A comparative medical genomics approach may facilitate the interpretation of rare missense variation.

PURPOSE: To determine the degree to which likely causal missense variants of single-locus traits in domesticated species have features suggestive of pathogenicity in a human genomic context. METHODS: We extracted missense variants from the Online Mendelian Inheritance in Animals database for nine animals (cat, cattle, chicken, dog, goat, horse, pig, rabbit and sheep), mapped coordinates to the human reference genome and annotated variants using genome analysis tools. We also searched a private commercial laboratory database of genetic testing results from >400 000 individuals with suspected rare disorders. RESULTS: Of 339 variants that were mappable to the same residue and gene in the human genome, 56 had been previously classified with respect to pathogenicity: 31 (55.4%) pathogenic/likely pathogenic, 1 (1.8%) benign/likely benign and 24 (42.9%) uncertain/other. The odds ratio for a pathogenic/likely pathogenic classification in ClinVar was 7.0 (95% CI 4.1 to 12.0, p<0.0001), compared with all other germline missense variants in these same 220 genes. The remaining 283 variants disproportionately had allele frequencies and REVEL scores that supported pathogenicity. CONCLUSION: Cross-species comparisons could facilitate the interpretation of rare missense variation. These results provide further support for comparative medical genomics approaches that connect big data initiatives in human and veterinary genetics.

De novo missense variants in exon 9 of SEPHS1 cause a neurodevelopmental condition with developmental delay, poor growth, hypotonia, and dysmorphic features.

Selenophosphate synthetase (SEPHS) plays an essential role in selenium metabolism. Two mammalian SEPHS paralogues, SEPHS1 and SEPHS2, share high sequence identity and structural homology with SEPHS. Here, we report nine individuals from eight families with developmental delay, growth and feeding problems, hypotonia, and dysmorphic features, all with heterozygous missense variants in SEPHS1. Eight of these individuals had a recurrent variant at amino acid position 371 of SEPHS1 (p.Arg371Trp, p.Arg371Gln, and p.Arg371Gly); seven of these variants were known to be de novo. Structural modeling and biochemical assays were used to understand the effect of these variants on SEPHS1 function. We found that a variant at residue Trp352 results in local structural changes of the C-terminal region of SEPHS1 that decrease the overall thermal stability of the enzyme. In contrast, variants of a solvent-exposed residue Arg371 do not impact enzyme stability and folding but could modulate direct protein-protein interactions of SEPSH1 with cellular factors in promoting cell proliferation and development. In neuronal SH-SY5Y cells, we assessed the impact of SEPHS1 variants on cell proliferation and ROS production and investigated the mRNA expression levels of genes encoding stress-related selenoproteins. Our findings provided evidence that the identified SEPHS1 variants enhance cell proliferation by modulating ROS homeostasis. Our study supports the hypothesis that SEPHS1 plays a critical role during human development and provides a basis for further investigation into the molecular mechanisms employed by SEPHS1. Furthermore, our data suggest that variants in SEPHS1 are associated with a neurodevelopmental disorder.

TELO2-related syndrome (You-Hoover-Fong syndrome): Description of 14 new affected individuals and review of the literature.

You-Hoover-Fong syndrome (YHFS) is an autosomal recessive condition caused by pathogenic variants in the TELO2 gene. Affected individuals were reported to have global developmental delay, intellectual disability, microcephaly, dysmorphic facial features, ocular involvement including cortical visual impairment, strabismus, cataract and rotatory nystagmus, movement disorder, hypertonia and spasticity, balance disturbance and ataxia, and abnormal sleep pattern. Other features reported include poor growth, cleft palate, cardiac malformations, epilepsy, scoliosis, and hearing loss. To date, 12 individuals with YHFS have been reported in the literature. Here we describe 14 new individuals with YHFS from 10 families. Their clinical presentation provides additional support of the phenotype recognized previously and delineates the clinical spectrum associated with YHFS syndrome. In addition, we present a review of the literature including follow-up data on four previously reported individuals with YHFS.

