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MAD2L1BP-encoded p31comet mediates Trip13-dependent disassembly of Mad2- and Rev7-containing complexes and, through this antagonism, promotes timely spindle assembly checkpoint (SAC) silencing, faithful chromosome segregation, insulin signaling, and homology-directed repair (HDR) of DNA double-strand breaks. We identified a homozygous MAD2L1BP nonsense variant, R253*, in 2 siblings with microcephaly, epileptic encephalopathy, and juvenile granulosa cell tumors of ovary and testis. Patient-derived cells exhibited high-grade mosaic variegated aneuploidy, slowed-down proliferation, and instability of truncated p31comet mRNA and protein. Corresponding recombinant p31comet was defective in Trip13, Mad2, and Rev7 binding and unable to support SAC silencing or HDR. Furthermore, C-terminal truncation abrogated an identified interaction of p31comet with tp53. Another homozygous truncation, R227*, detected in an early-deceased patient with low-level aneuploidy, severe epileptic encephalopathy, and frequent blood glucose elevations, likely corresponds to complete loss of function, as in Mad2l1bp-/- mice. Thus, human mutations of p31comet are linked to aneuploidy and tumor predisposition.

CSNK2B encodes for casein kinase II subunit beta (CK2β), the regulatory subunit of casein kinase II (CK2), which is known to mediate diverse cellular pathways. Variants in this gene have been recently identified as a cause of Poirier-Bienvenu neurodevelopmental syndrome (POBINDS), but functional evidence is sparse. Here, we report five unrelated individuals: two of them manifesting POBINDS, while three are identified to segregate a new intellectual disability-craniodigital syndrome (IDCS), distinct from POBINDS. The three IDCS individuals carried two different de novo missense variants affecting the same codon of CSNK2B . Both variants, NP_001311.3; p.Asp32His and NP_001311.3; p.Asp32Asn, lead to an upregulation of CSNK2B expression at transcript and protein level, along with global dysregulation of canonical Wnt signaling. We found impaired interaction of the two key players DVL3 and β-catenin with mutated CK2β. The variants compromise the kinase activity of CK2 as evident by a marked reduction of phosphorylated β-catenin and consequent absence of active β-catenin inside nuclei of the patient-derived lymphoblastoid cell lines (LCLs). In line with these findings, whole-transcriptome profiling of patient-derived LCLs harboring the NP_001311.3; p.Asp32His variant confirmed a marked difference in expression of genes involved in the Wnt signaling pathway. In addition, whole-phosphoproteome analysis of the LCLs of the same subject showed absence of phosphorylation for 313 putative CK2 substrates, enriched in the regulation of nuclear β-catenin and transcription of the target genes. Our findings suggest that discrete variants in CSNK2B cause dominant-negative perturbation of the canonical Wnt signaling pathway, leading to a new craniodigital syndrome distinguishable from POBINDS., Competing Interests: The authors declare no competing interests., (© 2022 The Authors.)

Nonsyndromic hearing loss is an extremely heterogeneous disorder. Thus, clinical diagnostics is challenging, in particular due to differences in the etiology of hearing loss between populations. With this study, we wanted to elucidate the genetic basis of hearing loss in 61 consanguineous Egyptian families. In 25 families, linkage analysis was used as a prescreening to identify regions for targeted sequencing of candidate genes. Initially, the coding regions of 12 and later of 94 genes associated with hearing loss were enriched and subjected to massively parallel sequencing (MPS) with diagnostic yields of 36% and 75%, respectively. Causative variants were identified in 48 families (79%). They were found in 23 different genes with the majority being located in MYO15A (15.3%), SLC26A4 (9.7%), GJB2 (8.3%), and MYO7A (6.4%). As many as 32 variants were novel ones at the time of detection. Five variants were shared by two, three, or even four families. Our study provides a first survey of the mutational spectrum of deaf patients in Egypt revealing less GJB2 variants than in many European populations. It underlines the value of targeted enrichment of well-selected deafness genes in combination with MPS in the diagnostics of this frequent and genetically heterogeneous disorder., (© 2020 The Authors. Clinical Genetics published by John Wiley & Sons Ltd.)

