Biology subjects

Weber, Y.

Publications and source records attributed to Weber, Y..

At least 19 recordsLinked to original sources

bioRxiv2026

Graph theory for the analysis of micro-electrode array recordings of human brain slices - framework and benchmarking

Micro-electrode array (MEA) recordings are widely used to characterize functional connectivity in neural cultures and have gained traction for the analysis of human brain slices. However, the impact of graph construction methodology on the resulting network topology has not been systematically quantified. Here, we benchmark three methods - shared spiking activity, Pearson cross-correlation, and the spike time tiling coefficient (STTC) - across 37 recordings from human cortical slice cultures classified into low, moderate, and high activity groups. We show that method choice alone produces large topological differences (Cohens d = 0.86-1.14 for clustering coefficient, d > 1.0 for node count), while higher-order features such as modularity remain stable. Each method exhibits a distinct sensitivity profile: shared spiking detects activity-dependent changes primarily through network size, correlation uniquely captures clustering differences, and STTC combines strong biological sensitivity with negligible parameter dependence across lag windows (all d < 0.1). Within shared spiking, z-score normalization dominates all other parameter choices (d > 1.0 versus bin size effects of d < 0.23), functioning as an implicit analytical null model that fundamentally reshapes the edge set rather than merely rescaling weights. Inter-method edge overlap is low (Jaccard index 0.08-0.45) and activity dependent, demonstrating that these methods identify substantially different connections from identical data. Our results reveal that methodological choices including construction method, threshold, and normalization introduce hidden degrees of freedom with effect sizes comparable to the biological signals being measured. We provide practical recommendations for parameter selection, reporting, and cross-method validation in MEA-based network neuroscience. Author SummaryWhen we record electrical activity from brain tissue using grids of electrodes, we can ask how different sites influence one another and map the tissue as a network of connections. Thanks to novel culturing methods, this approach is increasingly used to study human brain slices. However, deciding what is "connected" is not well defined. Researchers use several different methods, and it has never been clear how much this choice shapes the network they end up describing. Here we compared three widely used methods on 37 recordings from human cortical slices spanning a range of activity levels. We found that the method alone can change the apparent structure of the network as much as real biological differences do. The methods frequently disagreed about which connections exist and some technical choices, including normalization techniques, had surprisingly large effects. Because these hidden choices can rival the biological signal, we provide this benchmarking work with practical recommendations for selecting, reporting, and cross-checking methods, so that network studies of brain tissue become more transparent, comparable, and reproducible.

neuroscience↗

bioRxiv2025

Modeling AP2M1 Developmental and Epileptic Encephalopathy in Drosophila

Genetic defects in AP2M1, which encodes the -subunit of the adaptor protein complex 2 (AP-2) essential for clathrin-mediated endocytosis (CME), cause a rare form of developmental and epileptic encephalopathy (DEE). In this study, we modeled AP2M1-DEE in Drosophila melanogaster to gain deeper insights into the underlying disease mechanisms. Pan-neuronal knock-down of the Drosophila AP2M1 ortholog, AP-2{micro}, resulted in a consistent heat-sensitive paralysis phenotype and altered morphology in class IV dendritic arborization (c4da) neurons. Unexpectedly, affected flies were resistant to antiseizure medications and exhibited increased resistance to electrically induced seizures. A CRISPR-engineered fly line carrying the recurrent human disease variant p.Arg170Trp displayed a milder seizure resistance phenotype. While these findings contrast with the human phenotype, they align with previous studies on other CME-related genes in Drosophila. Our results suggest that hyperexcitability and seizures in AP2M1-DEE may stem from broader defects in neuronal development rather than direct synaptic dysfunction.

