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The solute carrier (SLC) superfamily encompasses >400 transmembrane transporters involved in the exchange of amino acids, nutrients, ions, metals, neurotransmitters and metabolites across biological membranes. SLCs are highly expressed in the mammalian brain; defects in nearly 100 unique SLC-encoding genes (OMIM: https://www.omim.org) are associated with rare Mendelian disorders including developmental and epileptic encephalopathy and severe neurodevelopmental disorders. Exome sequencing and family-based rare variant analyses on a cohort with neurodevelopmental disorders identified two siblings with developmental and epileptic encephalopathy and a shared deleterious homozygous splicing variant in SLC38A3. The gene encodes SNAT3, a sodium-coupled neutral amino acid transporter and a principal transporter of the amino acids asparagine, histidine, and glutamine, the latter being the precursor for the neurotransmitters GABA and glutamate. Additional subjects with a similar developmental and epileptic encephalopathy phenotype and biallelic predicted-damaging SLC38A3 variants were ascertained through GeneMatcher and collaborations with research and clinical molecular diagnostic laboratories. Untargeted metabolomic analysis was performed to identify novel metabolic biomarkers. Ten individuals from seven unrelated families from six different countries with deleterious biallelic variants in SLC38A3 were identified. Global developmental delay, intellectual disability, hypotonia, and absent speech were common features while microcephaly, epilepsy, and visual impairment were present in the majority. Epilepsy was drug-resistant in half. Metabolomic analysis revealed perturbations of glutamate, histidine, and nitrogen metabolism in plasma, urine, and CSF of selected subjects, potentially representing biomarkers of disease. Our data support the contention that SLC38A3 is a novel disease gene for developmental and epileptic encephalopathy and illuminate the likely pathophysiology of the disease as perturbations in glutamine homeostasis.

Au-Kline syndrome (AKS) is a neurodevelopmental disorder associated with multiple malformations and a characteristic facial gestalt. The first individuals ascertained carried de novo loss-of-function (LoF) variants in HNRNPK. Here, we report 32 individuals with AKS (26 previously unpublished), including 13 with de novo missense variants. We propose new clinical diagnostic criteria for AKS that differentiate it from the clinically overlapping Kabuki syndrome and describe a significant phenotypic expansion to include individuals with missense variants who present with subtle facial features and few or no malformations. Many gene-specific DNA methylation (DNAm) signatures have been identified for neurodevelopmental syndromes. Because HNRNPK has roles in chromatin and epigenetic regulation, we hypothesized that pathogenic variants in HNRNPK may be associated with a specific DNAm signature. Here, we report a unique DNAm signature for AKS due to LoF HNRNPK variants, distinct from controls and Kabuki syndrome. This DNAm signature is also identified in some individuals with de novo HNRNPK missense variants, confirming their pathogenicity and the phenotypic expansion of AKS to include more subtle phenotypes. Furthermore, we report that some individuals with missense variants have an "intermediate" DNAm signature that parallels their milder clinical presentation, suggesting the presence of an epi-genotype phenotype correlation. In summary, the AKS DNAm signature may help elucidate the underlying pathophysiology of AKS. This DNAm signature also effectively supported clinical syndrome delineation and is a valuable aid for variant interpretation in individuals where a clinical diagnosis of AKS is unclear, particularly for mild presentations. ispartof: AMERICAN JOURNAL OF HUMAN GENETICS vol:109 issue:10 pages:1867-1884 ispartof: location:United States status: published

The purpose of this study is to describe a Mendelian disorder of DNA damage repair. Phenotypic delineation of two families, one new and one previously published, with overlapping dysmorphic and neurodevelopmental features was undertaken. Functional characterization of DNA damage repair in fibroblasts obtained from the index individuals in each of the two families was pursued. We present new evidence of a distinct disorder caused by biallelic truncating variants in ZNF668 comprising microcephaly, growth deficiency, severe global developmental delay, brain malformation, and distinct facial dysmorphism. DNA damage repair defect was observed in fibroblasts of affected individuals. ZNF668 deficiency in humans results in a recognizable autosomal recessive disorder, which we propose to name ZNF668-related ZMAND (ZNF668-related brain malformation, microcephaly, abnormal growth, neurodevelopmental delay, and dysmorphism). Our results add to the growing list of Mendelian disorders of the DNA damage repair machinery.

