scholarly journals Classifying disease-associated variants using measures of protein activity and stability

2019 ◽  
Author(s):  
Michael Maglegaard Jepsen ◽  
Douglas M. Fowler ◽  
Rasmus Hartmann-Petersen ◽  
Amelie Stein ◽  
Kresten Lindorff-Larsen
Keyword(s):  
Protein Stability ◽  
Computational Methods ◽  
Genetic Disorders ◽  
Computational Modelling ◽  
Missense Variant ◽  
Loss Of Function ◽  
Protein Activity ◽  
Missense Variants ◽  
Tensin Homolog ◽  
Pathogenic Variants

AbstractDecreased cost of human exome and genome sequencing provides new opportunities for diagnosing genetic disorders, but we need better and more robust methods for interpreting sequencing results including determining whether and by which mechanism a specific missense variants may be pathogenic. Using the protein PTEN (phosphatase and tensin homolog) as an example, we show how recent developments in both experiments and computational modelling can be used to determine whether a missense variant is likely to be pathogenic. One approach relies on multiplexed experiments that enable determination of the effect of all possible individual missense variants in a cellular assay. Another approach is to use computational methods to predict variant effects. We compare two different multiplexed experiments and two computational methods to classify variant effects in PTEN. We distinguish between methods that focus on effects on protein stability and protein-specific methods that are more directly related to enzyme activity. Our results on PTEN suggest that ~60% of pathogenic variants cause loss of function because they destabilise the folded protein which is subsequently degraded. Methods that quantify a broader range of effects on PTEN activity perform better at predicting variant effects. Either experimental method performs better than the corresponding computational predictions, so that e.g. experiments that probe cellular abundance perform better at identifying pathogenic variants than predictions of thermodynamic stability. Our results suggest that loss of stability of PTEN is a key driver for disease, and we hypothesize that experiments and prediction methods that probe protein stability can be used to find variants with similar mechanisms in other genes.

scholarly journals Protein Stability Perturbation Contributes to the Loss of Function in Haploinsufficient Genes

2021 ◽  
Vol 8 ◽  
Author(s):  
Giovanni Birolo ◽  
Silvia Benevenuta ◽  
Piero Fariselli ◽  
Emidio Capriotti ◽  
Elisa Giorgio ◽  
...  
Keyword(s):  
Protein Stability ◽  
Missense Variant ◽  
Loss Of Function ◽  
Disease Biomarkers ◽  
Missense Variants ◽  
Pathogenic Variants ◽  
Predictive Correlation ◽  
Genome Modifications

Missense variants are among the most studied genome modifications as disease biomarkers. It has been shown that the “perturbation” of the protein stability upon a missense variant (in terms of absolute ΔΔG value, i.e., |ΔΔG|) has a significant, but not predictive, correlation with the pathogenicity of that variant. However, here we show that this correlation becomes significantly amplified in haploinsufficient genes. Moreover, the enrichment of pathogenic variants increases at the increasing protein stability perturbation value. These findings suggest that protein stability perturbation might be considered as a potential cofactor in diseases associated with haploinsufficient genes reporting missense variants.

scholarly journals Loss-of-function and missense variants in NSD2 cause decreased methylation activity and are associated with a distinct developmental phenotype

2021 ◽  
Author(s):  
Paolo Zanoni ◽  
Katharina Steindl ◽  
Deepanwita Sengupta ◽  
Pascal Joset ◽  
Angela Bahr ◽  
...  
Keyword(s):  
Loss Of Function ◽  
Mild Phenotype ◽  
Psychological Issues ◽  
Missense Variants ◽  
Mild Developmental Delay ◽  
Pathogenic Variants ◽  
In Silico Modeling ◽  
And Function ◽  
Methylation Activity

Abstract Purpose Despite a few recent reports of patients harboring truncating variants in NSD2, a gene considered critical for the Wolf–Hirschhorn syndrome (WHS) phenotype, the clinical spectrum associated with NSD2 pathogenic variants remains poorly understood. Methods We collected a comprehensive series of 18 unpublished patients carrying heterozygous missense, elongating, or truncating NSD2 variants; compared their clinical data to the typical WHS phenotype after pooling them with ten previously described patients; and assessed the underlying molecular mechanism by structural modeling and measuring methylation activity in vitro. Results The core NSD2-associated phenotype includes mostly mild developmental delay, prenatal-onset growth retardation, low body mass index, and characteristic facial features distinct from WHS. Patients carrying missense variants were significantly taller and had more frequent behavioral/psychological issues compared with those harboring truncating variants. Structural in silico modeling suggested interference with NSD2’s folding and function for all missense variants in known structures. In vitro testing showed reduced methylation activity and failure to reconstitute H3K36me2 in NSD2 knockout cells for most missense variants. Conclusion NSD2 loss-of-function variants lead to a distinct, rather mild phenotype partially overlapping with WHS. To avoid confusion for patients, NSD2 deficiency may be named Rauch–Steindl syndrome after the delineators of this phenotype.

scholarly journals A novel de novo DDX3X missense variant in a female with brachycephaly and intellectual disability: a case report

2021 ◽  
Vol 47 (1) ◽  
Author(s):  
Giada Moresco ◽  
Jole Costanza ◽  
Carlo Santaniello ◽  
Ornella Rondinone ◽  
Federico Grilli ◽  
...  
Keyword(s):  
Intellectual Disability ◽  
Clinical Presentation ◽  
De Novo ◽  
Clinical Signs ◽  
Missense Variant ◽  
Psychomotor Development ◽  
Helicase Domain ◽  
Protein Activity ◽  
Mrna Metabolism ◽  
Pathogenic Variants

