Biology subjects

Poduri, A.

Publications and source records attributed to Poduri, A..

16 recordsLinked to original sources

bioRxiv2026

CAMSAP3 loss of function models suggest causative role in generalized genetic epilepsy

Advancements in next generation sequencing have led to the discovery of hundreds of human epilepsy gene associations. Newly associated genes require functional validation to establish causation and to inform patient treatment in the clinic. A recent exome trio analysis identified predicted pathogenic variants in two patients with generalized epilepsy in the gene CAMSAP3. CAMSAP3 regulates non-centrosomal microtubule dynamics, and the acetylation necessary for normal axonal differentiation and migration. We show that overexpression of patient variants leads to protein degradation and dysregulation of microtubule acetylation in cultured HEK cells. Camsap3 knockout zebrafish also exhibit increased axonal microtubule acetylation as well as epileptic features such as seizure-like swimming behaviors, aberrant inhibitory interneuron development and epileptiform via local field potential. Together these data suggest that CAMSAP3 plays an important role in generalized genetic epilepsy.

neuroscience↗

bioRxiv2026

Activating Ras-MAPK pathway variants drive hippocampal clonal competition in human epilepsy

Mesial (a.k.a., medial) temporal lobe epilepsy (MTLE) is the most common focal epilepsy1,2 and, in drug-resistant cases, is treated by surgical removal of the anterior temporal lobe, which often shows neuronal loss and gliosis consistent with hippocampal sclerosis (HS)2. MTLE with HS has minimal contribution from germline genetic variation3, and is associated with prior precipitating insults such as prolonged childhood seizures and head trauma4-6. Somatic variants in Ras-MAPK pathway genes were recently reported in a few MTLE surgical specimens7,8, but their prevalence, clinical relevance, and underlying biological mechanisms remain unknown. Targeted duplex sequencing of hippocampal DNA from 462 surgical resections revealed significant enrichment of deleterious somatic variants in MTLE versus controls, with >40% of MTLE specimens harboring activating Ras-MAPK variants in PTPN11, NF1, BRAF, KRAS, and twelve genes not previously associated with focal epilepsy. Eight Ras-MAPK genes showed positive clonal selection in MTLE. Increased somatic variant burden predicted worse surgical outcome. Somatic Ras-MAPK variants at ultra-low (<0.5%) variant allele fractions were associated with older seizure onset and HS pathology, supporting a late prenatal or postnatal origin. Ras-MAPK variants in MTLE were enriched in cells derived from hippocampal progenitors--neurons, astrocytes, oligodendrocytes--in line with the known neuronal hyperexcitability and seizures induced by Ras-MAPK overactivation9,10; in contrast, Alzheimer disease hippocampi exhibited microglial enrichment of Ras-MAPK variants, consistent with prior reports11. Single-nucleus RNA sequencing showed increased expression of Ras-MAPK genes in neurons and upregulation of pathways mediating neurogenesis and neural development in MTLE. Functional validation of novel, recurrent PTPN11 variants confirmed gain-of-function, while cellular modeling in induced pluripotent stem cells demonstrated proliferative/survival advantages for mutant cells in mosaic culture. Overall, our data suggest that somatic Ras-MAPK variants and acquired risk factors may converge on clonal competition in the hippocampus to modulate epilepsy risk.

genetics↗

bioRxiv2025

Early death and neuronal abnormalities in depdc5 loss-of-function mosaic zebrafish models