Summary TP73 belongs to the TP53 family of transcription factors and has therefore been well studied in cancer research. Studies in mice, however, have revealed non-oncogenic activities related to multiciliogenesis. Utilizing whole-exome sequencing analysis in a cohort of individuals with a mucociliary clearance disorder and cortical malformation, we identified homozygous loss-of-function variants in TP73 in seven individuals from five unrelated families. All affected individuals exhibit a chronic airway disease as well as a brain malformation consistent with lissencephaly. We performed high-speed video microscopy, immunofluorescence analyses, and transmission electron microscopy in respiratory epithelial cells after spheroid or air liquid interface culture to analyze ciliary function, ciliary length, and number of multiciliated cells (MCCs). The respiratory epithelial cells studied display reduced ciliary length and basal bodies mislocalized within the cytoplasm. The number of MCCs is severely reduced, consistent with a reduced number of cells expressing the transcription factors crucial for multiciliogenesis (FOXJ1, RFX2). Our data demonstrate that autosomal-recessive deleterious variants in the TP53 family member TP73 cause a mucociliary clearance disorder due to a defect in MCC differentiation.

N-terminal acetyltransferases modify proteins by adding an acetyl moiety to the first amino acid and are vital for protein and cell function. The NatB complex acetylates 20% of the human proteome and is composed of the catalytic subunit NAA20 and the auxiliary subunit NAA25. In five individuals with overlapping phenotypes, we identified recessive homozygous missense variants in NAA20. Two different NAA20 variants were identified in affected individuals in two consanguineous families by exome and genome sequencing. Biochemical studies were employed to assess the impact of the NAA20 variants on NatB complex formation and catalytic activity. Two homozygous variants, NAA20 p.Met54Val and p.Ala80Val (GenBank: NM_016100.4, c.160A>G and c.239C>T), segregated with affected individuals in two unrelated families presenting with developmental delay, intellectual disability, and microcephaly. Both NAA20-M54V and NAA20-A80V were impaired in their capacity to form a NatB complex with NAA25, and in vitro acetylation assays revealed reduced catalytic activities toward different NatB substrates. Thus, both NAA20 variants are impaired in their ability to perform cellular NatB-mediated N-terminal acetylation. We present here a report of pathogenic NAA20 variants causing human disease and data supporting an essential role for NatB-mediated N-terminal acetylation in human development and physiology.

Purpose To identify novel genes associated with intellectual disability (ID) in four unrelated families. Methods Here, through exome sequencing and international collaboration, we report eight individuals from four unrelated families of diverse geographic origin with biallelic loss-of-function variants in UBE4A. Results Eight evaluated individuals presented with syndromic intellectual disability and global developmental delay. Other clinical features included hypotonia, short stature, seizures, and behavior disorder. Characteristic features were appreciated in some individuals but not all; in some cases, features became more apparent with age. We demonstrated that UBE4A loss-of-function variants reduced RNA expression and protein levels in clinical samples. Mice generated to mimic patient-specific Ube4a loss-of-function variant exhibited muscular and neurological/behavioral abnormalities, some of which are suggestive of the clinical abnormalities seen in the affected individuals. Conclusion These data indicate that biallelic loss-of-function variants in UBE4A cause a novel intellectual disability syndrome, suggesting that UBE4A enzyme activity is required for normal development and neurological function.

Rhabdomyolysis is a serious medical condition characterized by muscle injury, and there are recognized genetic causes especially in recurrent forms. The majority of these cases, however, remain unexplained. Here, we describe a patient with recurrent rhabdomyolysis in whom extensive clinical testing failed to identify a likely etiology. Whole-exome sequencing revealed a novel missense variant in MYH1, which encodes a major adult muscle fiber protein. Structural biology analysis revealed that the mutated residue is extremely well conserved and is located in the actin binding cleft. Furthermore, immediately adjacent mutations in that cleft in other myosins are pathogenic in humans. Our results are consistent with the finding that MYH1 is mutated in rhabdomyolysis in horses and suggest that this gene should be investigated in cases with recurrent rhabdomyolysis.

