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BackgroundO’Donnell-Luria-Rodan syndrome (ODLURO) is an autosomal-dominant neurodevelopmental disorder caused by pathogenic, mostly truncating variants in KMT2E. It was first described by O’Donnell-Luria et al in 2019 in a cohort of 38 patients. Clinical features encompass macrocephaly, mild intellectual disability (ID), autism spectrum disorder (ASD) susceptibility and seizure susceptibility.MethodsAffected individuals were ascertained at paediatric and genetic centres in various countries by diagnostic chromosome microarray or exome/genome sequencing. Patients were collected into a case cohort and were systematically phenotyped where possible.ResultsWe report 18 additional patients from 17 families with genetically confirmed ODLURO. We identified 15 different heterozygous likely pathogenic or pathogenic sequence variants (14 novel) and two partial microdeletions of KMT2E. We confirm and refine the phenotypic spectrum of the KMT2E-related neurodevelopmental disorder, especially concerning cognitive development, with rather mild ID and macrocephaly with subtle facial features in most patients. We observe a high prevalence of ASD in our cohort (41%), while seizures are present in only two patients. We extend the phenotypic spectrum by sleep disturbances.ConclusionOur study, bringing the total of known patients with ODLURO to more than 60 within 2 years of the first publication, suggests an unexpectedly high relative frequency of this syndrome worldwide. It seems likely that ODLURO, although just recently described, is among the more common single-gene aetiologies of neurodevelopmental delay and ASD. We present the second systematic case series of patients with ODLURO, further refining the mutational and phenotypic spectrum of this not-so-rare syndrome.

Rare genomic disorders (RGDs) confer elevated risk for neurodevelopmental psychiatric disorders. In this era of intense genomics discoveries, the landscape of RGDs is rapidly evolving. However, there has not been comparable progress to date in scalable, harmonized phenotyping methods. As a result, beyond associations with categorical diagnoses, the effects on dimensional traits remain unclear for many RGDs. The nature and specificity of RGD effects on cognitive and behavioral traits is an area of intense investigation: RGDs are frequently associated with more than one psychiatric condition, and those studied to date affect, to varying degrees, a broad range of developmental and cognitive functions. Although many RGDs have large effects, phenotypic expression is typically influenced by additional genomic and environmental factors. There is emerging evidence that using polygenic risk scores in individuals with RGDs offers opportunities to refine prediction, thus allowing for the identification of those at greatest risk of psychiatric illness. However, translation into the clinic is hindered by roadblocks, which include limited genetic testing in clinical psychiatry, and the lack of guidelines for following individuals with RGDs, who are at high risk of developing psychiatric symptoms. The Genes to Mental Health Network (G2MH) is a newly funded National Institute of Mental Health initiative that will collect, share, and analyze large-scale data sets combining genomics and dimensional measures of psychopathology spanning diverse populations and geography. The authors present here the most recent understanding of the effects of RGDs on dimensional behavioral traits and risk for psychiatric conditions and discuss strategies that will be pursued within the G2MH network, as well as how expected results can be translated into clinical practice to improve patient outcomes.

BackgroundClinical exome sequencing typically achieves diagnostic yields of 30%–57.5% in individuals with monogenic rare diseases. Undiagnosed diseases programmes implement strategies to improve diagnostic outcomes for these individuals.AimWe share the lessons learnt from the first 3 years of the Undiagnosed Diseases Program-Victoria, an Australian programme embedded within a clinical genetics service in the state of Victoria with a focus on paediatric rare diseases.MethodsWe enrolled families who remained without a diagnosis after clinical genomic (panel, exome or genome) sequencing between 2016 and 2018. We used family-based exome sequencing (family ES), family-based genome sequencing (family GS), RNA sequencing (RNA-seq) and high-resolution chromosomal microarray (CMA) with research-based analysis.ResultsIn 150 families, we achieved a diagnosis or strong candidate in 64 (42.7%) (37 in known genes with a consistent phenotype, 3 in known genes with a novel phenotype and 24 in novel disease genes). Fifty-four diagnoses or strong candidates were made by family ES, six by family GS with RNA-seq, two by high-resolution CMA and two by data reanalysis.ConclusionWe share our lessons learnt from the programme. Flexible implementation of multiple strategies allowed for scalability and response to the availability of new technologies. Broad implementation of family ES with research-based analysis showed promising yields post a negative clinical singleton ES. RNA-seq offered multiple benefits in family ES-negative populations. International data sharing strategies were critical in facilitating collaborations to establish novel disease–gene associations. Finally, the integrated approach of a multiskilled, multidisciplinary team was fundamental to having diverse perspectives and strategic decision-making.

JAG2 encodes the Notch ligand Jagged2. The conserved Notch signaling pathway contributes to the development and homeostasis of multiple tissues, including skeletal muscle. We studied an international cohort of 23 individuals with genetically unsolved muscular dystrophy from 13 unrelated families. Whole-exome sequencing identified rare homozygous or compound heterozygous JAG2 variants in all 13 families. The identified bi-allelic variants include 10 missense variants that disrupt highly conserved amino acids, a nonsense variant, two frameshift variants, an in-frame deletion, and a microdeletion encompassing JAG2. Onset of muscle weakness occurred from infancy to young adulthood. Serum creatine kinase (CK) levels were normal or mildly elevated. Muscle histology was primarily dystrophic. MRI of the lower extremities revealed a distinct, slightly asymmetric pattern of muscle involvement with cores of preserved and affected muscles in quadriceps and tibialis anterior, in some cases resembling patterns seen in POGLUT1-associated muscular dystrophy. Transcriptome analysis of muscle tissue from two participants suggested misregulation of genes involved in myogenesis, including PAX7. In complementary studies, Jag2 downregulation in murine myoblasts led to downregulation of multiple components of the Notch pathway, including Megf10. Investigations in Drosophila suggested an interaction between Serrate and Drpr, the fly orthologs of JAG1/JAG2 and MEGF10, respectively. In silico analysis predicted that many Jagged2 missense variants are associated with structural changes and protein misfolding. In summary, we describe a muscular dystrophy associated with pathogenic variants in JAG2 and evidence suggests a disease mechanism related to Notch pathway dysfunction.