DEPDC5 (DEP domain-containing protein 5) encodes a repressor of the mTORC1 signaling pathway. Variants in DEPDC5 are associated with a range of focal epilepsies, including mosaic variants associated with focal cortical dysplasia (FCD) and other focal brain malformations with brain-only somatic mosaic variants. To investigate the role of DEPDC5 in human epilepsy related to mosaic variants, we have generated mosaic depdc5 loss-of-function zebrafish models using homology-based constructs acutely targeting depdc5 and labeled with tdTomato to allow for visualization of the degree of mosaicism. The resulting mosaic depdc5 CRISPants demonstrated early larval death, with [~]50% of CRISPants (vs. 10% of controls) dead by 7 days post fertilization (dpf), analogous to the early death sometimes associated with human DEPDC5-related epilepsy. We compared depdc5 CRISPants with uninjected and scrambled controls from the same clutches. Body and head size were reduced in the depdc5 CRISPants. Analysis of swimming behavior showed a striking reduction in distance traveled and maximum velocity in the depdc5 CRISPants vs. controls. Based on visual confirmation of mutational load, we categorized CRISPants into depdc5+ vs. depdc5++, reflecting weak vs. strong tdTomato fluorescence. We observed that depdc5++ CRISPants had increased episodes of posture loss, suggesting increased seizure-like behavior related to higher percentages of mutant cells. Local field potential recordings revealed increased neuronal hyperexcitability in depdc5 CRISPants vs. controls. Acridine orange staining demonstrated early apoptosis in the CRISPants vs. controls. Our mosaic depdc5 CRISPants provide a clinically relevant model to study the role of mosaic DEPDC5-related epilepsy and early death.

neuroscience↗

bioRxiv2025

High-throughput imaging of GABA fluorescence as a functional assay for variants in the neurodevelopmental gene, SLC6A1

Pathogenic variants in SLC6A1, which encodes GABA Transporter 1 (GAT-1), are associated with developmental delay, autism, epilepsy (e.g., epilepsy with myoclonic astatic seizures [EMAS]), and possibly schizophrenia. Functional assays to establish pathogenicity of human variants is a key limiting factor in the clinical interpretation of genetic findings. Methods based on radioactive [3H]-GABA uptake come with significant regulatory concerns, cost, and workflow complexity, which could be resolved by an alternate assay. To address this issue, we developed a high-content fluorescence imaging assay of GAT-1-mediated GABA uptake using a genetically encoded GABA sensor, iGABA-Snfr. We demonstrated that pathogenic variants strongly reduced uptake (mean, -89.4% [95% CI, -71.5% to -107.3%]). Some variants of uncertain significance (VUS) were associated with reduced GABA uptake (G111R, S459R, V511M; mean, -101.2% [95% CI -81.1% to -121.3%]), whereas others showed only mild reduction (R211C, R566H, F242V, R419C; mean, -33.6% [95% CI -17.2% to -50.1%]), supporting variant reclassification. Variant-specific effects on iGABA were highly correlated with the results of the radioactive [3H]-GABA assay (R2=0.8095, p<0.0001). The molecular chaperone 4-phenylbutyric acid (4PBA) was associated with [~]35% increase in iGABA. This non-radioactive assay is suitable for functional validation and high-throughput screening to identify positive modulators of GAT-1.

neuroscience↗

bioRxiv2025

Characterization of the functional and clinical impacts of CACNA1A missense variants found in neurodevelopmental disorders

CACNA1A encodes the P/Q-type CaV2.1 calcium channels whose function underlies neuronal excitability, presynaptic neurotransmitter release, and Ca2+ signaling in neurons. Pathogenic variants in CACNA1A have been found in individuals with various neurological conditions, including hemiplegic migraine, epilepsy, developmental delay, and ataxia. Clinical presentations can vary significantly between patients, with limited information known about the underlying neurobiology of these different clinical patterns. Adding further complication, prior work on pathogenic missense variants has demonstrated variable impacts on CaV2.1 channel function, sometimes in opposite directions. As such, the relationships between specific coding variants, electrophysiological properties, and clinical phenotypes remain elusive. In this study, we determined the biophysical properties of an allelic series of 42 de novo missense CACNA1A variants discovered in a neurodevelopmental disorder cohort of more than 31,000 individuals, together with the most common eight coding variants found in the general population. We found that all but one de novo variant altered at least one aspect of the channel properties examined, and the majority (70%) of the variants reduced the channel current density. In addition, for variants that encode human CaV2.1 channels (hCaV2.1) with detectable currents, nearly 50% altered how channels respond to membrane potential, while common variations did not significantly change any channel biophysical properties. Coupled with our functional analyses and AlphaMissense prediction, we showed that CaV2.1 missense variants significantly underlie the risk of developmental epileptic encephalopathy. Subsequently, we examined the physiological impact of variant hCaV2.1 using NEURON simulations as an omnibus output of neuronal function and found that abnormal biophysical channel properties have a profound impact on Purkinje cell excitability. Most interestingly, we correlated the clinical phenotype with molecular consequences of missense variants provided by our comprehensive functional analyses and found that distinct CaV2.1 channel molecular function is significantly associated with different clinical outcomes. By analyzing an entire allelic series of CACNA1A de novo changes in a large cohort of individuals with neurodevelopmental disorders, we provide a powerful approach to dissecting the role of missense variants in CACNA1A channelopathy, which in turn may help pave the way for future precision medicine initiatives.