Contains fulltext : 283128.pdf (Publisher’s version ) (Open Access) Developmental and epileptic encephalopathy 35 (DEE 35) is a severe neurological condition caused by biallelic variants in ITPA, encoding inosine triphosphate pyrophosphatase, an essential enzyme in purine metabolism. We delineate the genotypic and phenotypic spectrum of DEE 35, analyzing possible predictors for adverse clinical outcomes. We investigated a cohort of 28 new patients and reviewed previously described cases, providing a comprehensive characterization of 40 subjects. Exome sequencing was performed to identify underlying ITPA pathogenic variants. Brain MRI (magnetic resonance imaging) scans were systematically analyzed to delineate the neuroradiological spectrum. Survival curves according to the Kaplan-Meier method and log-rank test were used to investigate outcome predictors in different subgroups of patients. We identified 18 distinct ITPA pathogenic variants, including 14 novel variants, and two deletions. All subjects showed profound developmental delay, microcephaly, and refractory epilepsy followed by neurodevelopmental regression. Brain MRI revision revealed a recurrent pattern of delayed myelination and restricted diffusion of early myelinating structures. Congenital microcephaly and cardiac involvement were statistically significant novel clinical predictors of adverse outcomes. We refined the molecular, clinical, and neuroradiological characterization of ITPase deficiency, and identified new clinical predictors which may have a potentially important impact on diagnosis, counseling, and follow-up of affected individuals.

Major Facilitator Superfamily Domain containing 2a (MFSD2A) is an essential endothelial lipid transporter at the blood-brain barrier. Biallelic variants affecting function in MFSD2A cause autosomal recessive primary microcephaly 15 (MCPH15, OMIM# 616486). We sought to expand our knowledge of the phenotypic spectrum of MCPH15 and demonstrate the underlying mechanism of inactivation of the MFSD2A transporter. We carried out detailed analysis of the clinical and neuroradiological features of a series of 27 MCPH15 cases, including eight new individuals from seven unrelated families. Genetic investigation was performed through exome sequencing (ES). Structural insights on the human Mfsd2a model and in-vitro biochemical assays were used to investigate the functional impact of the identified variants. All patients had primary microcephaly and severe developmental delay. Brain MRI showed variable degrees of white matter reduction, ventricular enlargement, callosal hypodysgenesis, and pontine and vermian hypoplasia. ES led to the identification of six novel biallelic MFSD2A variants (NG_053084.1, NM_032793.5: c.556+1G>A, c.748G>T; p.(Val250Phe), c.750_753del; p.(Cys251SerfsTer3), c.977G>A; p.(Arg326His), c.1386_1435del; p.(Gln462HisfsTer17), and c.1478C>T; p.(Pro493Leu)) and two recurrent variants (NM_032793.5: c.593C>T; p.(Thr198Met) and c.476C>T; p.(Thr159Met)). All these variants and the previously reported NM_032793.5: c.490C>A; p.(Pro164Thr) resulted in either reduced MFSD2A expression and/or transport activity. Our study further delineates the phenotypic spectrum of MCPH15, refining its clinical and neuroradiological characterization and supporting that MFSD2A deficiency causes early prenatal brain developmental disruption. We also show that poor MFSD2A expression despite normal transporter activity is a relevant pathomechanism in MCPH15.

In mammals, a subset of arginine tRNA isoacceptors are methylated in the anticodon loop by the METTL2 methyltransferase to form the 3-methylcytosine (m3C) modification. However, the mechanism by which METTL2 identifies specific tRNA arginine species for m3C formation as well as the biological role of m3C in mammals is unknown. Here, we show that human METTL2 forms a complex with DALR anticodon binding domain containing 3 (DALRD3) protein to recognize particular arginine tRNAs destined for m3C modification. DALRD3-deficient human cells exhibit nearly complete loss of the m3C modification in tRNA-Arg species. Notably, we identify a homozygous nonsense mutation in the DALRD3 gene that impairs m3C formation in human patients exhibiting developmental delay and early-onset epileptic encephalopathy. These findings uncover an unexpected function for the DALRD3 protein in the targeting of distinct arginine tRNAs for m3C modification and suggest a crucial biological role for DALRD3-dependent tRNA modification in proper neurological development., METTL2 methyltransferase is responsible for 3-methylcytosine modification of arginine tRNAs in mammals. Here the authors show that DALR anticodon binding domain containing 3 (DALRD3) forms a complex with METTL2 to recognize specific arginine tRNAs and find DALRD3 mutations in patients with developmental delay and early-onset epileptic encephalopathy.