Focal segmental glomerulosclerosis (FSGS) is the main pathology underlying steroid-resistant nephrotic syndrome (SRNS) and a leading cause of chronic kidney disease. Monogenic forms of pediatric SRNS are predominantly caused by recessive mutations, while the contribution of de novo variants (DNVs) to this trait is poorly understood. Using exome sequencing (ES) in a proband with FSGS/SRNS, developmental delay, and epilepsy, we discovered a nonsense DNV in TRIM8, which encodes the E3 ubiquitin ligase tripartite motif containing 8. To establish whether TRIM8 variants represent a cause of FSGS, we aggregated exome/genome-sequencing data for 2,501 pediatric FSGS/SRNS-affected individuals and 48,556 control subjects, detecting eight heterozygous TRIM8 truncating variants in affected subjects but none in control subjects (p = 3.28 × 10(−11)). In all six cases with available parental DNA, we demonstrated de novo inheritance (p = 2.21 × 10(−15)). Reverse phenotyping revealed neurodevelopmental disease in all eight families. We next analyzed ES from 9,067 individuals with epilepsy, yielding three additional families with truncating TRIM8 variants. Clinical review revealed FSGS in all. All TRIM8 variants cause protein truncation clustering within the last exon between residues 390 and 487 of the 551 amino acid protein, indicating a correlation between this syndrome and loss of the TRIM8 C-terminal region. Wild-type TRIM8 overexpressed in immortalized human podocytes and neuronal cells localized to nuclear bodies, while constructs harboring patient-specific variants mislocalized diffusely to the nucleoplasm. Co-localization studies demonstrated that Gemini and Cajal bodies frequently abut a TRIM8 nuclear body. Truncating TRIM8 DNVs cause a neuro-renal syndrome via aberrant TRIM8 localization, implicating nuclear bodies in FSGS and developmental brain disease.

PURPOSE: Somatic variants in tumor necrosis factor receptor-associated factor 7 (TRAF7) cause meningioma, while germline variants have recently been identified in seven patients with developmental delay and cardiac, facial, and digital anomalies. We aimed to define the clinical and mutational spectrum associated with TRAF7 germline variants in a large series of patients, and to determine the molecular effects of the variants through transcriptomic analysis of patient fibroblasts.METHODS: We performed exome, targeted capture, and Sanger sequencing of patients with undiagnosed developmental disorders, in multiple independent diagnostic or research centers. Phenotypic and mutational comparisons were facilitated through data exchange platforms. Whole-transcriptome sequencing was performed on RNA from patient- and control-derived fibroblasts.RESULTS: We identified heterozygous missense variants in TRAF7 as the cause of a developmental delay-malformation syndrome in 45 patients. Major features include a recognizable facial gestalt (characterized in particular by blepharophimosis), short neck, pectus carinatum, digital deviations, and patent ductus arteriosus. Almost all variants occur in the WD40 repeats and most are recurrent. Several differentially expressed genes were identified in patient fibroblasts.CONCLUSION: We provide the first large-scale analysis of the clinical and mutational spectrum associated with the TRAF7 developmental syndrome, and we shed light on its molecular etiology through transcriptome studies.

Exome and genome sequencing have become the tools of choice for rare disease diagnosis, leading to large amounts of data available for analyses. To identify causal variants in these datasets, powerful filtering and decision support tools that can be efficiently used by clinicians and researchers are required. To address this need, we developed seqr - an open source, web-based tool for family-based monogenic disease analysis that allows researchers to work collaboratively to search and annotate genomic callsets. To date, seqr is being used in several research pipelines and one clinical diagnostic lab. In our own experience through the Broad Institute Center for Mendelian Genomics, seqr has enabled analyses of over 10,000 families, supporting the diagnosis of more than 3,800 individuals with rare disease and discovery of over 300 novel disease genes. Here we describe a framework for genomic analysis in rare disease that leverages seqr’s capabilities for variant filtration, annotation, and causal variant identification, as well as support for research collaboration and data sharing. The seqr platform is available as open source software, allowing low-cost participation in rare disease research, and a community effort to support diagnosis and gene discovery in rare disease.

The precise regulation of gene expression is fundamental to neurodevelopment, plasticity, and cognitive function. While several studies have deeply profiled mRNA dynamics in the developing human brain, there is a fundamental gap in our understanding of accompanying translational regulation. We perform ribosome profiling from more than 70 human prenatal and adult cortex samples across ontogeny and into adulthood, mapping translation events at nucleotide resolution. In addition to characterizing the translational regulation of annotated open reading frames (ORFs), we identify thousands of previously unknown translation events, including small open reading frames (sORFs) that give rise to human- and/or brain-specific microproteins, many of which we independently verify using size-selected proteomics. Ribosome profiling in stem cell-derived human neuronal cultures further corroborates these findings and shows that several neuronal activity-induced long non-coding RNAs (lncRNAs), including LINC00473, a primate-specific lncRNA implicated in depression, encode previously undescribed microproteins. Physicochemical analysis of these brain microproteinss identifies a large class harboring arginine-glycine-glycine (RGG) repeats as strong candidates for regulating RNA metabolism. Moreover, we find that, collectively, these previously unknown human brain sORFs are enriched for variants associated with schizophrenia. In addition to significantly expanding the translational landscape of the developing brain, this atlas will serve as a rich resource for the annotation and functional interrogation of thousands of previously unknown brain-specific protein products.

