retardation, ataxia, gait disorder, and inability to speak. Through a combination of whole-exome sequencing and subsequent Sanger sequencing, a novel X-linked missense mutation, c.1882G>C (p.D628H) in the CASK gene, was identified in two siblings, as well as their mother and grandmother, who exhibited mild mental retardation. Other family members with negative genetic testing were normal. In silico analyses indicated that this missense mutation was predicted to reduce CASK protein stability, disrupt the SRC homology 3 (SH3) domain, and abolish its function. In summary, we identified a novel missense variate in CASK associated with MICPCH. Our work facilitates the diagnosis of the disease in this family and broadens the gene variant spectrum of the CASK in MICPCH patients.

Case report: A novel CASK mutation in a Chinese female child with microcephaly with pontine and cerebellar hypoplasia.

Objective: Microcephaly with pontine and cerebellar hypoplasia (MICPCH) is a rare X-linked dominant genetic disease, and most MICPCHs are ascribed to CASK mutations, while few are revealed in Chinese patients. This study aims to identify the pathogenic mutation in a Chinese proband with MICPCH. Methods: A 3-year-old female Chinese proband with MICPCH and her parents were included. Clinical data were collected from the medical records and recalled by the proband's mother. Whole genome sequencing and Sanger sequencing were used to find the pathogenic mutation of MICPCH. Results: The proband presented with postnatal progressive microcephaly, cerebellar hypoplasia, intellectual disability, motor and language development retardation and limb hypertonia. Genetic analysis indicated that there was a novel compound heterozygote nonsynonymous mutation, c.755T>C(p.Leu252Pro) in exon8 of CASK gene in the proband, but not in her parents. This CASK mutation has not been reported in other databases. Conclusion: This study broadens the mutation spectrum of the CASK gene and is of great value for precise prenatal diagnosis and genetic counseling.

Two heterozygous mutations in the calcium/calmodulin-dependent serine protein kinase gene (CASK) in cases with developmental disorders.

BACKGROUND: The calcium/calmodulin-dependent serine protein kinase gene (CASK) is an essential gene in mammals, critical for neurodevelopment. The purpose of this study is to expand the understanding of the diagnosis of CASK-linked disorders. MATERIALS/METHODS: From clinical and genetic mutational analyses, relevant data in 2 Han Chinese patients were collected and analyzed. Real-time quantitative PCR (RT-qPCR) was performed to investigate the CASK expression levels in the patients. The X-chromosome inactivation (XCI) patterns of the patients and their nuclear families were tested by quantitation of methylation of the polymorphic human androgen receptor (HUMARA) locus. RESULTS: Two Han Chinese patients both presented with intellectual disability (ID), microcephaly with pontine and cerebellar hypoplasia (MICPCH). Two de novo mutations of c.82C>T (p.Arg28*) and c.846C>G (p.Tyr282*) in CASK have been investigated and predicted to be deleterious, which have produced truncated proteins. The functional protein association network of STRING (http://string-db.org) generated three-dimensional (3D) atomic models based on protein sequences in CASK and two Arg28 and Tyr282 residues were marked. RT-qPCR showed lower copy numbers of CASK expression in the patients than in their parents, as well as the sex- and age- matched control groups. Patient 1 showed a skewed XCI pattern, while no related changes noted in patient 2. CONCLUSIONS: Patients carrying different nonsense variants may have different degrees of different clinical phenotypes. This study expands the spectrum of genotype and phenotype correlations of CASK-linked disorders in the Han Chinese ethnicity and provides new insights into the molecular mechanism.