Arthrogryposis multiplex congenita (AMC) is an important birth defect with a significant genetic contribution. Many syndromic forms of AMC have been described, but remain unsolved at the molecular level. In this report, we describe a novel syndromic form of AMC in two multiplex consanguineous families from Saudi Arabia and Oman. The phenotype is highly consistent, and comprises neurogenic arthrogryposis, microcephaly, brain malformation (absent corpus callosum), optic atrophy, limb fractures, profound global developmental delay, and early lethality. Whole-exome sequencing revealed a different homozygous truncating variant in SCYL2 in each of the two families. SCYL2 is a component of clathrin-coated vesicles, and deficiency of its mouse ortholog results in a severe neurological phenotype that largely recapitulates the phenotype observed in our patients. Our results suggest that severe neurogenic arthrogryposis with brain malformation is the human phenotypic consequence of SCYL2 loss of function mutations.

MEDNIK syndrome (mental retardation, enteropathy, deafness, peripheral neuropathy, ichthyosis, and keratoderma) is an autosomal-recessive disorder caused by bi-allelic mutations in AP1S1, encoding the small σ subunit of the AP-1 complex. Central to the pathogenesis of MEDNIK syndrome is abnormal AP-1-mediated trafficking of copper transporters; this abnormal trafficking results in a hybrid phenotype combining the copper-deficiency-related characteristics of Menkes disease and the copper-toxicity-related characteristics of Wilson disease. We describe three individuals from two unrelated families in whom a MEDNIK-like phenotype segregates with two homozygous null variants in AP1B1, encoding the large β subunit of the AP-1 complex. Similar to individuals with MEDNIK syndrome, the affected individuals we report display abnormal copper metabolism, evidenced by low plasma copper and ceruloplasmin, but lack evidence of copper toxicity in the liver. Functional characterization of fibroblasts derived from affected individuals closely resembles the abnormal ATP7A trafficking described in MEDNIK syndrome both at baseline and in response to copper treatment. Taken together, our results expand the list of inborn errors of copper metabolism.

Summary BCAS3 microtubule-associated cell migration factor (BCAS3) is a large, highly conserved cytoskeletal protein previously proposed to be critical in angiogenesis and implicated in human embryogenesis and tumorigenesis. Here, we established BCAS3 loss-of-function variants as causative for a neurodevelopmental disorder. We report 15 individuals from eight unrelated families with germline bi-allelic loss-of-function variants in BCAS3. All probands share a global developmental delay accompanied by pyramidal tract involvement, microcephaly, short stature, strabismus, dysmorphic facial features, and seizures. The human phenotype is less severe compared with the Bcas3 knockout mouse model and cannot be explained by angiogenic defects alone. Consistent with being loss-of-function alleles, we observed absence of BCAS3 in probands’ primary fibroblasts. By comparing the transcriptomic and proteomic data based on probands’ fibroblasts with those of the knockout mouse model, we identified similar dysregulated pathways resulting from over-representation analysis, while the dysregulation of some proposed key interactors could not be confirmed. Together with the results from a tissue-specific Drosophila loss-of-function model, we demonstrate a vital role for BCAS3 in neural tissue development.

Neurodevelopmental disorders (NDD) are genetically and phenotypically heterogeneous conditions due to defects in genes involved in development and function of the nervous system. Individuals with NDD, in addition to their primary neurodevelopmental phenotype, may also have accompanying syndromic features that can be very helpful diagnostically especially those with recognizable facial appearance. In this study, we describe ten similarly affected individuals from six unrelated families of different ethnic origins having bi-allelic truncating variants in TMEM94, which encodes for an uncharacterized transmembrane nuclear protein that is highly conserved across mammals. The affected individuals manifested with global developmental delay/intellectual disability, and dysmorphic facial features including triangular face, deep set eyes, broad nasal root and tip and anteverted nostrils, thick arched eye brows, hypertrichosis, pointed chin, and hypertelorism. Birthweight in the upper normal range was observed in most, and all but one had congenital heart defects (CHD). Gene expression analysis in available cells from affected individuals showed reduced expression of TMEM94. Global transcriptome profiling using microarray and RNA sequencing revealed several dysregulated genes essential for cell growth, proliferation and survival that are predicted to have an impact on cardiotoxicity hematological system and neurodevelopment. Loss of Tmem94 in mouse model generated by CRISPR/Cas9 was embryonic lethal and led to craniofacial and cardiac abnormalities and abnormal neuronal migration pattern, suggesting that this gene is important in craniofacial, cardiovascular, and nervous system development. Our study suggests the genetic etiology of a recognizable dysmorphic syndrome with NDD and CHD and highlights the role of TMEM94 in early development.