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↗

bioRxiv2024

Comprehensive Analysis of Human Dendritic Spine Morphology and Density: Exploring Diversity of Human Dendritic Spines

Dendritic spines, small protrusions on neuronal dendrites, play a crucial role in brain function by changing shape and size in response to neural activity. So far, in depth analysis of dendritic spines in human brain tissue is lacking. This study presents a comprehensive analysis of human dendritic spine morphology and density using a unique dataset from human brain tissue from 27 patients (8 females, 19 males, aged 18-71) undergoing tumor or epilepsy surgery at three neurosurgery sites. We used acute slices and organotypic brain slice cultures to examine dendritic spines, classifying them into the three main morphological subtypes: Mushroom, Thin, and Stubby, via 3D reconstruction using ZEISS arivis Pro software. A deep learning model, trained on 39 diverse datasets, automated spine segmentation and 3D reconstruction, achieving a 74% F1-score and reducing processing time by over 50%. We show significant differences in spine density by sex, dendrite type, and tissue condition. Females had higher spine densities than males, and apical dendrites were denser in spines than basal ones. Acute tissue showed higher spine densities compared to cultured human brain tissue. With time in culture, Mushroom spines decreased, while Stubby and Thin spine percentages increased, particularly from 7-9 to 14 days in vitro, reflecting potential synaptic plasticity changes. Our study underscores the importance of using human brain tissue to understand unique synaptic properties and shows that integrating deep learning with traditional methods enables efficient large-scale analysis, revealing key insights into sex- and tissue-specific dendritic spine dynamics relevant to neurological diseases. New and NoteworthyThis study presents a dataset of nearly 4,000 morphologically reconstructed human dendritic spines across different ages, gender, and tissue conditions. The dataset was further used to evaluate a deep learning algorithm for three-dimensional spine reconstruction, offering a scalable method for semi-automated spine analysis across various tissues and microscopy setups. The findings enhance understanding of human neurology, indicating potential connections between spine morphology, brain function, and the mechanisms of neurological and psychiatric diseases.

neuroscience↗

bioRxiv2024

Lipid hydrogen isotope compositions primarily reflect growth water in the model archaeon Sulfolobus acidocaldarius

The stable hydrogen isotope composition ({delta}2H) of lipid biomarkers can track environmental processes and remain stable over geologically relevant time scales, enabling studies of past climate, hydrology, and ecology. Most research has focused on lipids from the domain Eukarya (e.g., plant waxes, long-chain alkanes), and the potential of prokaryotic lipid biomarkers from the domain Archaea to offer unique insights into environments not captured by eukaryotic lipids remains unclear. Here, we investigate the H-isotope composition of biphytanes in Sulfolobus acidocaldarius, a model thermoacidophile and obligate heterotroph. We conducted a series of experiments that varied temperature, pH, shaking rate, electron acceptor availability, or electron donor flux. From these experiments, we quantified the lipid/water H-isotope fractionation (2{varepsilon}L/W) values for core biphytane chains derived from tetraether lipids. The 2{varepsilon}L/W values are consistently negative (-230 to - 180 {per thousand}) and are relatively invariant across all experiments despite a 20-fold change in doubling times and 2-fold change in lipid cyclization. The magnitude and relative invariance of 2{varepsilon}L/W values are consistent with studies on other heterotrophic archaea and suggests archaeal lipids may be faithful recorders of the {delta}2H composition of growth water. Our study highlights the potential of archaeal lipid {delta}2H as a hydrological proxy, offering new insights into environments where traditional proxies, such as plant-derived lipids, are not available, including extreme environments and extraterrestrial settings. ImportanceReconstructing past climates is crucial for understanding Earths environmental history and its responses to changing conditions. This study examines Sulfolobus acidocaldarius, a thermoacidophilic archaeon that thrives in extreme environments like hot springs. These microorganisms incorporate hydrogen water in the growth environment into membrane lipids, creating hydrogen isotope signatures that can reflect hydroclimate conditions. Our findings show that these hydrogen isotope ratios remain consistent even under varying temperature, pH, oxygen levels, and electron donor fluxes, indicating a stable fractionation between lipids and water. This invariance suggests that S. acidocaldarius lipids could serve as a robust proxy for reconstructing ancient water H-isotope values, especially in extreme environments where traditional proxies, such as plant waxes, are absent. This research has broader implications for planetary-scale reconstructions, including potential applications in studying past climates on other planets, such as Mars, where similar microorganisms may have existed in hydrothermal conditions.