De novo KCNA6 variants with attenuated KV 1.6 channel deactivation in patients with epilepsy.

OBJECTIVE: Mutations in the genes encoding neuronal ion channels are a common cause of Mendelian neurological diseases. We sought to identify novel de novo sequence variants in cases with early infantile epileptic phenotypes and neurodevelopmental anomalies. METHODS: Following clinical diagnosis, we performed whole exome sequencing of the index cases and their parents. Identified channel variants were expressed in Xenopus oocytes and their functional properties assessed using two-electrode voltage clamp. RESULTS: We identified novel de novo variants in KCNA6 in four unrelated individuals variably affected with neurodevelopmental disorders and seizures with onset in the first year of life. Three of the four identified mutations affect the pore-lining S6 α-helix of KV 1.6. A prominent finding of functional characterization in Xenopus oocytes was that the channel variants showed only minor effects on channel activation but slowed channel closure and shifted the voltage dependence of deactivation in a hyperpolarizing direction. Channels with a mutation affecting the S6 helix display dominant effects on channel deactivation when co-expressed with wild-type KV 1.6 or KV 1.1 subunits. SIGNIFICANCE: This is the first report of de novo nonsynonymous variants in KCNA6 associated with neurological or any clinical features. Channel variants showed a consistent effect on channel deactivation, slowing the rate of channel closure following normal activation. This specific gain-of-function feature is likely to underlie the neurological phenotype in our patients. Our data highlight KCNA6 as a novel channelopathy gene associated with early infantile epileptic phenotypes and neurodevelopmental anomalies.

De novo missense variants in the E3 ubiquitin ligase adaptor KLHL20 cause a developmental disorder with intellectual disability, epilepsy, and autism spectrum disorder.

PURPOSE: KLHL20 is part of a CUL3-RING E3 ubiquitin ligase involved in protein ubiquitination. KLHL20 functions as the substrate adaptor that recognizes substrates and mediates the transfer of ubiquitin to the substrates. Although KLHL20 regulates neurite outgrowth and synaptic development in animal models, a role in human neurodevelopment has not yet been described. We report on a neurodevelopmental disorder caused by de novo missense variants in KLHL20. METHODS: Patients were ascertained by the investigators through Matchmaker Exchange. Phenotyping of patients with de novo missense variants in KLHL20 was performed. RESULTS: We studied 14 patients with de novo missense variants in KLHL20, delineating a genetic syndrome with patients having mild to severe intellectual disability, febrile seizures or epilepsy, autism spectrum disorder, hyperactivity, and subtle dysmorphic facial features. We observed a recurrent de novo missense variant in 11 patients (NM_014458.4:c.1069G>A p.[Gly357Arg]). The recurrent missense and the 3 other missense variants all clustered in the Kelch-type ß-propeller domain of the KLHL20 protein, which shapes the substrate binding surface. CONCLUSION: Our findings implicate KLHL20 in a neurodevelopmental disorder characterized by intellectual disability, febrile seizures or epilepsy, autism spectrum disorder, and hyperactivity.

De novo variants in genes regulating stress granule assembly associate with neurodevelopmental disorders.

Stress granules (SGs) are cytoplasmic assemblies in response to a variety of stressors. We report a new neurodevelopmental disorder (NDD) with common features of language problems, intellectual disability, and behavioral issues caused by de novo likely gene-disruptive variants in UBAP2L, which encodes an essential regulator of SG assembly. Ubap2l haploinsufficiency in mouse led to social and cognitive impairments accompanied by disrupted neurogenesis and reduced SG formation during early brain development. On the basis of data from 40,853 individuals with NDDs, we report a nominally significant excess of de novo variants within 29 genes that are not implicated in NDDs, including 3 essential genes (G3BP1, G3BP2, and UBAP2L) in the core SG interaction network. We validated that NDD-related de novo variants in newly implicated and known NDD genes, such as CAPRIN1, disrupt the interaction of the core SG network and interfere with SG formation. Together, our findings suggest the common SG pathology in NDDs.