Background: Biallelic PTPRQ pathogenic variants have been previously reported as causative for autosomal recessive non-syndromic hearing loss. In 2018 the first heterozygous PTPRQ variant has been implicated in the development of autosomal dominant non-syndromic hearing loss (ADNSHL) in a German family. The study presented the only, so far known, PTPRQ pathogenic variant (c.6881G>A) in ADNSHL. It is located in the last PTPRQ coding exon and introduces a premature stop codon (p.Trp2294*)., Methods: A five-generation Polish family with ADNSHL was recruited for the study (n = 14). Thorough audiological, neurotological and imaging studies were carried out to precisely define the phenotype. Genomic DNA was isolated from peripheral blood samples or buccal swabs of available family members. Clinical exome sequencing was conducted for the proband. Family segregation analysis of the identified variants was performed using Sanger sequencing. Single nucleotide polymorphism array on DNA samples from the Polish and the original German family was used for genome-wide linkage analysis., Results: Combining clinical exome sequencing and family segregation analysis, we have identified the same (NM_001145026.2:c.6881G>A, NP_001138498.1:p.Trp2294*) PTPRQ alteration in the Polish ADNSHL family. Using genome-wide linkage analysis, we found that the studied family and the original German family derive from a common ancestor. Deep phenotyping of the affected individuals showed that in contrast to the recessive form, the PTPRQ-related ADNSHL is not associated with vestibular dysfunction. In both families ADNSHL was progressive, affected mainly high frequencies and had a variable age of onset., Conclusion: Our data provide the first confirmation of PTPRQ involvement in ADNSHL. The finding strongly reinforces the inclusion of PTPRQ to the small set of genes leading to both autosomal recessive and dominant hearing loss.

We previously reported that inactivation of the transmembrane taurine transporter (TauT or solute carrier 6a6) causes early retinal degeneration in mice. Compatible with taurine's indispensability for cell volume homeostasis, protein stabilization, cytoprotection, antioxidation, and immuno- and neuromodulation, mice develop multisystemic dysfunctions (hearing loss; liver fibrosis; and behavioral, heart, and skeletal muscle abnormalities) later on. Here, by genetic, cell biologic, in vivo 1 H-magnetic resonance spectroscopy and molecular dynamics simulation studies, we conducted in-depth characterization of a novel disorder: human TAUT deficiency. Loss of TAUT function due to a homozygous missense mutation caused panretinal degeneration in 2 brothers. TAUT p.A78E H-taurine uptake by peripheral blood mononuclear cells was reduced by 95%, and taurine levels were severely reduced in plasma, skeletal muscle, and brain. Extraocular dysfunctions were not yet detected, but significantly increased urinary excretion of 8-oxo-7,8-dihydroguanosine indicated generally enhanced (yet clinically unapparent) oxidative stress and RNA oxidation, warranting continuous broad surveillance.-Preising, M. N., Görg, B., Friedburg, C., Qvartskhava, N., Budde, B. S., Bonus, M., Toliat, M. R., Pfleger, C., Altmüller, J., Herebian, D., Beyer, M., Zöllner, H. J., Wittsack, H.-J., Schaper, J., Klee, D., Zechner, U., Nürnberg, P., Schipper, J., Schnitzler, A., Gohlke, H., Lorenz, B., Häussinger, D., Bolz, H. J. Biallelic mutation of human 3 H-taurine uptake by peripheral blood mononuclear cells was reduced by 95%, and taurine levels were severely reduced in plasma, skeletal muscle, and brain. Extraocular dysfunctions were not yet detected, but significantly increased urinary excretion of 8-oxo-7,8-dihydroguanosine indicated generally enhanced (yet clinically unapparent) oxidative stress and RNA oxidation, warranting continuous broad surveillance.-Preising, M. N., Görg, B., Friedburg, C., Qvartskhava, N., Budde, B. S., Bonus, M., Toliat, M. R., Pfleger, C., Altmüller, J., Herebian, D., Beyer, M., Zöllner, H. J., Wittsack, H.-J., Schaper, J., Klee, D., Zechner, U., Nürnberg, P., Schipper, J., Schnitzler, A., Gohlke, H., Lorenz, B., Häussinger, D., Bolz, H. J. Biallelic mutation of human SLC6A6 encoding the taurine transporter TAUT is linked to early retinal degeneration.