Autophagy is the major intracellular degradation route in mammalian cells. Systemic ablation of core autophagy-related (ATG) genes in mice leads to embryonic or perinatal lethality, and conditional models show neurodegeneration. Impaired autophagy has been associated with a range of complex human diseases, yet congenital autophagy disorders are rare.We performed a genetic, clinical, and neuroimaging analysis involving five families. Mechanistic investigations were conducted with the use of patient-derived fibroblasts, skeletal muscle-biopsy specimens, mouse embryonic fibroblasts, and yeast.We found deleterious, recessive variants in human ATG7, a core autophagy-related gene encoding a protein that is indispensable to classical degradative autophagy. Twelve patients from five families with distinct ATG7 variants had complex neurodevelopmental disorders with brain, muscle, and endocrine involvement. Patients had abnormalities of the cerebellum and corpus callosum and various degrees of facial dysmorphism. These patients have survived with impaired autophagic flux arising from a diminishment or absence of ATG7 protein. Although autophagic sequestration was markedly reduced, evidence of basal autophagy was readily identified in fibroblasts and skeletal muscle with loss of ATG7. Complementation of different model systems by deleterious ATG7 variants resulted in poor or absent autophagic function as compared with the reintroduction of wild-type ATG7.We identified several patients with a neurodevelopmental disorder who have survived with a severe loss or complete absence of ATG7, an essential effector enzyme for autophagy without a known functional paralogue. (Funded by the Wellcome Centre for Mitochondrial Research and others.).

Autism spectrum disorder (ASD) is a constellation of neurodevelopmental disorders with high phenotypic and genetic heterogeneity, complicating the discovery of causative genes. Through a forward genetics approach selecting for defective vocalization in mice, we identified Kdm5a as a candidate ASD gene. To validate our discovery, we generated a Kdm5a knockout mouse model (Kdm5a-/-) and confirmed that inactivating Kdm5a disrupts vocalization. In addition, Kdm5a-/- mice displayed repetitive behaviors, sociability deficits, cognitive dysfunction, and abnormal dendritic morphogenesis. Loss of KDM5A also resulted in dysregulation of the hippocampal transcriptome. To determine if KDM5A mutations cause ASD in humans, we screened whole exome sequencing and microarray data from a clinical cohort. We identified pathogenic KDM5A variants in nine patients with ASD and lack of speech. Our findings illustrate the power and efficacy of forward genetics in identifying ASD genes and highlight the importance of KDM5A in normal brain development and function.

PURPOSE: To investigate the effect of PLXNA1 variants on the phenotype of patients with autosomal dominant and recessive inheritance patterns and to functionally characterize the zebrafish homologs plxna1a and plxna1b during development. METHODS: We assembled ten patients from seven families with biallelic or de novo PLXNA1 variants. We describe genotype-phenotype correlations, investigated the variants by structural modeling, and used Morpholino knockdown experiments in zebrafish to characterize the embryonic role of plxna1a and plxna1b. RESULTS: Shared phenotypic features among patients include global developmental delay (9/10), brain anomalies (6/10), and eye anomalies (7/10). Notably, seizures were predominantly reported in patients with monoallelic variants. Structural modeling of missense variants in PLXNA1 suggests distortion in the native protein. Our zebrafish studies enforce an embryonic role of plxna1a and plxna1b in the development of the central nervous system and the eye. CONCLUSION: We propose that different biallelic and monoallelic variants in PLXNA1 result in a novel neurodevelopmental syndrome mainly comprising developmental delay, brain, and eye anomalies. We hypothesize that biallelic variants in the extracellular Plexin-A1 domains lead to impaired dimerization or lack of receptor molecules, whereas monoallelic variants in the intracellular Plexin-A1 domains might impair downstream signaling through a dominant-negative effect.

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.