FIBCD1 is an endocytic GAG receptor associated with a novel neurodevelopmental disorder.

Whole-exome sequencing of two patients with idiopathic complex neurodevelopmental disorder (NDD) identified biallelic variants of unknown significance within FIBCD1, encoding an endocytic acetyl group-binding transmembrane receptor with no known function in the central nervous system. We found that FIBCD1 preferentially binds and endocytoses glycosaminoglycan (GAG) chondroitin sulphate-4S (CS-4S) and regulates GAG content of the brain extracellular matrix (ECM). In silico molecular simulation studies and GAG binding analyses of patient variants determined that such variants are loss-of-function by disrupting FIBCD1-CS-4S association. Gene knockdown in flies resulted in morphological disruption of the neuromuscular junction and motor-related behavioural deficits. In humans and mice, FIBCD1 is expressed in discrete brain regions, including the hippocampus. Fibcd1 KO mice exhibited normal hippocampal neuronal morphology but impaired hippocampal-dependent learning. Further, hippocampal synaptic remodelling in acute slices from Fibcd1 KO mice was deficient but restored upon enzymatically modulating the ECM. Together, we identified FIBCD1 as an endocytic receptor for GAGs in the brain ECM and a novel gene associated with an NDD, revealing a critical role in nervous system structure, function and plasticity.

Mutation update: Variants of the ENPP1 gene in pathologic calcification, hypophosphatemic rickets, and cutaneous hypopigmentation with punctate keratoderma.

ENPP1 encodes ENPP1, an ectonucleotidase catalyzing hydrolysis of ATP to AMP and inorganic pyrophosphate (PPi), and an endogenous plasma protein physiologically preventing ectopic calcification of connective tissues. Mutations in ENPP1 have been reported in association with a range of human genetic diseases. In this mutation update, we provide a comprehensive review of all the pathogenic variants, likely pathogenic variants, and variants of unknown significance in ENPP1 associated with three autosomal recessive disorders-generalized arterial calcification of infancy (GACI), autosomal recessive hypophosphatemic rickets type 2 (ARHR2), and pseudoxanthoma elasticum (PXE), as well as with a predominantly autosomal dominant disorder-Cole disease. The classification of all variants is determined using the latest ACMG guidelines. A total of 140 ENPP1 variants were curated consisting of 133 previously reported variants and seven novel variants, with missense variants being the most prevalent (70.0%, 98/140). While the pathogenic variants are widely distributed in the ENPP1 gene of patientsgen without apparent genotype-phenotype correlation, eight out of nine variants associated with Cole disease are confined to the somatomedin-B-like (SMB) domains critical for homo-dimerization of the ENPP1 protein.

Semaphorin-Plexin Signaling: From Axonal Guidance to a New X-Linked Intellectual Disability Syndrome.

BACKGROUND: Semaphorins and plexins are ligands and cell surface receptors that regulate multiple neurodevelopmental processes such as axonal growth and guidance. PLXNA3 is a plexin gene located on the X chromosome that encodes the most widely expressed plexin receptor in fetal brain, plexin-A3. Plexin-A3 knockout mice demonstrate its role in semaphorin signaling in vivo. The clinical manifestations of semaphorin/plexin neurodevelopmental disorders have been less widely explored. This study describes the neurological and neurodevelopmental phenotypes of boys with maternally inherited hemizygous PLXNA3 variants. METHODS: Data-sharing through GeneDx and GeneMatcher allowed identification of individuals with autism or intellectual disabilities (autism/ID) and hemizygous PLXNA3 variants in collaboration with their physicians and genetic counselors, who completed questionnaires about their patients. In silico analyses predicted pathogenicity for each PLXNA3 variant. RESULTS: We assessed 14 boys (mean age, 10.7 [range 2 to 25] years) with maternally inherited hemizygous PLXNA3 variants and autism/ID ranging from mild to severe. Other findings included fine motor dyspraxia (92%), attention-deficit/hyperactivity traits, and aggressive behaviors (63%). Six patients (43%) had seizures. Thirteen boys (93%) with PLXNA3 variants showed novel or very low allele frequencies and probable damaging/disease-causing pathogenicity in one or more predictors. We found a genotype-phenotype correlation between PLXNA3 cytoplasmic domain variants (exons 22 to 32) and more severe neurodevelopmental disorder phenotypes (P < 0.05). CONCLUSIONS: We report 14 boys with maternally inherited, hemizygous PLXNA3 variants and a range of neurodevelopmental disorders suggesting a novel X-linked intellectual disability syndrome. Greater understanding of PLXNA3 variant pathogenicity in humans will require additional clinical, computational, and experimental validation.