To uncover the genotype underlying early-onset cone-rod dystrophy and central nummular macular atrophic lesion in 2 siblings from an endogamous Arab family, we performed targeted next-generation sequencing (NGS) of 44 retinal dystrophy genes, whole-exome sequencing (WES) and genome-wide linkage analysis. Targeted NGS and WES in the index patient highlighted 2 homozygous variants, a CCDC66 frameshift deletion and a novel missense NMNAT1 variant, c.500G>A (p.Asn167Ser). Linkage and segregation analysis excluded the CCDC66 variant and confirmed the NMNAT1 mutation. Biallelic NMNAT1 mutations cause Leber congenital amaurosis with a central nummular macular atrophic lesion (LCA9). The NMNAT1 mutation reported here underlied cone-rod dystrophy rather than LCA but the fundus lesion was compatible with that of LCA9 patients, highlighting that such a fundus appearance should raise suspicion for biallelic mutations in NMNAT1 when in the context of any retinal dystrophy. Although Ccdc66 mutations have been proposed to cause retinal disease in dogs, our results and public databases challenge CCDC66 as a candidate gene for human retinal dystrophy., (© 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

We describe a large family with disproportionate short stature and bone dysplasia from Nias in which we observed differences in severity when comparing the phenotypes of affected individuals from two remote branches. We conducted a linkage scan in the more severely affected family branch and determined a critical interval of 4.7 cM on chromosome 11. Sequencing of the primary candidate gene TBX10 did not reveal a disease-causing variant. When performing whole exome sequencing we noticed a homozygous missense variant in B3GAT3, c.419C>T [p.(Pro140Leu)]. B3GAT3 encodes β-1,3-glucuronyltransferase-I (GlcAT-I). GlcAT-I catalyzes an initial step of proteoglycan synthesis and the mutation p. (Pro140Leu) lies within the donor substrate-binding subdomain of the catalytic domain. In contrast to the previously published mutation in B3GAT3, c.830G>A [p.(Arg277Gln)], no heart phenotype could be detected in our family. Functional studies revealed a markedly reduced GlcAT-I activity in lymphoblastoid cells from patients when compared to matched controls. Moreover, relative numbers of glycosaminoglycan (GAG) side chains were decreased in patient cells. We found that Pro140Leu-mutant GlcAT-I cannot efficiently transfer GlcA to the linker region trisaccharide. This failure results in a partial deficiency of both chondroitin sulfate and heparan sulfate chains. Since the phenotype of the Nias patients differs from the Larsen-like syndrome described for patients with mutation p.(Arg277Gln), we suggest mutation B3GAT3:p.(Pro140Leu) to cause a different type of GAG linkeropathy showing no involvement of the heart.