Recently, the genetic cause of HIDEA syndrome (hypotonia, hypoventilation, intellectual disability, dysautonomia, epilepsy, and eye abnormalities) was identified as biallelic pathogenic variants in P4HTM, which encodes an atypical member of the prolyl 4-hydroxylases (P4Hs) family of enzymes. We report seven patients from four new families in whom HIDEA was only diagnosed after whole-exome sequencing (WES) revealed novel disease-causing variants in P4HTM. We note the variable phenotypic expressivity of the syndrome except for cognitive impairment/developmental delay, and hypotonia, which seem to be consistent findings. One patient only presented with hypotonia, developmental delay, and abnormal eye movements, which highlights the challenge in diagnosing milder cases with this new syndrome. Other notable features include mild facial dysmorphism, obesity, and brain dysmyelination and atrophy. We conclude that HIDEA is a highly variable syndrome and suspect that a large fraction of patients will be diagnosed via reverse phenotyping after recessive P4HTM variants are identified by agnostic genomic sequencing assays.

Additional file 6. Figure S1. Pedigree of a family which is mapped to a single locus and identified a very deep (+ 335) variant in ABCB4 causing cholestatic disease. Figure S2. A) A sashimi plot showing base-level densities of reads mapped to a genomic region containing exons 11 and 12 of KCTD3 transcripts from three samples. The x-axis represents the genomic coordinate in hg19. The y-axis represents per-base read counts, and the range is specified in the upper-left corner of the plot for each sample. Arcs connecting exons represent splice junction reads. The horizontal bar lines on the bottom indicate isoforms (exons as rectangle boxes and introns as line with arrow heads). The distribution in blue shows the sample with aberrant KCTD3 transcript, while the other two distributions are from randomly selected samples of lymphocytes (red from a patient and green from the GTEx cohort). B) A sashimi plot showing base-level densities of reads mapped to a genomic region in the positive control cases.

Additional file 1. Supplemental file 1. Causal gene prediction comparison for RNA-Seq data.

Pediatric cataract is highly heterogeneous clinically and etiologically. While mostly isolated, cataract can be part of many multisystem disorders, further complicating the diagnostic process. In this study, we applied genomic tools in the form of a multi-gene panel as well as whole-exome sequencing on unselected cohort of pediatric cataract (166 patients from 74 families). Mutations in previously reported cataract genes were identified in 58% for a total of 43 mutations, including 15 that are novel. GEMIN4 was independently mutated in families with a syndrome of cataract, global developmental delay with or without renal involvement. We also highlight a recognizable syndrome that resembles galactosemia (a fulminant infantile liver disease with cataract) caused by biallelic mutations in CYP51A1. A founder mutation in RIC1 (KIAA1432) was identified in patients with cataract, brain atrophy, microcephaly with or without cleft lip and palate. For non-syndromic pediatric cataract, we map a novel locus in a multiplex consanguineous family on 4p15.32 where exome sequencing revealed a homozygous truncating mutation in TAPT1. We report two further candidates that are biallelically inactivated each in a single cataract family: TAF1A (cataract with global developmental delay) and WDR87 (non-syndromic cataract). In addition to positional mapping data, we use iSyTE developmental lens expression and gene-network analysis to corroborate the proposed link between the novel candidate genes and cataract. Our study expands the phenotypic, allelic and locus heterogeneity of pediatric cataract. The high diagnostic yield of clinical genomics supports the adoption of this approach in this patient group.

We report four unrelated children with homozygous loss-of-function variants in TASP1 and an overlapping phenotype comprising developmental delay with hypotonia and microcephaly, feeding difficulties with failure-to-thrive, recurrent respiratory infections, cardiovascular malformations, cryptorchidism, happy demeanor, and distinctive facial features. Two children had a homozygous founder deletion encompassing exons 5–11 of TASP1, the third had a homozygous missense variant, c.701 C>T (p.Thr234Met), affecting the active site of the encoded enzyme, and the fourth had a homozygous nonsense variant, c.199 C>T (p.Arg67*). TASP1 encodes taspase 1 (TASP1), which is responsible for cleaving, thus activating, the lysine methyltransferases KMT2A and KMT2D, which are essential for histone methylation and transcription regulation. The consistency of the phenotype, the critical biological function of TASP1, the deleterious nature of the TASP1 variants, and the overlapping features with Wiedemann–Steiner and Kabuki syndromes respectively caused by pathogenic variants in KMT2A and KMT2D all support that TASP1 is a disease-related gene.