MYT1L-associated neurodevelopmental disorder: description of 40 new cases and literature review of clinical and molecular aspects.

Pathogenic variants of the myelin transcription factor-1 like (MYT1L) gene include heterozygous missense, truncating variants and 2p25.3 microdeletions and cause a syndromic neurodevelopmental disorder (OMIM#616,521). Despite enrichment in de novo mutations in several developmental disorders and autism studies, the data on clinical characteristics and genotype-phenotype correlations are scarce, with only 22 patients with single nucleotide pathogenic variants reported. We aimed to further characterize this disorder at both the clinical and molecular levels by gathering a large series of patients with MYT1L-associated neurodevelopmental disorder. We collected genetic information on 40 unreported patients with likely pathogenic/pathogenic MYT1L variants and performed a comprehensive review of published data (total = 62 patients). We confirm that the main phenotypic features of the MYT1L-related disorder are developmental delay with language delay (95%), intellectual disability (ID, 70%), overweight or obesity (58%), behavioral disorders (98%) and epilepsy (23%). We highlight novel clinical characteristics, such as learning disabilities without ID (30%) and feeding difficulties during infancy (18%). We further describe the varied dysmorphic features (67%) and present the changes in weight over time of 27 patients. We show that patients harboring highly clustered missense variants in the 2-3-ZNF domains are not clinically distinguishable from patients with truncating variants. We provide an updated overview of clinical and genetic data of the MYT1L-associated neurodevelopmental disorder, hence improving diagnosis and clinical management of these patients.

CDK19-related disorder results from both loss-of-function and gain-of-function de novo missense variants.

PURPOSE: To expand the recent description of a new neurodevelopmental syndrome related to alterations in CDK19. METHODS: Individuals were identified through international collaboration. Functional studies included autophosphorylation assays for CDK19 Gly28Arg and Tyr32His variants and in vivo zebrafish assays of the CDK19G28R and CDK19Y32H. RESULTS: We describe 11 unrelated individuals (age range: 9 months to 14 years) with de novo missense variants mapped to the kinase domain of CDK19, including two recurrent changes at residues Tyr32 and Gly28. In vitro autophosphorylation and substrate phosphorylation assays revealed that kinase activity of protein was lower for p.Gly28Arg and higher for p.Tyr32His substitutions compared with that of the wild-type protein. Injection of CDK19 messenger RNA (mRNA) with either the Tyr32His or the Gly28Arg variants using in vivo zebrafish model significantly increased fraction of embryos with morphological abnormalities. Overall, the phenotype of the now 14 individuals with CDK19-related disorder includes universal developmental delay and facial dysmorphism, hypotonia (79%), seizures (64%), ophthalmologic anomalies (64%), and autism/autistic traits (56%). CONCLUSION: CDK19 de novo missense variants are responsible for a novel neurodevelopmental disorder. Both kinase assay and zebrafish experiments showed that the pathogenetic mechanism may be more diverse than previously thought.

Novel NUDT2 variant causes intellectual disability and polyneuropathy.