microbiology↗

bioRxiv2024

Temporal Dynamics of Neocortical Development in Organotypic Mouse Cultures: A Comprehensive Analysis

Murine organotypic brain slice cultures have been widely used in neuroscientific research and are offering the opportunity to study neuronal function under normal and disease conditions. Despite the brought application, the mechanisms governing the maturation of immature cortical circuits in vitro are not well understood. In this study, we present a detailed investigation into the development of the neocortex in vitro. Utilizing a holistic approach, we studied organotypic whole-hemisphere brain slice cultures from postnatal mice and tracked the development of the somatosensory area over a five-week period. Our analysis revealed the maturation of passive and active intrinsic properties of pyramidal cells together with their morphology, closely resembling in vivo development. Detailed Multi-electrode array (MEA) electrophysiological assessments and RNA expression profiling demonstrated stable network properties by two weeks in culture, followed by the transition of spontaneous activity towards more complex patterns including high-frequency oscillations. However, weeks 4 and 5 exhibited increased variability and initial signs of neuronal loss, highlighting the importance of considering developmental stages in experimental design. This comprehensive characterization is vital for understanding the temporal dynamics of the neocortical development in vitro, with implications for neuroscientific research methodologies, particularly in the investigation of diseases such as epilepsy and other neurodevelopmental disorders.

neuroscience↗

bioRxiv2024

Treatment of a genetic liver disease in mice through transient prime editor expression

Prime editing is a versatile genome editing technology that does not rely on DNA double-strand break formation and homology-directed repair (HDR). This makes it a promising tool for correcting pathogenic mutations in tissues consisting predominantly of postmitotic cells, such as the liver. While recent studies have already demonstrated proof-of-concept for in vivo prime editing, the use of viral delivery vectors resulted in prolonged prime editor (PE) expression, posing challenges for clinical application. Here, we developed an in vivo prime editing approach where we delivered the pegRNA using self-complementary adeno-associated viral (scAAV) vectors and the prime editor using nucleoside-modified mRNA encapsulated in lipid nanoparticles (LNPs). This methodology led to transient expression of the PE for 48h and 26% editing at the Dnmt1 locus using AAV doses of 2.5x1013 vector genomes (vg)/kg and a single dose of 3mg/kg mRNA-LNP. When targeting the pathogenic mutation in the Pahenu2 mouse model of phenylketonuria (PKU), we achieved 4.3% gene correction using an AAV dose of 2.5x1013 vg/kg and three doses of 2 mg/kg mRNA-LNP. Editing was specific to the liver and the intended locus, and was sufficient to reduce blood L-phenylalanine (Phe) levels from over 1500 {micro}mol/l to below the therapeutic threshold of 600 {micro}mol/l. Our study demonstrates the feasibility of in vivo gene correction in the liver with transient PE expression, bringing prime editing closer to clinical application.

bioengineering↗

bioRxiv2023

The hydrogen isotope signatures of autotrophy versus heterotrophy recorded in archaeal tetraether lipids