The phenotypic variability associated with pathogenic variants in Lysine Acetyltransferase 6B (KAT6B, a.k.a. MORF, MYST4) results in several interrelated syndromes including Say‐Barber‐Biesecker‐Young‐Simpson Syndrome and Genitopatellar Syndrome. Here we present 20 new cases representing 10 novel KAT6B variants. These patients exhibit a range of clinical phenotypes including intellectual disability, mobility and language difficulties, craniofacial dysmorphology, and skeletal anomalies. Given the range of features previously described for KAT6B‐related syndromes, we have identified additional phenotypes including concern for keratoconus, sensitivity to light or noise, recurring infections, and fractures in greater numbers than previously reported. We surveyed clinicians to qualitatively assess the ways families engage with genetic counselors upon diagnosis. We found that 56% (10/18) of individuals receive diagnoses before the age of 2 years (median age = 1.96 years), making it challenging to address future complications with limited accessible information and vast phenotypic severity. We used CRISPR to introduce truncating variants into the KAT6B gene in model cell lines and performed chromatin accessibility and transcriptome sequencing to identify key dysregulated pathways. This study expands the clinical spectrum and addresses the challenges to management and genetic counseling for patients with KAT6B‐related disorders., We describe 20 new cases harboring the KAT6B spectrum of disorders, which range from Say‐Barber‐Biesecker‐Young‐Simpson (SBBYSS) to Genitopatellar (GPS) syndrome or an intermediate phenotype. In our holistic approach, we expand the genotypic and phenotypic spectrum of KAT6B spectrum of disorders. Furthermore, we provide extensive clinical phenotyping, explore the impact of genetic counseling for these complex syndromes, and examine molecular mechanisms in RNA‐seq data in an in vitro cell model of truncating KAT6B mutations.

Genetic variants that inactivate protein-coding genes are a powerful source of information about the phenotypic consequences of gene disruption: genes that are crucial for the function of an organism will be depleted of such variants in natural populations, whereas non-essential genes will tolerate their accumulation. However, predicted loss-of-function variants are enriched for annotation errors, and tend to be found at extremely low frequencies, so their analysis requires careful variant annotation and very large sample sizes

The genetic causes of global developmental delay (GDD) and intellectual disability (ID) are diverse and include variants in numerous ion channels and transporters. Loss-of-function variants in all five endosomal/lysosomal members of the CLC family of Cl− channels and Cl−/H+ exchangers lead to pathology in mice, humans, or both. We have identified nine variants in CLCN3, the gene encoding CIC-3, in 11 individuals with GDD/ID and neurodevelopmental disorders of varying severity. In addition to a homozygous frameshift variant in two siblings, we identified eight different heterozygous de novo missense variants. All have GDD/ID, mood or behavioral disorders, and dysmorphic features; 9/11 have structural brain abnormalities; and 6/11 have seizures. The homozygous variants are predicted to cause loss of ClC-3 function, resulting in severe neurological disease similar to the phenotype observed in Clcn3−/− mice. Their MRIs show possible neurodegeneration with thin corpora callosa and decreased white matter volumes. Individuals with heterozygous variants had a range of neurodevelopmental anomalies including agenesis of the corpus callosum, pons hypoplasia, and increased gyral folding. To characterize the altered function of the exchanger, electrophysiological analyses were performed in Xenopus oocytes and mammalian cells. Two variants, p.Ile607Thr and p.Thr570Ile, had increased currents at negative cytoplasmic voltages and loss of inhibition by luminal acidic pH. In contrast, two other variants showed no significant difference in the current properties. Overall, our work establishes a role for CLCN3 in human neurodevelopment and shows that both homozygous loss of ClC-3 and heterozygous variants can lead to GDD/ID and neuroanatomical abnormalities.

Wahlster, Verboon, et al. identify and functionally characterize a complex structural variant in a family with thrombocytopenia that results in a WAC-ANKRD26 fusion transcript encoding a shortened form of ANKRD26, whose expression during hematopoiesis is deregulated, thereby resulting in disease., Advances in genome sequencing have resulted in the identification of the causes for numerous rare diseases. However, many cases remain unsolved with standard molecular analyses. We describe a family presenting with a phenotype resembling inherited thrombocytopenia 2 (THC2). THC2 is generally caused by single nucleotide variants that prevent silencing of ANKRD26 expression during hematopoietic differentiation. Short-read whole-exome and genome sequencing approaches were unable to identify a causal variant in this family. Using long-read whole-genome sequencing, a large complex structural variant involving a paired-duplication inversion was identified. Through functional studies, we show that this structural variant results in a pathogenic gain-of-function WAC-ANKRD26 fusion transcript. Our findings illustrate how complex structural variants that may be missed by conventional genome sequencing approaches can cause human disease.

Variants in multiple tubulin genes have been implicated in neurodevelopmental disorders, including malformations of cortical development (MCD) and congenital fibrosis of the extraocular muscles (CFEOM). Distinct missense variants in the beta-tubulin encoding genes TUBB3 and TUBB2B cause MCD, CFEOM, or both, suggesting substitution-specific mechanisms. Variants in the alpha tubulin-encoding gene TUBA1A have been associated with MCD, but not with CFEOM. Using exome sequencing (ES) and genome sequencing (GS), we identified 3 unrelated probands with CFEOM who harbored novel heterozygous TUBA1A missense variants c.1216C>G, p.(His406Asp); c.467G>A, p.(Arg156His); and c.1193T>G, p.(Met398Arg). MRI revealed small oculomotor-innervated muscles and asymmetrical caudate heads and lateral ventricles with or without corpus callosal thinning. Two of the three probands had MCD. Mutated amino acid residues localize either to the longitudinal interface at which α and β tubulins heterodimerize (Met398, His406) or to the lateral interface at which tubulin protofilaments interact (Arg156), and His406 interacts with the motor domain of kinesin-1. This series of individuals supports TUBA1A variants as a cause of CFEOM and expands our knowledge of tubulinopathies.