neuroscience↗

bioRxiv2025

Phenotypic analysis of 11,125 trio exomes in neurodevelopmental disorders

Genomic sequencing is widely used to identify causative genetic changes in neurodevelopmental disorders, such as autism, intellectual disability, and epilepsy. Most neurodevelopmental disorders also present with diverse clinical features, and delineating the interaction between causative genetic changes and phenotypic features is a key prerequisite for developing personalized therapies. However, assessing clinical features at a scale that parallels genomic sequencing remains challenging. Here, we standardize phenotypic information across 11,125 patient-parent trios with exome sequencing data using biomedical ontologies, analyzing 674,767 phenotypic terms. We find that individuals with de novo variants in 69 out of 261 neurodevelopmental genes exhibit statistically significant clinical similarities with distinct phenotypic fingerprints. We also observe that phenotypic relatedness follows a gradient, spanning from highly similar to dissimilar phenotypes, with intra-gene similarities suggesting clinically distinct subgroups for seven neurodevelopmental genes. For most genetic etiologies, only a small subset of highly phenotypically similar individuals carried de novo variants in the same gene, highlighting the heterogeneous and complex clinical landscape of neurodevelopmental disorders. Our study provides a large-scale overview of the dynamic relationship between genotypes and phenotypes in neurodevelopmental disorders, underscoring how the inherent complexity of these conditions can be deciphered through approaches that integrate genomic and phenotypic data.

genetics↗

bioRxiv2025

Enhanced proconvulsant sensitivity, not spontaneous rapid swimming activity, is a robust correlate of scn1lab loss-of-function in stable mutant and F0 crispant hypopigmented zebrafish expressing GCaMP6s

Zebrafish models of genetic epilepsy benefit from the ability to assess disease-relevant knock-out alleles with numerous tools, including genetically encoded calcium indicators (GECIs) and hypopigmentation alleles to improve visualization. However, there may be unintended effects of these manipulations on the phenotypes under investigation. There is also debate regarding the use of stable loss-of-function (LoF) alleles in zebrafish, due to genetic compensation (GC). In the present study, we applied a method for combined movement and calcium fluorescence profiling to the study of a zebrafish model of SCN1A, the main gene associated with Dravet syndrome, which encodes the voltage-gated sodium channel alpha1 subunit (Nav1.1). We evaluated for spontaneous and proconvulsant-induced seizure-like activity associated with scn1lab LoF mutations in larval zebrafish expressing a neuronally-driven GECI (elavl3:GCaMP6s) and a nacre mutation causing a common pigmentation defect. In parallel studies of stable scn1labs552 mutants and F0 crispant larvae generated using a CRISPR/Cas9 multi-sgRNA approach, we find that neither stable nor acute F0 larvae recapitulate the previously reported seizure-like rapid swimming phenotype nor does either group show spontaneous calcium events meeting criteria for seizure-like activity based on a logistic classifier trained on movement and fluorescence features of proconvulsant-induced seizures. This constitutes two independent lines of evidence for a suppressive effect against the scn1lab phenotype, possibly due to the GCaMP6s-derived genetic background (AB) or nacre hypopigmentation. In response to the proconvulsant pentylenetetrazole (PTZ), we see evidence of a separate suppressive effect affecting all conspecific larvae derived from the stable scn1labs552 line, independent of genotype, possibly related to a maternal effect of scn1lab LoF in mutant parents or the residual TL background. Nonetheless, both stable and F0 crispant fish show enhanced sensitivity to PTZ relative to conspecific larvae, suggesting that proconvulsant sensitivity provides a more robust readout of scn1lab LoF under our experimental conditions. Our study underscores the unexpected challenges associated with the combination of common zebrafish tools with disease alleles in the phenotyping of zebrafish models of genetic epilepsy. Our work further highlights the advantages of using F0 crispants and the evaluation of proconvulsant sensitivity as complementary approaches that faithfully reflect the shared gene-specific pathophysiology underlying spontaneous seizures in stable mutant lines. Future work to understand the molecular mechanisms by which scn1lab-related seizures and PTZ-related hyperexcitability are suppressed under these conditions may shed light on factors contributing to variability in preclinical models of epilepsy more generally and may identify genetic modifiers relevant to Dravet syndrome.