The stable hydrogen isotope composition of archaeal lipids is emerging as a potential paleoenvironmental proxy, adding to the well-established application of plant leaf wax-derived n-alkanes in paleohydrological reconstruction. A handful of studies reported relatively invariant and depleted hydrogen isotope compositions for archaeal lipids despite the range of different organisms and growth conditions explored. However, how modes of metabolism and physiological state (growth phase) affect the hydrogen isotope signatures of archaeal lipids remains poorly understood, limiting our ability to interpret archaeal lipid biomarker records from the environment. Here we conducted water isotope label experiments with a metabolically flexible and well-studied model archaeon Archaeoglobus fulgidus and quantified the hydrogen isotope fractionation between lipids and water in response to different carbon substrates and electron donor-acceptor pairs. The 2H/1H fractionation between lipids and water ({varepsilon}L/W) was overall negative, ranging from -280 to -226 {per thousand}, and overlapped with the ranges observed for other archaea in previous studies. Isotope flux-balance model results suggest that [≥]80 % and [≥]50 % of lipid-bound H in A. fulgidus cultures directly reflect water isotope compositions (i.e., not via organic substrate or H2) during autotrophy and heterotrophy, respectively. The model results also suggest the final saturation during isoprenoid lipid biosynthesis catalyzed by a flavoenzyme geranylgeranyl reductase likely contributes to the large 2H/1H fractionation observed in this study, consistent with previous isotope flux-balance model results for a different archaeon. Finally, we synthesized available data to compare {varepsilon}L/W patterns across all three domains of life: Bacteria, Archaea and Eukarya. Emerging patterns between archaeal and eukaryotic lipids are consistent with the notion of highly fractionating geranylgeranyl reductase, and the patterns between archaeal and bacterial lipids suggest that the general state of energy limitation may also contribute to large, negative values of {varepsilon}L/W observed in prokaryotic lipids. Altogether, these findings lend further support for the potential of archaeal lipid {varepsilon}L/W as a paleohydrological proxy and provide a broader insight into the 2H/1H fractionation mechanisms potentially shared among prokaryotic and eukaryotic lipid biomarkers.

microbiology↗

bioRxiv2023

Ultrahigh throughput evolution of tryptophan synthase in droplets via an aptamer-biosensor

Tryptophan synthase catalyzes the synthesis of a wide array of non-canonical amino acids and is an attractive target for directed evolution. Droplet microfluidics offers an ultrahigh throughput approach to directed evolution (>107 experiments per day), enabling the search for biocatalysts in wider regions of sequence space with reagent consumption minimized to the picoliter volume (per library member). While the majority of screening campaigns in this format on record relied on an optically active reaction product, a new assay is needed for tryptophan synthase. Tryptophan is not fluorogenic in the visible light spectrum and thus falls outside the scope of conventional droplet microfluidic read-outs which are incompatible with UV light detection at high throughput. Here, we engineer a tryptophan DNA aptamer into a biosensor to quantitatively report on tryptophan production in droplets. The utility of the biosensor was validated by identifying 5-fold improved tryptophan synthases from [~]100,000 protein variants. More generally this work establishes the use of DNA-aptamer sensors with a fluorogenic read-out in widening the scope of droplet microfluidic evolution.

biochemistry↗

bioRxiv2023

Mode of carbon and energy metabolism shifts lipid composition in the thermoacidophile Acidianus

The degree of cyclization, or ring index (RI), in archaeal glycerol dibiphytanyl glycerol tetraether (GDGT) lipids was long thought to reflect homeoviscous adaptation to temperature. However, more recent experiments show that other factors (e.g., pH, growth phase, and energy flux) can also affect membrane composition. The main objective of this study was to investigate the effect of carbon and energy metabolism on membrane cyclization. To do so we cultivated Acidianus sp. DS80, a metabolically flexible and thermoacidophilic archaeon, on different electron donor, acceptor and carbon source combinations (S0/Fe3+/CO2, H2/Fe3+/CO2, H2/S0/CO2, or H2/S0/glucose). We show that differences in energy and carbon metabolism can result in over a full unit of change in RI in the thermoacidophile Acidianus sp. DS80. The patterns in RI correlated with the normalized electron transfer rate between electron donor and acceptor and did not always align with thermodynamic predictions of energy yield. In light of this, we discuss other factors that may affect the kinetics of cellular energy metabolism: electron transfer chain (ETC) efficiency, location of ETC reaction components (cytoplasmic vs. extracellular), and the physical state of electron donors and acceptors (gas vs. solid). Furthermore, assimilation of a more reduced form of carbon during heterotrophy appears to decrease the demand for reducing equivalents during lipid biosynthesis, resulting in lower RI. Together, these results point to the fundamental role of the cellular energy state in dictating GDGT cyclization, with those cells experiencing greater energy limitation synthesizing more cyclized GDGTs. ImportanceSome archaea make unique membrane-spanning lipids with different numbers of five or six membered rings in the core structure that modulate membrane fluidity and permeability. Changes in membrane core lipid composition reflect fundamental adaptation strategies of archaea in response to stress, but multiple environmental and physiological factors may affect the needs for membrane fluidity and permeability. In this study, we tested how Acidianus sp. DS80 changed its core lipid composition when grown with different electron donor/acceptor pairs. We show that changes in energy and carbon metabolisms significantly affected the relative abundance of rings in the core lipids of DS80. These observations highlight the need to better constrain metabolic parameters, in addition to environmental factors, that may influence changes in membrane physiology in Archaea. Such consideration would be particularly important for studying archaeal lipids from habitats that experience frequent environmental fluctuations and/or where metabolically diverse archaea thrive.