BACKGROUND: Deficiency of adenosine deaminase 2 (DADA2) is an autoinflammatory disease caused by deleterious ADA2 variants. The frequency of these variants in the general population, and hence the expected disease prevalence, remain unknown. OBJECTIVE: We aim to characterize the functional impact and carrier frequency of ADA2 variants. METHODS: We performed functional studies and in silico analysis on 163 ADA2 variants, including DADA2-associated variants and population variants identified in the Genome Aggregation Database (gnomAD). We estimated the carrier rate using the aggregate frequency of deleterious variants. RESULTS: Functional studies of ADA2 variants revealed that 77/85 (91%) of DADA2-associated variants reduced ADA2 enzymatic function by > 75%. Analysis of 100 ADA2 variants in gnomAD showed a full spectrum of impact on ADA2 function, rather than a dichotomy of benign versus deleterious variants. We found several in silico algorithms that effectively predicted the impact of ADA2 variants with high sensitivity and specificity, and confirmed a correlation between the residual function of ADA2 variants in vitro and the plasma ADA2 activity of individuals carrying these variants (n = 45; r = 0.649; p < 0.0001). Using < 25% residual enzymatic activity as the cut-off to define potential pathogenicity, integration of our results with gnomAD population data revealed an estimated carrier frequency of at least 1 in 236 individuals, corresponding to an expected DADA2 disease prevalence of ~1 in 222,000 individuals. CONCLUSION: Functional annotation guides the interpretation of ADA2 variants to create a framework that enables estimation of DADA2 carrier frequency and disease prevalence.

Correction to: Nature Communications https://doi.org/10.1038/s41467-019-10717-9, published online 27 May 2020.The original version of this Article omitted from the Genome Aggregation Database consortium the member Marquis P. Vawter, from the Department of Psychiatry & Human Behavior, University of California Irvine, Irvine, CA, USA. Additionally, the following was added to the Author Contributions: ‘All authors listed under The Genome Aggregation Database Consortium contributed to the generation of the primary data incorporated into the gnomAD resource’. This has been corrected in both the PDF and HTML versions of the Article.

Dubowitz syndrome (DubS) is considered a recognizable syndrome characterized by a distinctive facial appearance and deficits in growth and development. There have been over 200 individuals reported with Dubowitz or a "Dubowitz-like" condition, although no single gene has been implicated as responsible for its cause. We have performed exome (ES) or genome sequencing (GS) for 31 individuals clinically diagnosed with DubS. After genome-wide sequencing, rare variant filtering and computational and Mendelian genomic analyses, a presumptive molecular diagnosis was made in 13/27 (48%) families. The molecular diagnoses included biallelic variants in SKIV2L, SLC35C1, BRCA1, NSUN2; de novo variants in ARID1B, ARID1A, CREBBP, POGZ, TAF1, HDAC8, and copy-number variation at1p36.11(ARID1A), 8q22.2(VPS13B), Xp22, and Xq13(HDAC8). Variants of unknown significance in known disease genes, and also in genes of uncertain significance, were observed in 7/27 (26%) additional families. Only one gene, HDAC8, could explain the phenotype in more than one family (N = 2). All but two of the genomic diagnoses were for genes discovered, or for conditions recognized, since the introduction of next-generation sequencing. Overall, the DubS-like clinical phenotype is associated with extensive locus heterogeneity and the molecular diagnoses made are for emerging clinical conditions sharing characteristic features that overlap the DubS phenotype.

Hundreds of thousands of genetic variants have been reported to cause severe monogenic diseases, but the probability that a variant carrier will develop the disease (termed penetrance) is unknown for virtually all of them. Additionally, the clinical utility of common polygenetic variation remains uncertain. Using exome sequencing from 77,184 adult individuals (38,618 multi-ancestral individuals from a type 2 diabetes case-control study and 38,566 participants from the UK Biobank, for whom genotype array data were also available), we applied clinical standard-of-care gene variant curation for eight monogenic metabolic conditions. Rare variants causing monogenic diabetes and dyslipidemias displayed effect sizes significantly larger than the top 1% of the corresponding polygenic scores. Nevertheless, penetrance estimates for monogenic variant carriers averaged below 60% in both studies for all conditions except monogenic diabetes. We assessed additional epidemiologic and genetic factors contributing to risk prediction, demonstrating that inclusion of common polygenic variation significantly improved biomarker estimation for two monogenic dyslipidemias.