Genetic testing in pediatric cholestasis can be very informative but genetic causes have not been fully characterized. Exome sequencing and positional mapping in seven families with cholestatic liver disease and negative clinical testing for known disease genes. KIF12, which encodes a microtubule motor protein with a tentative role in cell polarity, was found to harbor three homozygous likely deleterious variants in three families with sclerosing cholangitis. KIF12 expression is dependent on HNF-1β, deficiency which is known to cause bile duct dysmorphogenesis associated with loss of KIF12 expression. In another extended family, we mapped an apparently novel syndrome of sclerosing cholangitis, short stature, hypothyroidism, and abnormal tongue pigmentation in two cousins to a homozygous variant in PPM1F (POPX2), a regulator of kinesin-mediated ciliary transport. In the fifth family, a syndrome of normal gamma glutamyltransferase (GGT) cholestasis and hearing loss was found to segregate with a homozygous truncating variant in USP53, which encodes an interactor with TJP2. In the sixth family, we mapped a novel syndrome of transient neonatal cholestasis, intellectual disability, and short stature to a homozygous variant in LSR, an important regulator of liver development. In the last family of three affected siblings, a novel syndrome of intractable itching, hypercholanemia, short stature, and intellectual disability was mapped to a single locus that contains a homozygous truncating variant in WDR83OS (C19orf56), known to interact with ATP13A2 and BSEP. Our results expand the genetic heterogeneity of pediatric cholestatic liver disease and highlight the vulnerability of bile homeostasis to a wide range of molecular perturbations.

Purpose Establishing links between Mendelian phenotypes and genes enables the proper interpretation of variants therein. Autozygome, a rich source of homozygous variants, has been successfully utilized for the high throughput identification of novel autosomal recessive disease genes. Here, we highlight the utility of the autozygome for the high throughput confirmation of previously published tentative links to diseases. Methods Autozygome and exome analysis of patients with suspected Mendelian phenotypes. All variants were classified according to the American College of Medical Genetics and Genomics guidelines. Results We highlight 30 published candidate genes (ACTL6B, ADAM22, AGTPBP1, APC, C12orf4, C3orf17 (NEPRO), CENPF, CNPY3, COL27A1, DMBX1, FUT8, GOLGA2, KIAA0556, LENG8, MCIDAS, MTMR9, MYH11, QRSL1, RUBCN, SLC25A42, SLC9A1, TBXT, TFG, THUMPD1, TRAF3IP2, UFC1, UFM1, WDR81, XRCC2, ZAK) in which we identified homozygous likely deleterious variants in patients with compatible phenotypes. We also identified homozygous likely deleterious variants in 18 published candidate genes (ABCA2, ARL6IP1, ATP8A2, CDK9, CNKSR1, DGAT1, DMXL2, GEMIN4, HCN2, HCRT, MYO9A, PARS2, PLOD3, PREPL, SCLT1, STX3, TXNRD2, WIPI2) although the associated phenotypes are sufficiently different from the original reports that they represent phenotypic expansion or potentially distinct allelic disorders. Conclusions Our results should facilitate the timely relabeling of these candidate disease genes in relevant databases to improve the yield of clinical genomic sequencing.

Infertility is a relatively common disorder of the reproductive system and remains unexplained in many cases. In vitro fertilization techniques have uncovered previously unrecognized infertility phenotypes, including oocyte maturation arrest, the molecular etiology of which remains largely unknown. We report two families affected by female-limited infertility caused by oocyte maturation failure. Positional mapping and whole-exome sequencing revealed two homozygous, likely deleterious variants in PATL2, each of which fully segregates with the phenotype within the respective family. PATL2 encodes a highly conserved oocyte-specific mRNP repressor of translation. Previous data have shown the strict requirement for PATL2 in oocyte-maturation in model organisms. Data gathered from the families in this study suggest that the role of PATL2 is conserved in humans and expand our knowledge of the factors that are necessary for female meiosis.

To describe our experience with a large cohort (411 patients from 288 families) of various forms of skeletal dysplasia who were molecularly characterized. Detailed phenotyping and next-generation sequencing (panel and exome). Our analysis revealed 224 pathogenic/likely pathogenic variants (54 (24%) of which are novel) in 123 genes with established or tentative links to skeletal dysplasia. In addition, we propose 5 genes as candidate disease genes with suggestive biological links (WNT3A, SUCO, RIN1, DIP2C, and PAN2). Phenotypically, we note that our cohort spans 36 established phenotypic categories by the International Skeletal Dysplasia Nosology, as well as 18 novel skeletal dysplasia phenotypes that could not be classified under these categories, e.g., the novel C3orf17-related skeletal dysplasia. We also describe novel phenotypic aspects of well-known disease genes, e.g., PGAP3-related Toriello–Carey syndrome–like phenotype. We note a strong founder effect for many genes in our cohort, which allowed us to calculate a minimum disease burden for the autosomal recessive forms of skeletal dysplasia in our population (7.16E-04), which is much higher than the global average. By expanding the phenotypic, allelic, and locus heterogeneity of skeletal dysplasia in humans, we hope our study will improve the diagnostic rate of patients with these conditions.