microbiology↗

bioRxiv2023

Prime editing of the beta-1 adrenoceptor in the brain reprograms mouse behavior

Prime editing is a highly versatile genome editing technology that holds great potential for treating genetic diseases1, 2. While in vivo prime editing has recently been conducted in the brain, liver, heart, and retina3-6, application of this technology to modulate neural circuits in the brain has not been reported yet. Here, we employ adeno-associated viral vectors to deliver optimized intein-split prime editors into the brain of mice. Delivery into newborn pups via intracerebroventricular injection resulted in up to 44.0% editing at the Dnmt1 locus in the cortex (on average 34.8{+/-}9.8% after 6 months). In addition, we obtained up to 28.1% editing at the Adrb1 locus in the cortex (on average 14.7{+/-}11.6% after 6 months). The introduced Adrb1A187V mutation is a naturally occurring variant of the {beta}1-adrenergic receptor, which has previously been linked to increased activity and natural short sleep7. Similarly, we observed an increase in the activity and exploratory behavior of treated animals. This study demonstrates the potential of prime editing for treating genetic diseases in the central nervous system and for reprogramming molecular pathways that modulate animal behavior.

genetics↗

bioRxiv2022

Enhancing the performance of Magnets photosensors through directed evolution

Photosensory protein domains are the basis of optogenetic protein engineering. These domains originate from natural sources where they fulfill specific functions ranging from the protection against photooxidative damage to circadian rhythms. When used in synthetic biology, the features of these photosensory domains can be specifically tailored towards the application of interest, enabling their full exploitation for optogenetic regulation in basic research and applied bioengineering. In this work, we develop and apply a simple, yet powerful, directed evolution and high-throughput screening strategy that allows us to alter the most fundamental property of the widely used nMag/pMag photodimerization system: its light sensitivity. We identify a set of mutations located within the photosensory domains, which either increase or decrease the light sensitivity at sub-saturating light intensities, while also improving the dark-to-light fold change in certain variants. For some of these variants, photosensitivity and expression levels could be changed independently, showing that the shape of the light-activity dose-response curve can be tuned and adjusted. We functionally characterize the variants in vivo in bacteria on the single-cell and the population levels. We further show that a subset of these variants can be transferred into the mOptoT7 for gene expression regulation in mammalian cells. We demonstrate increased gene expression levels for low light intensities, resulting in reduced potential phototoxicity in long-term experiments. Our findings expand the applicability of the widely used Magnets photosensors by enabling a tuning towards the needs of specific optogenetic regulation strategies. More generally, our approach will aid optogenetic approaches by making the adaptation of photosensor properties possible to better suit specific experimental or bioprocess needs.

synthetic biology↗

bioRxiv2021

Water column stability as an important factor controlling nitrite-dependent anaerobic methane oxidation in stratified lake basins