Current clinical guidelines recommend three genetic tests for the assessment of fetal structural anomalies: karyotype to detect microscopically-visible balanced and unbalanced chromosomal rearrangements, chromosomal microarray (CMA) to detect sub-microscopic copy number variants (CNVs), and exome sequencing (ES) to identify individual nucleotide changes in coding sequence. Advances in genome sequencing (GS) analysis suggest that it is poised to displace the sequential application of all three conventional tests to become a single diagnostic approach for the assessment of fetal structural anomalies. However, systematic benchmarking is required to assure that GS can capture the full mutational spectrum associated with fetal structural anomalies and to accurately quantify the added diagnostic yield of GS. We applied a novel GS analytic framework that included the discovery, filtration, and interpretation of nine classes of genomic variation to 7,195 individuals. We assessed the sensitivity of GS to detect diagnostic variants (pathogenic or likely pathogenic) from three standard-of-care tests using 1,612 autism spectrum disorder quartet families (ASD; n=6,448) with matched GS, ES, and CMA data, and validated these findings in 46 fetuses with a clinically reportable variant originally identified by karyotype, CMA, or ES. We then assessed the added diagnostic yield of GS in 249 trios (n=747) comprising a fetus with a structural anomaly detected by ultrasound and two unaffected parents that were pre-screened with a combination of all three standard-of-care tests. Across both cohorts, our GS analytic framework identified 98.2% of all diagnostic variants detected by standard-of-care tests, including 100% of those originally detected by CMA (n=88) and ES (n=61), as well as 78.6% (n=11/14) of the chromosomal rearrangements identified by karyotype. The diagnostic yield from GS was 7.8% across all 1,612 ASD probands, almost two-fold more than CMA (4.4%) and three-fold more than ES (3.0%). We also demonstrated that the yield of ES can approach that of GS when CNVs are captured with high sensitivity from exome data (7.4% vs. 7.8%, respectively). In 249 pre-screened fetuses with structural anomalies, GS provided an additional diagnostic yield of 0.4% beyond the combination of all three tests (karyotype, CMA, and ES). Applying our benchmarking results to existing data indicates that GS can achieve an overall diagnostic yield of 46.1% in unselected fetuses with fetal structural anomalies, providing an estimated 17.2% increase in diagnostic yield over karyotype, 14.1% over CMA, and 36.1% over ES when sequence variants are assessed, and 4.1% when CNVs are also identified from exome data. In this study we demonstrate that GS is sensitive to the detection of almost all pathogenic variation captured by karyotype, CMA, and ES, provides a superior diagnostic yield than any individual test by a wide margin, and contributes a modest increase in diagnostic yield beyond the combination of all three tests. We also outline several strategies to aid the interpretation of GS variants that are cryptic to conventional technologies, which we anticipate will be increasingly encountered as comprehensive variant identification from GS is performed. Taken together, these data suggest GS warrants consideration as a first-tier diagnostic approach for fetal structural anomalies.

Animal development and homeostasis depend on precise temporal and spatial intercellular signaling. Components shared between signaling pathways, generally thought to decrease specificity, paradoxically can also provide a solution to pathway coordination. Here we show that the Bone Morphogenetic Protein (BMP) and Wnt signaling pathways share Apcdd1 as a common inhibitor and that Apcdd1 is a taxon-restricted gene with novel domains and signaling functions. Previously, we showed that Apcdd1 inhibits Wnt signaling, here we find that Apcdd1 potently inhibits BMP signaling in body axis formation and neural differentiation in chicken, frog, zebrafish, and humans. Our results from experiments and modeling suggest that Apcdd1 may coordinate the outputs of two signaling pathways central to animal development and human disease.Significance StatementApcdd1is a taxon-restricted gene that inhibits both BMP and Wnt intercellular signaling pathways in multiple organisms including mice, frog, zebrafish, and chicken. It encodes a bi-functional protein with a novel protein domain that can bind to Wnt and BMP receptors and block downstream signaling.

Structural variants (SVs) rearrange large segments of DNA1 and can have profound consequences in evolution and human disease2,3. As national biobanks, disease-association studies, and clinical genetic testing have grown increasingly reliant on genome sequencing, population references such as the Genome Aggregation Database (gnomAD)4 have become integral in the interpretation of single-nucleotide variants (SNVs)5. However, there are no reference maps of SVs from high-coverage genome sequencing comparable to those for SNVs. Here we present a reference of sequence-resolved SVs constructed from 14,891 genomes across diverse global populations (54% non-European) in gnomAD. We discovered a rich and complex landscape of 433,371 SVs, from which we estimate that SVs are responsible for 25–29% of all rare protein-truncating events per genome. We found strong correlations between natural selection against damaging SNVs and rare SVs that disrupt or duplicate protein-coding sequence, which suggests that genes that are highly intolerant to loss-of-function are also sensitive to increased dosage6. We also uncovered modest selection against noncoding SVs in cis-regulatory elements, although selection against protein-truncating SVs was stronger than all noncoding effects. Finally, we identified very large (over one megabase), rare SVs in 3.9% of samples, and estimate that 0.13% of individuals may carry an SV that meets the existing criteria for clinically important incidental findings7. This SV resource is freely distributed via the gnomAD browser8 and will have broad utility in population genetics, disease-association studies, and diagnostic screening., A large empirical assessment of sequence-resolved structural variants from 14,891 genomes across diverse global populations in the Genome Aggregation Database (gnomAD) provides a reference map for disease-association studies, population genetics, and diagnostic screening.

Rare disease investigators constantly face challenges in identifying additional cases to build evidence for gene‐disease causality. The Matchmaker Exchange (MME) addresses this limitation by providing a mechanism for matching patients across genomic centers via a federated network. The MME has revolutionized searching for additional cases by making it possible to query across institutional boundaries, so that what was once a laborious and manual process of contacting researchers is now automated and computable. However, while the MME network is beginning to scale, the growth of additional nodes is limited by the lack of easy‐to‐use solutions that can be implemented by any rare disease database owner, even one without significant software engineering resources. Here we describe matchbox, which is an open‐source, platform‐independent, portable bridge between any given rare disease genomic center and the MME network, which has already led to novel gene discoveries. We also describe how matchbox greatly reduces the barrier to participation by overcoming challenges for new databases to join the MME. Refereed/Peer-reviewed

Acute idiopathic hyperammonemia in an adult patient is a life-threatening condition often resulting in a rapid progression to irreversible cerebral edema and death. While ammonia-scavenging therapies lower blood ammonia levels, in comparison, clearance of waste nitrogen from the brain may be delayed. Therefore, we used magnetic resonance spectroscopy (MRS) to monitor cerebral glutamine levels, the major reservoir of ammonia, in a gastric bypass patient with hyperammonemic coma undergoing therapy with N-carbamoyl glutamate and the ammonia-scavenging agents, sodium phenylacetate and sodium benzoate. Improvement in mental status mirrored brain glutamine levels, as coma persisted for 48h after plasma ammonia normalized. We hypothesize that the slower clearance for brain glutamine levels accounts for the delay in improvement following initiation of treatment in cases of chronic hyperammonemia. We propose MRS to monitor brain glutamine as a noninvasive approach to be utilized for diagnostic and therapeutic monitoring purposes in adult patients presenting with idiopathic hyperammonemia.