To ensure efficient genome duplication, cells have evolved numerous factors that promote unperturbed DNA replication and protect, repair and restart damaged forks. Here we identify downstream neighbor of SON (DONSON) as a novel fork protection factor and report biallelic DONSON mutations in 29 individuals with microcephalic dwarfism. We demonstrate that DONSON is a replisome component that stabilizes forks during genome replication. Loss of DONSON leads to severe replication-associated DNA damage arising from nucleolytic cleavage of stalled replication forks. Furthermore, ATM- and Rad3-related (ATR)-dependent signaling in response to replication stress is impaired in DONSON-deficient cells, resulting in decreased checkpoint activity and the potentiation of chromosomal instability. Hypomorphic mutations in DONSON substantially reduce DONSON protein levels and impair fork stability in cells from patients, consistent with defective DNA replication underlying the disease phenotype. In summary, we have identified mutations in DONSON as a common cause of microcephalic dwarfism and established DONSON as a critical replication fork protein required for mammalian DNA replication and genome stability.

Dominant SCN1B mutations are known to cause several epilepsy syndromes in humans. Only 2 epilepsy patients to date have been reported to have recessive mutations in SCN1B as the likely cause of their phenotype. Here, we confirm the recessive inheritance of 2 novel SCN1B mutations in 5 children from 3 families with developmental epileptic encephalopathy. The recessive inheritance and early death in these patients is consistent with the Dravet-like phenotype observed in Scn1b-/- mice. The 'negative' clinical exome in one of these families highlights the need to consider recessive mutations in the interpretation of variants in typically dominant genes.

Intellectual disability (ID) is a measurable phenotypic consequence of genetic and environmental factors. In this study, we prospectively assessed the diagnostic yield of genomic tools (molecular karyotyping, multi-gene panel and exome sequencing) in a cohort of 337 ID subjects as a first-tier test and compared it with a standard clinical evaluation performed in parallel. Standard clinical evaluation suggested a diagnosis in 16% of cases (54/337) but only 70% of these (38/54) were subsequently confirmed. On the other hand, the genomic approach revealed a likely diagnosis in 58% (n=196). These included copy number variants in 14% (n=54, 15% are novel), and point mutations revealed by multi-gene panel and exome sequencing in the remaining 43% (1% were found to have Fragile-X). The identified point mutations were mostly recessive (n=117, 81%), consistent with the high consanguinity of the study cohort, but also X-linked (n=8, 6%) and de novo dominant (n=19, 13%). When applied directly on all cases with negative molecular karyotyping, the diagnostic yield of exome sequencing was 60% (77/129). Exome sequencing also identified likely pathogenic variants in three novel candidate genes (DENND5A, NEMF and DNHD1) each of which harbored independent homozygous mutations in patients with overlapping phenotypes. In addition, exome sequencing revealed de novo and recessive variants in 32 genes (MAMDC2, TUBAL3, CPNE6, KLHL24, USP2, PIP5K1A, UBE4A, TP53TG5, ATOH1, C16ORF90, SLC39A14, TRERF1, RGL1, CDH11, SYDE2, HIRA, FEZF2, PROCA1, PIANP, PLK2, QRFPR, AP3B2, NUDT2, UFC1, BTN3A2, TADA1, ARFGEF3, FAM160B1, ZMYM5, SLC45A1, ARHGAP33 and CAPS2), which we highlight as potential candidates on the basis of several lines of evidence, and one of these genes (SLC39A14) was biallelically inactivated in a potentially treatable form of hypermanganesemia and neurodegeneration. Finally, likely causal variants in previously published candidate genes were identified (ASTN1, HELZ, THOC6, WDR45B, ADRA2B and CLIP1), thus supporting their involvement in ID pathogenesis. Our results expand the morbid genome of ID and support the adoption of genomics as a first-tier test for individuals with ID.