Background: Grebe dysplasia, Hunter-Thompson dysplasia, and du Pan dysplasia constitute a spectrum of skeletal dysplasias inherited as an autosomal recessive trait characterized by short stature, severe acromesomelic shortening of the limbs, and normal axial skeleton. The majority of patients with these disorders have biallelic loss-of-function mutations of GDF5. In single instances, Grebe dysplasia and a Grebe dysplasia-like phenotype with genital anomalies have been shown to be caused by mutations in BMPR1B, encoding a GDF5 receptor., Methods: We clinically and radiologically characterised an acromesomelic chondrodysplasia in an adult woman born to consanguineous parents. We sequenced GDF5 and BMPR1B on DNA of the proposita. We performed 3D structural analysis and luciferase reporter assays to functionally investigate the identified BMPR1B mutation., Results: We extend the genotype-phenotype correlation in the acromesomelic chondrodysplasias by showing that the milder du Pan dysplasia can be caused by a hypomorphic BMPR1B mutation. We show that the homozygous c.91C>T, p.(Arg31Cys) mutation causing du Pan dysplasia leads to a significant loss of BMPR1B function, but to a lesser extent than the previously reported p.Cys53Arg mutation that results in the more severe Grebe dysplasia., Conclusions: The phenotypic severity gradient of the clinically and radiologically related acromesomelic chondrodysplasia spectrum of skeletal disorders may be due to the extent of functional impairment of the ligand-receptor pair GDF5-BMPR1B.

Abstract Targeted re-sequencing such as gene panel sequencing (GPS) has become very popular in medical genetics, both for research projects and in diagnostic settings. The technical principles of the different enrichment methods have been reviewed several times before; however, new enrichment products are constantly entering the market, and researchers are often puzzled about the requirement to take decisions about long-term commitments, both for the enrichment product and the sequencing technology. This review summarizes important considerations for the experimental design and provides helpful recommendations in choosing the best sequencing strategy for various research projects and diagnostic applications.

Aims: The Schubert-Bornschein type of complete congenital stationary night blindness (CSNB) is a genetically heterogeneous retinal disorder. It is characterised by a non-progressive disease course, often associated with high myopia and nystagmus. So far, mutations in two genes, NYX (nyctalopin) and GRM6 (metabotropic glutamate receptor 6) have been associated with this form of CSNB. The purpose of this study was to identify the genetic defect in affected male patients from Flemish families with complete CSNB., Methods: Probands with CSNB from three large Flemish families underwent ophthalmological examination. DNA was extracted from peripheral blood, and the coding region of NYX along with parts of the 5'UTR and 3'UTR and intronic regions covering the splice sites were PCR amplified and sequenced., Results: In the affected individuals of three Flemish families with the complete form of CSNB a novel NYX mutation, c.855delG was identified. This deletion is predicted to lead to a frameshift mutation, p.Asp286ThrfsX62 causing a premature stop codon., Conclusion: Previously, both single families with different mutations in NYX as well as different families with an identical mutation, suggestive of a founder mutation, have been described. The c.855delG deletion in NYX seems to be a common mutation associated with CSNB in the Flemish population from Belgium. Thus, we suggest performing diagnostic testing for CSNB in the Flemish population initially directed towards the identification of this mutation. Subsequent screening for other mutations in NYX or GRM6 could be performed as a second step.

Pontocerebellar hypoplasias (PCH) represent a group of neurodegenerative autosomal recessive disorders with prenatal onset, atrophy or hypoplasia of the cerebellum, hypoplasia of the ventral pons, microcephaly, variable neocortical atrophy and severe mental and motor impairments. In two subtypes, PCH2 and PCH4, we identified mutations in three of the four different subunits of the tRNA-splicing endonuclease complex. Our findings point to RNA processing as a new basic cellular impairment in neurological disorders.