The dispensability of individual genes for viability has interested generations of geneticists. For some genes it is essential to maintain two functional chromosomal copies, while others may tolerate the loss of one or both copies. Exome sequence data from 60,706 individuals provide sufficient observations of rare protein truncating variants (PTVs) to make genome-wide estimates of selection against heterozygous loss of gene function. The cumulative frequency of rare deleterious PTVs is primarily determined by the balance between incoming mutations and purifying selection rather than genetic drift. This enables the estimation of the genome-wide distribution of selection coefficients for heterozygous PTVs and corresponding Bayesian estimates for individual genes. The strength of selection can discriminate the severity, age of onset, and mode of inheritance in Mendelian exome sequencing cases. We find that genes under the strongest selection are enriched in embryonic lethal mouse knockouts, putatively cell-essential genes, Mendelian disease genes, and regulators of transcription. Screening by essentiality, we find a large set of genes under strong selection that likely have critical function but have not yet been extensively annotated in published literature.

Contains fulltext : 206572.pdf (Publisher’s version ) (Open Access) We delineate a KMT2E-related neurodevelopmental disorder on the basis of 38 individuals in 36 families. This study includes 31 distinct heterozygous variants in KMT2E (28 ascertained from Matchmaker Exchange and three previously reported), and four individuals with chromosome 7q22.2-22.23 microdeletions encompassing KMT2E (one previously reported). Almost all variants occurred de novo, and most were truncating. Most affected individuals with protein-truncating variants presented with mild intellectual disability. One-quarter of individuals met criteria for autism. Additional common features include macrocephaly, hypotonia, functional gastrointestinal abnormalities, and a subtle facial gestalt. Epilepsy was present in about one-fifth of individuals with truncating variants and was responsive to treatment with anti-epileptic medications in almost all. More than 70% of the individuals were male, and expressivity was variable by sex; epilepsy was more common in females and autism more common in males. The four individuals with microdeletions encompassing KMT2E generally presented similarly to those with truncating variants, but the degree of developmental delay was greater. The group of four individuals with missense variants in KMT2E presented with the most severe developmental delays. Epilepsy was present in all individuals with missense variants, often manifesting as treatment-resistant infantile epileptic encephalopathy. Microcephaly was also common in this group. Haploinsufficiency versus gain-of-function or dominant-negative effects specific to these missense variants in KMT2E might explain this divergence in phenotype, but requires independent validation. Disruptive variants in KMT2E are an under-recognized cause of neurodevelopmental abnormalities.

Missense variant interpretation is challenging. Essential regions for protein function are conserved among gene family members, and genetic variants within these regions are potentially more likely to confer risk to disease. Here, we generated 2,871 gene family protein sequence alignments involving 9,990 genes and performed missense variant burden analyses to identify novel essential protein regions. We mapped 2,219,811 variants from the general population into these alignments and compared their distribution with 65,034 missense variants from patients. With this gene family approach, we identified 398 regions enriched for patient variants spanning 33,887 amino acids in 1,058 genes. As a comparison, testing the same genes individually we identified less patient variant enriched regions involving only 2,167 amino acids and 180 genes. Next, we selected de novo variants from 6,753 patients with neurodevelopmental disorders and 1,911 unaffected siblings, and observed a 5.56-fold enrichment of patient variants in our identified regions (95% C.I. =2.76-Inf, p-value = 6.66×10−8). Using an independent ClinVar variant set, we found missense variants inside the identified regions are 111-fold more likely to be classified as pathogenic in comparison to benign classification (OR = 111.48, 95% C.I = 68.09-195.58, p-value < 2.2e−16). All patient variant enriched regions identified (PERs) are available online through a user-friendly platform for interactive data mining, visualization and download at http://per.broadinstitute.org. In summary, our gene family burden analysis approach identified novel patient variant enriched regions in protein sequences. This annotation can empower variant interpretation.

Meiotic nondisjunction and resulting aneuploidy can lead to severe health consequences in humans. Aneuploidy rescue can restore euploidy but may result in uniparental disomy (UPD), the inheritance of both homologs of a chromosome from one parent with no representative copy from the other. Current understanding of UPD is limited to ∼3,300 case subjects for which UPD was associated with clinical presentation due to imprinting disorders or recessive diseases. Thus, the prevalence of UPD and its phenotypic consequences in the general population are unknown. We searched for instances of UPD across 4,400,363 consented research participants from the personal genetics company 23andMe, Inc., and 431,094 UK Biobank participants. Using computationally detected DNA segments identical-by-descent (IBD) and runs of homozygosity (ROH), we identified 675 instances of UPD across both databases. We estimate that UPD is twice as common as previously thought, and we present a machine-learning framework to detect UPD using ROH. While we find a nominally significant association between UPD of chromosome 22 and autism risk, we do not find significant associations between UPD and deleterious traits in the 23andMe database.

We describe an infant with a phenotype typical of early onset Marfan syndrome whose genetic evaluation, including Sanger sequencing and deletion/duplication testing of FBN1 and exome sequencing, was negative. Ultimately, genome sequencing revealed a deletion missed on prior testing, demonstrating the unique utility of genome sequencing for molecular genetic diagnosis.