Exome or genome sequencing was performed to identify the genetic etiology for the clinical presentation of global developmental delay, intellectual disability, and sensorimotor neuropathy with associated distal weakness in two unrelated families. A homozygous frameshift variant c.186delA (p.A63Qfs*3) in the NUDT2 gene was identified in cases 1 and 2 from one family and a third case from another family. Variants in NUDT2 were previously shown to cause intellectual disability, but here we expand the phenotype by demonstrating its association with distal upper and lower extremity weakness due to a sensorimotor polyneuropathy with demyelinating and/or axonal features.

Enhanced MAPK1 Function Causes a Neurodevelopmental Disorder within the RASopathy Clinical Spectrum.

Signal transduction through the RAF-MEK-ERK pathway, the first described mitogen-associated protein kinase (MAPK) cascade, mediates multiple cellular processes and participates in early and late developmental programs. Aberrant signaling through this cascade contributes to oncogenesis and underlies the RASopathies, a family of cancer-prone disorders. Here, we report that de novo missense variants in MAPK1, encoding the mitogen-activated protein kinase 1 (i.e., extracellular signal-regulated protein kinase 2, ERK2), cause a neurodevelopmental disease within the RASopathy phenotypic spectrum, reminiscent of Noonan syndrome in some subjects. Pathogenic variants promote increased phosphorylation of the kinase, which enhances translocation to the nucleus and boosts MAPK signaling in vitro and in vivo. Two variant classes are identified, one of which directly disrupts binding to MKP3, a dual-specificity protein phosphatase negatively regulating ERK function. Importantly, signal dysregulation driven by pathogenic MAPK1 variants is stimulus reliant and retains dependence on MEK activity. Our data support a model in which the identified pathogenic variants operate with counteracting effects on MAPK1 function by differentially impacting the ability of the kinase to interact with regulators and substrates, which likely explains the minor role of these variants as driver events contributing to oncogenesis. After nearly 20 years from the discovery of the first gene implicated in Noonan syndrome, PTPN11, the last tier of the MAPK cascade joins the group of genes mutated in RASopathies.

New roles for the external globus pallidus in basal ganglia circuits and behavior.

The development of methodology to identify specific cell populations and circuits within the basal ganglia is rapidly transforming our ability to understand the function of this complex circuit. This mini-symposium highlights recent advances in delineating the organization and function of neural circuits in the external segment of the globus pallidus (GPe). Although long considered a homogeneous structure in the motor-suppressing "indirect-pathway," the GPe consists of a number of distinct cell types and anatomical subdomains that contribute differentially to both motor and nonmotor features of behavior. Here, we integrate recent studies using techniques, such as viral tracing, transgenic mice, electrophysiology, and behavioral approaches, to create a revised framework for understanding how the GPe relates to behavior in both health and disease.

Control of muscle synergies by cortical ensembles.

Since its introduction in the early 1980s, the concept of a "preferred direction" for neuronal discharge has proven to be a powerful means of studying diverse properties of individual neurons in the motor areas of the brain. More recently, the activity recorded from ensembles of neurons, each with an identified preferred direction, has been used to predict hand movement, both off-line, and in real-time. Our recent experiments have addressed similar issues, but have focused on the relation between primary motor cortical discharge and muscle activity, rather than limb kinematics. We recently introduced the concept of a "muscle-space" preferred direction (PD(M)), that is analogous to the familiar hand-space preferred direction (PD(H)). In this manuscript, we show that there is considerable variety in the direction of these PD(M) vectors across neurons, but that for a given task and neuron, two successive measurements of PD(M) are very similar. We found that these vectors tend to form clusters in particular regions of the muscle space that may reflect neurons that control synergistically important groups of muscles. We have also shown that the discharge measured from neural ensembles can be used to predict the activity of individual muscles, in much the way that kinematic signals have been predicted by other groups. In fact, the accuracy of these predictions is similar to that of kinematic signals, despite the stochastic nature and greater bandwidth of the EMG signals. PD(M)s represent a divergence from one neuron to numerous muscles, while the prediction of muscle activity represents convergence from many neurons to individual muscles. We are continuing to investigate the nature of this complex matrix of functional interconnections.