neuroscience↗

bioRxiv2024

Machine learning enables high-throughput, low-replicate screening for novel anti-seizure targets and compounds using combined movement and calcium fluorescence in larval zebrafish

AbstractIdentifying new, more efficacious anti-seizure medications (ASMs) is challenging, partly due to limitations in animal-based assays. Zebrafish (Danio rerio) can serve as a model of chemical and genetic seizures, but methods for detecting seizure-like activity in zebrafish, though powerful, have been hampered by low sensitivity (locomotor/behavioral assays) or low-throughput (tectal electrophysiology or calcium fluorescence microscopy). To address these issues, we developed a novel approach to assay seizure-like activity using combined locomotor and calcium fluorescence features, measured simultaneously from unrestrained larval zebrafish using a 96-well fluorescent plate reader. Using custom software to track fish movement and changes in fluorescence (deltaF/F0) from high-speed time-series (12.6Hz), we trained logistic classifiers using elastic net regression to distinguish seizure-like activity from non-seizure related changes based on event-specific and subject-specific features in response to the GABAAR antagonist, pentylenetetrazole (PTZ). We demonstrate that a classifier trained on combined movement and fluorescence data achieves high accuracy ("PTZ M+F"; area-under-curve receiver-operator characteristic (AUC-ROC): 0.98; F1 score: 0.912) and out-performs classifiers trained on movement ("PTZ M"; AUC-ROC: 0.9, F1: 0.9) or fluorescence features alone ("PTZ F"; AUC-ROC 0.96; F1: 0.87). The rate of classified seizure-like events increases as a dose-response to PTZ (serial dose escalation, 0, 2.5mM, 15mM) and is strongly suppressed by ASM treatment (valproic acid, VPA; tiagabine, TGB). At high-dose PTZ, we show that VPA reduces seizure-like activity defined by either "PTZ M+F" or "PTZ M" classifiers. Meanwhile, TGB selectively reduces events defined by the "PTZ M+F" classifier, paralleling previous reports that TGB reduces electrographic but not locomotor seizures and highlighting the potential for our approach to combine features of previously orthogonal assays. Using ASM benchmark data, we employ bootstrap simulation to calculate the expected statistical power of our method as a function of sample size. We demonstrate that anti-seizure responses (robust strictly standardized mean difference, RSSMD, versus control) with magnitudes similar to those associated with VPA or TGB can be reliably detected (true positive rate (TPR) > 90%) with as few as N=4 biological replicates per group, while maintaining a 5% false positive rate. In a prospective test screen with 3-6 replicates per group and on-plate controls, the anti-seizure effect of 4 out of 5 tested ASMs (CBZ, LEV, LZP, TGB) was detected. In summary, we demonstrate a simple high-throughput approach to whole organism anti-seizure phenotyping combining two previously reported metrics to facilitate screens for novel anti-seizure interventions in zebrafish.

pharmacology and toxicology↗

bioRxiv2024

Zebrafish models of candidate human epilepsy-associated genes provide evidence of hyperexcitability