Excessive skin wrinkling and cutis laxa are seen in many genetic conditions and overlapping features can make a clinical diagnosis difficult. Here we report on 22 Omani patients from 11 consanguineous families with the diagnosis of wrinkly skin syndrome (WSS, OMIM 278250) or geroderma osteodysplasticum hereditaria (GO, OMIM 231070). The WSS phenotype evolves during early childhood and includes a generalized and excessive skin wrinkling, dental problems, herniae, foot deformities, hip dislocations, growth retardation, and a large anterior fontanelle. The facial gestalt is characterized by a broad nasal bridge, hypertelorism, and downslanting palpebral fissures. We were unable to differentiate between WSS and cutis laxa with growth and developmental delay (CLGDD, OMIM 219200) suggesting that both can be considered as one entity. Distinct hallmarks of GO were skin wrinkling limited to the dorsum of hands and feet and to the abdomen, normal fontanelles, maxillary hypoplasia, bowed long bones, and osteopenia with frequent fractures. In contrast to the attenuation of the skin phenotype with age in WSS, adult patients with GO appeared prematurely aged. A serum sialotransferrin type 2 pattern was found in all four WSS patients tested. Apolipoprotein CIII (a marker for O-glycosylation) was normal suggesting that WSS is frequently associated with a N-protein glycosylation defect, probably at the level of processing (CDG-II). All four investigated GO patients showed normal sialotransferrin patterns. The known loci for cutis laxa and WSS on 2q31, 5q23-q31, 7q11, 11q13, and 14q32 were excluded. We suggest that WSS and GO are distinct entities with overlapping features., (Copyright 2008 Wiley-Liss, Inc.)

Noncompaction of the ventricular myocardium (NVM) is the morphological hallmark of a rare familial or sporadic unclassified heart disease of heterogeneous origin. NVM results presumably from a congenital developmental error and has been traced back to single point mutations in various genes. The objective of this study was to determine the underlying genetic defect in a large German family suffering from NVM. Twenty four family members were clinically assessed using advanced imaging techniques. For molecular characterization, a genome-wide linkage analysis was undertaken and the disease locus was mapped to chromosome 14ptel-14q12. Subsequently, two genes of the disease interval, MYH6 and MYH7 (encoding the alpha- and beta-myosin heavy chain, respectively) were sequenced, leading to the identification of a previously unknown de novo missense mutation, c.842G>C, in the gene MYH7. The mutation affects a highly conserved amino acid in the myosin subfragment-1 (R281T). In silico simulations suggest that the mutation R281T prevents the formation of a salt bridge between residues R281 and D325, thereby destabilizing the myosin head. The mutation was exclusively present in morphologically affected family members. A few members of the family displayed NVM in combination with other heart defects, such as dislocation of the tricuspid valve (Ebstein's anomaly, EA) and atrial septal defect (ASD). A high degree of clinical variability was observed, ranging from the absence of symptoms in childhood to cardiac death in the third decade of life. The data presented in this report provide first evidence that a mutation in a sarcomeric protein can cause noncompaction of the ventricular myocardium.

Variation between inbred mice of susceptibility to experimental Trypanosoma cruzi infection has frequently been described, but the immunogenetic background is poorly understood. The outcross of the susceptible parental mouse strains C57BL/6 (B6) and DBA/2 (D2), B6D2F1 (F1) mice, is highly resistant to this parasite. In the present study we show by quantitative PCR that the increase of tissue parasitism during the early phase of infection is comparable up to day 11 between susceptible B6 and resistant F1 mice. A reduction of splenic parasite burdens occurs thereafter in both strains but is comparatively retarded in susceptible mice. Splenic microarchitecture is progressively disrupted with loss of follicles and B lymphocytes in B6 mice, but not in F1 mice. By genotyping of additional backcross offspring we corroborate our earlier findings that susceptibility maps to three loci on Chromosomes 5, 13 and 17. Analysis of gene expression of spleen cells from infected B6 and F1 mice with microarrays identifies about 0.3% of transcripts that are differentially expressed. Assuming that differential susceptibility is mediated by altered gene expression, we propose that the following differentially expressed transcripts from these loci are strong candidates for the observed phenotypic variation: H2-Ealpha, H2-D1, Ng23, Msh5 and Tubb5 from Chromosome 17; and Cxcl11, Bmp2k and Spp1 from Chromosome 5. Our results indicate that innate mechanisms are not of primary relevance to resistance of F1 mice to T. cruzi infection, and that differential susceptibility to experimental infection with this protozoan pathogen is not paralleled by extensive variation of the transcriptome.