SummaryMeiotic nondisjunction and resulting aneuploidy can lead to severe health consequences in humans. Aneuploidy rescue can restore euploidy but may result in uniparental disomy (UPD), the inheritance of both homologs of a chromosome from one parent with no representative copy from the other. Current understanding of UPD is limited to ~3,300 cases for which UPD was associated with clinical presentation due to imprinting disorders or recessive diseases. Thus, the prevalence of UPD and its phenotypic consequences in the general population are unknown. We searched for instances of UPD in over four million consented research participants from the personal genetics company 23andMe, Inc., and 431,094 UK Biobank participants. Using computationally detected DNA segments identical-by-descent (IBD) and runs of homozygosity (ROH), we identified 675 instances of UPD across both databases. Here we present the first characterization of UPD prevalence in the general population, a machine-learning framework to detect UPD using ROH, and a novel association between autism and UPD of chromosome 22.

SummaryGenetic variants that inactivate protein-coding genes are a powerful source of information about the phenotypic consequences of gene disruption: genes critical for an organism’s function will be depleted for such variants in natural populations, while non-essential genes will tolerate their accumulation. However, predicted loss-of-function (pLoF) variants are enriched for annotation errors, and tend to be found at extremely low frequencies, so their analysis requires careful variant annotation and very large sample sizes1. Here, we describe the aggregation of 125,748 exomes and 15,708 genomes from human sequencing studies into the Genome Aggregation Database (gnomAD). We identify 443,769 high-confidence pLoF variants in this cohort after filtering for sequencing and annotation artifacts. Using an improved human mutation rate model, we classify human protein-coding genes along a spectrum representing tolerance to inactivation, validate this classification using data from model organisms and engineered human cells, and show that it can be used to improve gene discovery power for both common and rare diseases.

Identifying genes and variants contributing to rare disease phenotypes and Mendelian conditions informs biology and medicine, yet potential phenotypic consequences for variation of >75% of the ~20,000 annotated genes in the human genome are lacking. Technical advances to assess rare variation genome-wide, particularly exome sequencing (ES), enabled establishment in the United States of the National Institutes of Health (NIH)-supported Centers for Mendelian Genomics (CMGs) and have facilitated collaborative studies resulting in novel “disease gene” discoveries. Pedigree-based genomic studies and rare variant analyses in families with suspected Mendelian conditions have led to the elucidation of hundreds of novel disease genes and highlighted the impact of de novo mutational events, somatic variation underlying nononcologic traits, incompletely penetrant alleles, phenotypes with high locus heterogeneity, and multilocus pathogenic variation. Herein, we highlight CMG collaborative discoveries that have contributed to understanding both rare and common diseases and discuss opportunities for future discovery in single-locus Mendelian disorder genomics. Phenotypic annotation of all human genes; development of bioinformatic tools and analytic methods; exploration of non-Mendelian modes of inheritance including reduced penetrance, multilocus variation, and oligogenic inheritance; construction of allelic series at a locus; enhanced data sharing worldwide; and integration with clinical genomics are explored. Realizing the full contribution of rare disease research to functional annotation of the human genome, and further illuminating human biology and health, will lay the foundation for the Precision Medicine Initiative.

Diamond-Blackfan anemia (DBA) is a rare bone marrow failure disorder that affects 1 in 100,000 to 200,000 live births and has been associated with mutations in components of the ribosome. In order to characterize the genetic landscape of this genetically heterogeneous disorder, we recruited a cohort of 472 individuals with a clinical diagnosis of DBA and performed whole exome sequencing (WES). Overall, we identified rare and predicted damaging mutations in likely causal genes for 78% of individuals. The majority of mutations were singletons, absent from population databases, predicted to cause loss of function, and in one of 19 previously reported genes encoding for a diverse set of ribosomal proteins (RPs). Using WES exon coverage estimates, we were able to identify and validate 31 deletions in DBA associated genes. We also observed an enrichment for extended splice site mutations and validated the diverse effects of these mutations using RNA sequencing in patientderived cell lines. Leveraging the size of our cohort, we observed several robust genotype-phenotype associations with congenital abnormalities and treatment outcomes. In addition to comprehensively identifying mutations in known genes, we further identified rare mutations in 7 previously unreported RP genes that may cause DBA. We also identified several distinct disorders that appear to phenocopy DBA, including 9 individuals with biallelicCECR1mutations that result in deficiency of ADA2. However, no new genes were identified at exome-wide significance, suggesting that there are no unidentified genes containing mutations readily identified by WES that explain > 5% of DBA cases. Overall, this comprehensive report should not only inform clinical practice for DBA patients, but also the design and analysis of future rare variant studies for heterogeneous Mendelian disorders.

A Correction to this paper has been published: https://doi.org/10.1038/s41586-020-03176-6.

Rare gene knockouts in adult humans On average, most people's genomes contain approximately 100 completely nonfunctional genes. These loss-of-function (LOF) mutations tend to be rare and/or occur only as a single copy within individuals. Narasimhan et al. investigated LOF in a Pakistani population with high levels of consanguinity. Examining LOF alleles that were identical by descent, they found, as expected, an absence of homozygote LOF for certain protein-coding genes. However, they also identified many homozygote LOF alleles with no apparent deleterious phenotype, including some that were expected to confer genetic disease. Indeed, one family had lost the recombination-associated gene PRDM9 . Science , this issue p. 474

Clinical exome sequencing (CES) is increasingly being utilized; however, a large proportion of patients remain undiagnosed, creating a need for a systematic approach to increase the diagnostic yield. We have reanalyzed CES data for a clinically heterogeneous cohort of 102 probands with likely Mendelian conditions, including 74 negative cases and 28 cases with candidate variants, but reanalysis requested by clinicians. Reanalysis was performed by an interdisciplinary team using a validated custom-built pipeline, “Variant Explorer Pipeline” (VExP). This reanalysis approach and results were compared with existing literature. Reanalysis of candidate variants from CES in 28 cases revealed 1 interpretation that needed to be reclassified. A confirmed or potential genetic diagnosis was identified in 24 of 75 CES-negative/reclassified cases (32.0%), including variants in known disease-causing genes (n = 6) or candidate genes (n = 18). This yield was higher compared with similar studies demonstrating the utility of this approach. In summary, reanalysis of negative CES in a research setting enhances diagnostic yield by about a third. This study suggests the need for comprehensive, continued reanalysis of exome data when molecular diagnosis is elusive.