Hundreds of novel candidate human epilepsy-associated genes have been identified thanks to advancements in next-generation sequencing and large genome-wide association studies, but establishing genetic etiology requires functional validation. We generated a list of >2200 candidate epilepsy-associated genes, of which 81 were determined suitable for the generation of loss-of-function zebrafish models via CRISPR/Cas9 gene editing. Of those 81 crispants, 48 were successfully established as stable mutant lines and assessed for seizure-like swim patterns in a primary F2 screen. Evidence of seizure-like behavior was present in 5 (arfgef1, kcnd2, kcnv1, ubr5, wnt8b) of the 48 mutant lines assessed. Further characterization of those 5 lines provided evidence for epileptiform activity via electrophysiology in kcnd2 and wnt8b mutants. Additionally, arfgef1 and wnt8b mutants showed a decrease in the number of inhibitory interneurons in the optic tectum of larval animals. Furthermore, RNAseq revealed convergent transcriptional abnormalities between mutant lines, consistent with their developmental defects and hyperexcitable phenotypes. These zebrafish models provide strongest experimental evidence supporting the role of ARFGEF1, KCND2, and WNT8B in human epilepsy and further demonstrate the utility of this model system for evaluating candidate human epilepsy genes. HighlightsO_LIZebrafish models generated by CRISPR/Cas9 gene editing display seizure-like swim patterns in five candidate human epilepsy genes: arfgef1, kcnd2, kcnv1, ubr5, wnt8b. C_LIO_LILocal field potential abnormalities recorded from kcnd2 and wnt8b crispants provide additional evidence of hyperexcitability. C_LIO_LIArfgef1 and wnt8b mutant larvae have fewer inhibitory interneurons than wild type in the optic tectum. C_LIO_LICRISPR-generated mutants of epilepsy genes displayed convergent transcriptional dysregulation, consistent with developmental abnormalities and their hyperexcitability phenotype. C_LI

genetics↗

bioRxiv2021

Mosaic and non-mosaic pcdh19 mutation leads to neuronal hyperexcitability in zebrafish

Epilepsy is one of the most common neurological disorders. The X-linked gene PCDH19 is associated with sporadic and familial epilepsy in humans, typically with early-onset clustering seizures and intellectual disability in females but not in so-called carrier males, suggesting that mosaic PCDH19 expression is required to produce epilepsy. To characterize the role of loss of PCDH19 function in epilepsy, we generated zebrafish with truncating pcdh19 variants. Evaluating zebrafish larvae for electrophysiological abnormalities, we observed hyperexcitability phenotypes in both mosaic and non-mosaic pcdh19+/- and pcdh19-/- mutant larvae. Thus, we demonstrate that the key feature of epilepsy--network hyperexcitability--can be modeled effectively in zebrafish, even though overt spontaneous seizure-like swim patterns were not observed. Further, zebrafish with non-mosaic pcdh19 mutation displayed reduced numbers of inhibitory interneurons suggesting a potential cellular basis for the observed hyperexcitability. Our findings in both mosaic and non-mosaic pcdh19 mutant zebrafish challenge the prevailing theory that mosaicism governs all PCDH19-related phenotypes and point to interneuron-mediated mechanisms underlying these phenotypes.

neuroscience↗

bioRxiv2019

Modeling and treating GRIN2A developmental and epileptic encephalopathy in mice

NMDA receptors (NMDAR) play crucial roles in excitatory synaptic transmission. Rare variants of GRIN2A, which encodes the GluN2A NMDAR subunit, are associated with several intractable neurodevelopmental disorders, including developmental and epileptic encephalopathy (DEE). A de novo missense variant, p.Ser644Gly (c.1930A>G), was identified in a child with DEE, and Grin2a knockin mice were generated to model and extend understanding of this intractable childhood disease. Homozygous and heterozygous mutant mice exhibit altered hippocampal morphology at two weeks of age, and homozygotes exhibit lethal tonic-clonic seizures in the third week. Heterozygous adult mice display a variety of distinct features, including resistance to electrically induced partial seizures, as well as hyperactivity and repetitive and reduced anxiety behaviors. Multielectrode recordings of mutant neuronal networks reveal hyperexcitability and altered bursting and synchronicity. When expressed in heterologous cells, mutant receptors exhibit enhanced NMDAR agonist potency and slow deactivation following rapid removal of glutamate, as occurs at synapses. Consistent with these observations, NMDAR-mediated synaptic currents in hippocampal slices from mutant mice show a prolonged deactivation time course. Standard antiepileptic drug monotherapy was ineffective in the patient, but combined treatment of NMDAR antagonists with antiepileptic drugs substantially reduced the seizure burden albeit without appreciable developmental improvement. Chronic treatment of homozygous mutant mouse pups with NMDAR antagonists delayed the onset of lethal seizures but did not prevent them. These studies illustrate the power of modeling severe neurodevelopmental seizure disorders using multiple experimental modalities and suggest their extended utility in identifying and evaluating new therapies.