Anaerobic oxidation of methane (AOM) with nitrate/nitrite as the terminal electron acceptor may play an important role in mitigating methane emissions from lacustrine environments to the atmosphere. We investigated AOM in the water column of two connected but hydrodynamically contrasting basins of a south-alpine lake in Switzerland (Lake Lugano). The North Basin is permanently stratified with year-round anoxic conditions below 120 m water depth, while the South Basin undergoes seasonal stratification with the development of bottom water anoxia during summer. We show that below the redoxcline of the North Basin a substantial fraction of methane was oxidized coupled to nitrite reduction by Candidatus Methylomirabilis. Incubation experiments with 14CH4 and concentrated biomass from showed at least 43-52%-enhanced AOM rates with added nitrate/nitrite as electron acceptor. Multiannual time series data on the population dynamics of Candidatus Methylomirabilis in the North Basin following an exceptional mixing event in 2005/2006 revealed their requirement for lasting stable low redox-conditions to establish. In the South Basin, on the other hand, we did not find molecular evidence for nitrite-dependent methane oxidizing bacteria. Our data suggest that here the dynamic mixing regime with fluctuating redox conditions is not conducive to the development of a stable population of relatively slow-growing Candidatus Methylomirabilis, despite a hydrochemical framework that seems more favorable for nitrite-dependent AOM than in the North Basin. We predict that the importance of N-dependent AOM in freshwater lakes will likely increase in future because of longer thermal stratification periods and reduced mixing caused by global warming.

microbiology↗

bioRxiv2020

Multiple environmental parameters impact core lipid cyclization in Sulfolobus acidocaldarius

Environmental reconstructions based on microbial lipids require understanding the coupling between environmental conditions and membrane physiology. The paleotemperature proxy TEX86 is built on the observation that archaea alter the number of five- and six-membered rings in the hydrophobic core of their glycerol dibiphytanyl glycerol tetraether (GDGT) membrane lipids when growing at different temperatures. However, recent work with these archaea also highlights a role for other factors, such as pH or energy availability in determining the degree of core lipid cyclization. To better understand the role of these variables we cultivated a model Crenarchaeon, Sulfolobus acidocaldarius, over a range in temperature, pH, oxygen flux, or agitation speed, and quantified the changes in growth rate, biomass yield, and core lipid compositions. The average degree of cyclization in core lipids correlated with growth rate under most conditions. When considered alongside other experimental findings from both the thermoacidophilic and mesoneutrophilic archaea, the results suggest the cyclization of archaeal lipids records a universal response to energy availability at the cellular level. Although we isolated the effects of individual parameters, there remains a need for multi-factor experiments (e.g., pH + temperature + redox) to establish a robust framework to interpret biomarker records of environmental change.

microbiology↗

bioRxiv2019

Energy flux controls tetraether lipid cyclization in Sulfolobus acidocaldarius

Microorganisms regulate the composition of their membranes in response to environmental cues. Many archaea maintain the fluidity and permeability of their membranes by adjusting the number of cyclic moieties within the cores of their glycerol dibiphytanyl glycerol tetraether (GDGT) lipids. Cyclized GDGTs increase membrane packing and stability, which has been shown to help cells survive shifts in temperature and pH. However, the extent of this cyclization also varies with growth phase and electron acceptor or donor limitation. These observations indicate a relationship between energy metabolism and membrane composition. Here we show that the average degree of GDGT cyclization increases with doubling time in continuous cultures of the thermoacidophile Sulfolobus acidocaldarius (DSM 639). This is consistent with the behavior of a mesoneutrophile, Nitrosopumilus maritimus SCM1. Together, these results demonstrate that archaeal GDGT distributions can shift in response to electron donor flux and energy availability, independent of pH or temperature. Paleoenvironmental reconstructions based on GDGTs thus capture the energy available to microbes, which encompasses fluctuations in temperature and pH, as well as electron donor and acceptor availability. The ability of Archaea to adjust membrane composition and packing may be an important strategy that enables survival during episodes of energy stress.\n\nSignificance StatementMicrobial lipid membranes protect and isolate a cell from its environment while regulating the flow of energy and nutrients to metabolic reaction centers within. We demonstrate that membrane lipids change as a function of energy flux using a well-studied archaeon that thrives in acidic hot springs and observe an increase in membrane packing as energy becomes more limited. These observations are consistent with chemostat experiments utilizing a low temperature, neutral pH, marine archaeon. This strategy appears to regulate membrane homeostasis is common across GDGT-producing lineages, demonstrating that diverse taxa adjust membrane composition in response to chronic energy stress.