Recent work by Shah and colleagues demonstrated that many variants in the ClinVar database are misclassified, and that disease-specific allele frequency (AF) thresholds can identify wrongly classified alleles by flagging variants that are too prevalent in the population to be causative of rare penetrant disease. While we agree with the main conclusions of this work, the authors compare their AF filtering approach to our recently published method, concluding that the method we advanced 9may be prone to removing potentially pathogenic variants9. This is incorrect. Here we demonstrate that our approach is robust, and further illustrate the power of disease-specific AF thresholds for investigating the genetic architecture of disease.

We describe 2 children with cobalamin G disease, a disorder of vitamin B

The etiology of intellectual disabilities (ID) remains unknown for the majority of patients. Due to reduced reproductive fitness in many individuals with ID, de novo mutations account for a significant portion of severe ID. The ATP-dependent SWI/SNF chromatin modifier has been linked with neurodevelopmental disorders including ID and autism. ARID2 is an intrinsic component of polybromo-associated BAF (PBAF), the SWI/SNF subcomplex. In this study, we used clinical whole exome sequencing (WES) in proband-parent-trios to identify the etiology of ID. We identified four independent, novel, loss of function variants in ARID2 gene in four patients, three of which were confirmed to be de novo. The patients all have ID and share other clinical characteristics including attention deficit hyperactivity disorder, short stature, dysmorphic facial features, and Wormian bones. All four novel variants are predicted to lead to a premature termination with the loss of the two conservative zinc finger motifs. This is the first report of mutations in ARID2 associated with developmental delay and ID.

We review recent understanding of the pathophysiology, molecular biology, and management of Turner syndrome.Sophisticated genetic techniques are able to detect mosaicism in one-third of individuals previously thought to have monosomy X. Prenatal detection using maternal blood should permit noninvasive detection of most fetuses with an X chromosome abnormality. Disproportionate growth with short limbs has been documented in this condition, and a target gene of short stature homeobox, connective tissue growth factor (Ctgf), has been described. Liver disease is more common in Turner syndrome than previously recognized. Most girls have gonadal failure. Spontaneous puberty and menarche is more commonly seen in girls with XX mosaicism. Low-dose estrogen replacement therapy may be given early to induce a more normal onset and tempo of puberty. Oocyte donation for assisted reproduction carries a substantial risk, particularly if the woman has known cardiac or aortic disease. Neurodevelopmental differences in Turner syndrome are beginning to be correlated with differences in brain anatomy.An increased understanding of the molecular basis for aspects of this disorder is now developing. In addition, a renewed focus on health maintenance through the life span should provide better general and targeted healthcare for these girls and women.

This software repository provides a pipeline for converting raw ClinVar data files into analysis-friendly tab-delimited tables, and also provides these tables for the most recent ClinVar release. Separate tables are generated for genome builds GRCh37 and GRCh38 as well as for mono-allelic variants and complex multi-allelic variants. Additionally, the tables are augmented with allele frequencies from the ExAC and gnomAD datasets as these are often consulted when analyzing ClinVar variants. Overall, this work provides ClinVar data in a format that is easier to work with and can be directly loaded into a variety of popular analysis tools such as R, python pandas, and SQL databases.

A major goal of biomedicine is to understand the function of every gene in the human genome.1 Loss-of-function (LoF) mutations can disrupt both copies of a given gene in humans and phenotypic analysis of such ‘human knockouts’ can provide insight into gene function. Consanguineous unions are more likely to result in offspring who carry LoF mutations in a homozygous state. In Pakistan, consanguinity rates are notably high.2 Here, we sequenced the protein-coding regions of 10,503 adult participants in the Pakistan Risk of Myocardial Infarction Study (PROMIS) designed to understand the determinants of cardiometabolic diseases in South Asians.3 We identified individuals carrying predicted LoF (pLoF) mutations in the homozygous state, and performed phenotypic analysis involving >200 biochemical and disease traits. We enumerated 49,138 rare (

Rapid whole-exome sequencing (rWES) is used in critically ill newborn infants to inform about diagnosis, clinical management, and prognosis. Here we report a male newborn infant with hydrops, pancytopenia, and acute liver failure who was listed for liver transplantation. Given the acuity of the presentation, the procedure-related morbidity and mortality, and lack of diagnosis, we used rWES in the proband and both parents with a turnaround time of 10 business days. rWES returned one maternally inherited, likely pathogenic and one paternally inherited, likely pathogenic variant in NPC1, suggestive of a diagnosis of Niemann–Pick disease type C (NPC). Interestingly, a diagnosis of NPC was entertained prior to rWES, but deemed unlikely in light of absent cholesterol storage on liver biopsy and near-normal oxysterol levels in dried blood. The diagnosis of NPC was confirmed on filipin stain in fibroblasts demonstrating defective cholesterol trafficking. NPC is a slowly progressive neurodegenerative disorder that may also affect the liver with overall poor prognosis. It was decided to take the infant off the transplant list and transfer to palliative care, where he died after 4 wk. This case highlights the utility of rWES in an acute clinical setting for several domains of precision medicine including (1) diagnosis, (2) prognosis and outcome, (3) management and therapy, and (4) utilization of resources.