neuroscience↗

bioRxiv2019

High throughput Characterization of KCNB1 variants Associated with Developmental and Epileptic Encephalopathy

Pathogenic variants in KCNB1, encoding the voltage-gated potassium channel Kv2.1, are associated with developmental and epileptic encephalopathies (DEE). Previous functional studies on a limited number of KCNB1 variants indicated a range of molecular mechanisms by which variants affect channel function, including loss of voltage sensitivity, loss of ion selectivity, and reduced cell-surface expression. We evaluated a series of 17 KCNB1 variants associated with DEE or neurodevelopmental disorder (NDD) to rapidly ascertain channel dysfunction using high-throughput functional assays. Specifically, we investigated the biophysical properties and cell-surface expression of variant Kv2.1 channels expressed in heterologous cells using high-throughput automated electrophysiology and immunocytochemistry-flow cytometry. Pathogenic variants exhibited diverse functional defects, including altered current density and shifts in the voltage-dependence of activation and/or inactivation, as homotetramers or when co-expressed with wild-type Kv2.1. Quantification of protein expression also identified variants with reduced total Kv2.1 expression or deficient cell-surface expression.\n\nOur study establishes a platform for rapid screening of functional defects of KCNB1 variants associated with DEE and other NDDs, which will aid in establishing KCNB1 variant pathogenicity and may enable discovery of targeted strategies for therapeutic intervention based on molecular phenotype.

genetics↗

bioRxiv2018

The role of sodium channels in sudden unexpected death in pediatrics

Sudden Unexpected Death in Pediatrics (SUDP) is a tragic condition with hypothesized multifactorial etiology. While there is recent evidence implicating genes related to cardiac arrhythmia and epilepsy as genetic risk factors contributing to some cases of SUDP, the underlying mechanisms of SUDP remain under active investigation. SUDP encompasses Sudden Infant Death Syndrome (SIDS) and Sudden Unexplained Death in Childhood (SUDC), affecting children under and over 1 year of age, respectively. The presence of developmental hippocampal malformations in many children with SIDS and SUDC suggests that a subset of patients may share epilepsy-related mechanisms with Sudden Unexplained Death in Epilepsy Patients (SUDEP). Pathogenic variants in both epilepsy- and arrhythmia-related sodium channel genes have recently been identified in patients with SIDS, SUDC, and SUDEP.\n\nWe performed a candidate gene analysis for genes encoding sodium channel subunits in whole exome sequencing (WES) data from 73 SUDP patients. After a thorough literature review, we mapped all reported SUDP-associated sodium channel variants alongside variants from the population on a structural protein model to evaluate whether patient variants clustered in important protein domains compared to controls.\n\nIn our cohort, 13 variants met criteria for pathogenicity or potential pathogenicity. While SCN1A, SCN1B, and SCN5A have established disease associations, we also considered variants in the paralogs SCN3A, SCN4A and SCN9A. Overall, the patient-associated variants clustered at conserved amino acid sites across the sodium channel gene family that do not tolerate variation in these genes.\n\nThis study provides a molecular overview of sodium channel variants present in cases with SUDP and reveals key amino acid sites that do not tolerate variation across the SCN paralog family. Further research will lead to an improved understanding of the contribution of sodium channels to SUDP, with a goal of one day implementing prevention strategies to avoid untimely deaths in at-risk children.\n\nAuthor SummaryThe sudden unexplained death of an infant or a child is a tragic event, which is likely caused by the complex interaction of multiple factors. Besides environmental factors, genes related to epilepsy and cardiac arrhythmia have been identified as risk factors. The sodium channel family encompasses genes, related to both cardiac arrhythmia as well as epilepsy, whose proteins share structural homology. We evaluated sodium channel gene variants in our cohort, examined all known variants in sodium genes in SUDP patients from the literature, and mapped patient variants alongside variants from the population on a 3D protein model. The patient variants clustered at conserved amino acid sites with low rates of variation in the general population, not only in the particular gene involved but also in the gene family. This study illustrates that sodium channel variants contribute to the complex phenotype of sudden death in pediatrics, suggesting complex mechanisms of neurologic and/or cardiac dysfunction contributing to death.