microbiology↗

bioRxiv2019

Targeted gene sequencing in 6994 individuals with neurodevelopmental disorder with epilepsy

PurposeWe aimed to gain insight into frequencies of genetic variants in genes implicated in neurodevelopmental disorder with epilepsy (NDD+E) by investigating large cohorts of patients in a diagnostic setting.\n\nMethodsWe analyzed variants in NDD+E using epilepsy gene panel sequencing performed between 2013 and 2017 by two large diagnostic companies. We compared variant frequencies in 6,994 panels to other 8,588 recently published panels as well as exome-wide de novo variants in 1,942 individuals with NDD+E and 10,937 controls.\n\nResultsGenes with highest frequencies of ultra-rare variants in NDD+E comprised SCN1A, KCNQ2, SCN2A, CDKL5, SCN8A and STXBP1, concordant with the two other epilepsy cohorts we investigated. Only 46% of the analysed 262 dominant and X-linked panel genes contained ultra-rare variants in patients. Among genes with contradictory evidence of association with epilepsy CACNB4, CLCN2, EFHC1, GABRD, MAGI2 and SRPX2 showed equal frequencies in cases and controls.\n\nConclusionWe show that improvement of panel design increased diagnostic yield over time, but panels still display genes with low or no diagnostic yield. With our data, we hope to improve current diagnostic NDD+E panel design and provide a resource of ultra-rare variants in individuals with NDD+E to the community.

genetics↗

bioRxiv2017

Characterization of Glycosylphosphatidylinositol Biosynthesis Defects by Clinical Features, Flow Cytometry, and Automated Image Analysis

BackgroundGlycosylphosphatidylinositol Biosynthesis Defects (GPIBDs) cause a group of phenotypically overlapping recessive syndromes with intellectual disability, for which pathogenic mutations have been described in 16 genes of the corresponding molecular pathway. An elevated serum activity of alkaline phosphatase (AP), a GPI-linked enzyme, has been used to assign GPIBDs to the phenotypic series of Hyperphosphatasia with Mental Retardation Syndrome (HPMRS) and to distinguish them from another subset of GPIBDs, termed Multiple Congenital Anomalies Hypotonia Seizures syndrome (MCAHS). However, the increasing number of individuals with a GPIBD shows that hyperphosphatasia is a variable feature that is not ideal for a clinical classification.\n\nMethodsWe studied the discriminatory power of multiple GPI-linked substrates that were assessed by flow cytometry in blood cells and fibroblasts of 39 and 14 individuals with a GPIBD, respectively. On the phenotypic level, we evaluated the frequency of occurrence of clinical symptoms and analyzed the performance of computer-assisted image analysis of the facial gestalt in 91 individuals.\n\nResultsWe found that certain malformations such as Morbus Hirschsprung and Diaphragmatic defects are more likely to be associated with particular gene defects (PIGV, PGAP3, PIGN). However, especially at the severe end of the clinical spectrum of HPMRS, there is a high phenotypic overlap with MCAHS. Elevation of AP has also been documented in some of the individuals with MCAHS, namely those with PIGA mutations. Although the impairment of GPI-linked substrates is supposed to play the key role in the pathophysiology of GPIBDs, we could not observe gene-specific profiles for flow cytometric markers or a correlation between their cell surface levels and the severity of the phenotype. In contrast, it was facial recognition software that achieved the highest accuracy in predicting the disease-causing gene in a GPIBD.\n\nConclusionsDue to the overlapping clinical spectrum of both, HPMRS and MCAHS, in the majority of affected individuals, the elevation of AP and the reduced surface levels of GPI-linked markers in both groups, a common classification as GPIBDs is recommended. The effectiveness of computer-assisted gestalt analysis for the correct gene inference in a GPIBD and probably beyond is remarkable and illustrates how the information contained in human faces is pivotal in the delineation of genetic entities.

genetics↗

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↗