genetics↗

bioRxiv2017

Molecular map of GNAO1-related disease phenotypes and reactions to treatment

The GNAO1 gene codes for the most commonly expressed G protein in the central nervous system. Pathogenic GNAO1 variants result in early-onset neurological phenotypes, sometimes with distinct epilepsy or movement disorder, and sometimes with both mani-festations in the same patient. The existing extensive knowledge about G-protein coupled receptor (GPCR) signaling provides the input needed to describe quantitatively how mutations modify the GPCR signal. This in turn allows rational interpretation of distinct phenotypes arising from mutations in GNAO1. In this work we outline a model that enables understanding of clinical phenotypes at a molecular level. The mutations affecting the catalytic pocket of GNAO1, we show, result in the improper withdrawal of the signal, and give rise to epileptic phenotypes (EPs). The converse is not true - some pure EPs are caused by mutations with no obvious impact on catalysis. Mutations close to the interface with GNAO1s downstream effector block the signal propagation in that direction, and manifest as a movement disorder phenotype without epilepsy. Quantifying the reported reaction to therapy highlights the tendency of the latter group to be unresponsive to the therapies currently in use. We argue, however, that the majority of clinically described mutations can impact several aspects of GNAO1 function at once, resulting in the continuum of phenotypes observed in patients. The reasoning based on GNAO1 signaling model provides a precision medicine paradigm to aid clinicians in selecting effective categories of medication, and in addition, can suggest pragmatic targets for future therapies.

molecular biology↗

bioRxiv2017

Gene family information facilitates variant interpretation and identification of disease-associated genes

Differentiating risk-conferring from benign missense variants, and therefore optimal calculation of gene-variant burden, represent a major challenge in particular for rare and genetic heterogeneous disorders. While orthologous gene conservation is commonly employed in variant annotation, approximately 80% of known disease-associated genes are paralogs and belong to gene families. It has not been thoroughly investigated how gene family information can be utilized for disease gene discovery and variant interpretation. We developed a paralog conservation score to empirically evaluate whether paralog conserved or nonconserved sites of in-human paralogs are important for protein function. Using this score, we demonstrate that disease-associated missense variants are significantly enriched at paralog conserved sites across all disease groups and disease inheritance models tested. Next, we assessed whether gene family information could assist in discovering novel disease-associated genes. We subsequently developed a gene family de novo enrichment framework that identified 43 exome-wide enriched gene families including 98 de novo variant carrying genes in more than 10k neurodevelopmental disorder patients. 33 gene family enriched genes represent novel candidate genes which are brain expressed and variant constrained in neurodevelopmental disorders.

genetics↗

bioRxiv2017

The Spectrum Of De Novo Variants In Neurodevelopmental Disorders With Epilepsy

Epilepsy is a frequent feature of neurodevelopmental disorders (NDD) but little is known about genetic differences between NDD with and without epilepsy. We analyzed de novo variants (DNV) in 6753 parent-offspring trios ascertained for different NDD. In the subset of 1942 individuals with NDD with epilepsy, we identified 33 genes with a significant excess of DNV, of which SNAP25 and GABRB2 had previously only limited evidence for disease association. Joint analysis of all individuals with NDD also implicated CACNA1E as a novel disease gene. Comparing NDD with and without epilepsy, we found missense DNV, DNV in specific genes, age of recruitment and severity of intellectual disability to be associated with epilepsy. We further demonstrate to what extent our results impact current genetic testing as well as treatment, emphasizing the benefit of accurate genetic diagnosis in NDD with epilepsy.

genetics↗