Abstract Background De novo pathogenic variants in the DDX3X gene are reported to account for 1–3% of unexplained intellectual disability (ID) in females, leading to the rare disease known as DDX3X syndrome (MRXSSB, OMIM #300958). Besides ID, these patients manifest a variable clinical presentation, which includes neurological and behavioral defects, and abnormal brain MRIs. Case presentation We report a 10-year-old girl affected by delayed psychomotor development, delayed myelination, and polymicrogyria (PMG). We identified a novel de novo missense mutation in the DDX3X gene (c.625C > G) by whole exome sequencing (WES). The DDX3X gene encodes a DEAD-box ATP-dependent RNA-helicase broadly implicated in gene expression through regulation of mRNA metabolism. The identified mutation is located just upstream the helicase domain and is suggested to impair the protein activity, thus resulting in the altered translation of DDX3X-dependent mRNAs. The proband, presenting with the typical PMG phenotype related to the syndrome, does not show other clinical signs frequently reported in presence of missense DDX3X mutations that are associated with a most severe clinical presentation. In addition, she has brachycephaly, never described in female DDX3X patients, and macroglossia, that has never been associated with the syndrome. Conclusions This case expands the knowledge of DDX3X pathogenic variants and the associated DDX3X syndrome phenotypic spectrum.

BDV-syndrome: An emerging syndrome with profound obesity and neurodevelopmental delay resembling Prader-Willi syndrome

2021 ◽  
Author(s):  
Elisabeth Bosch ◽  
Moritz Hebebrand ◽  
Bernt Popp ◽  
Theresa Penger ◽  
Bettina Behring ◽  
...  
Keyword(s):  
Clinical Features ◽  
Large Scale ◽  
Hypogonadotropic Hypogonadism ◽  
Computational Modelling ◽  
Prader Willi Syndrome ◽  
Loss Of Function ◽  
Human Phenotype ◽  
Neurodevelopmental Delay ◽  
Missense Variants ◽  
Severely Obese

Abstract Context CPE encodes carboxypeptidase E, an enzyme which converts proneuropeptides and propeptide hormones to bioactive forms. It is widely expressed in the endocrine and central nervous system. To date, four individuals from two families with core clinical features including morbid obesity, neurodevelopmental delay and hypogonadotropic hypogonadism, harbouring biallelic loss-of-function CPE variants, were reported. Objective We describe four affected individuals from three unrelated consanguineous families, two siblings of Syrian, one of Egyptian and one of Pakistani descent, all harbouring novel homozygous CPE loss-of-function variants. Methods After excluding Prader-Willi syndrome, exome sequencing was performed in both Syrian siblings. The variants identified in the other two individuals were reported as research variants in a large scale exome study and in ClinVar database. Computational modelling of all possible missense alterations allowed assessing CPE tolerance to missense variants. Results All affected individuals were severely obese with neurodevelopmental delay and other endocrine anomalies. Three individuals from two families shared the same CPE homozygous truncating variant c.361C>T, p.(Arg121*), while the fourth carried the c.994del, p.(Ser333Alafs*22) variant. Comparison of clinical features with previously described cases and standardization according to the Human Phenotype Ontology indicated a recognisable clinical phenotype, which we termed Blakemore-Durmaz-Vasileiou (BDV) syndrome. Computational analysis indicated high conservation of CPE domains and intolerance to missense changes. Conclusions Biallelic truncating CPE variants are associated with BDV syndrome, a clinically recognisable monogenic recessive syndrome with childhood-onset obesity, neurodevelopmental delay, hypogonadotropic hypogonadism and hypothyroidism. BDV syndrome resembles Prader-Willi syndrome. Our findings suggested that missense variants may also be clinically relevant.

scholarly journals A combined in silico, in vitro and clinical approach to characterise novel pathogenic missense variants in PRPF31 in retinitis pigmentosa

2018 ◽  
Author(s):  
Gabrielle Wheway ◽  
Liliya Nazlamova ◽  
Nervine Meshad ◽  
Samantha Hunt ◽  
Nicola Jackson ◽  
...  
Keyword(s):  
Retinitis Pigmentosa ◽  
In Silico ◽  
Autosomal Dominant ◽  
Missense Variant ◽  
Incomplete Penetrance ◽  
Clinical Approach ◽  
Loss Of Function ◽  
Autosomal Dominant Retinitis Pigmentosa ◽  
Missense Variants

AbstractAt least six different proteins of the spliceosome, including PRPF3, PRPF4, PRPF6, PRPF8, PRPF31 and SNRNP200, are mutated in autosomal dominant retinitis pigmentosa (adRP). These proteins have recently been shown to localise to the base of the connecting cilium of the retinal photoreceptor cells, elucidating this form of RP as a retinal ciliopathy. In the case of loss-of-function variants in these genes, pathogenicity can easily be ascribed. In the case of missense variants, this is more challenging. Furthermore, the exact molecular mechanism of disease in this form of RP remains poorly understood.In this paper we take advantage of the recently published cryo EM-resolved structure of the entire human spliceosome, to predict the effect of a novel missense variant in one component of the spliceosome; PRPF31, found in a patient attending the genetics eye clinic at Bristol Eye Hospital. Monoallelic variants in PRPF31 are a common cause of autosomal dominant retinitis pigmentosa (adRP) with incomplete penetrance. We use in vitro studies to confirm pathogenicity of this novel variant PRPF31 c.341T>A, p.Ile114Asn.This work demonstrates how in silico modelling of structural effects of missense variants on cryo-EM resolved protein complexes can contribute to predicting pathogenicity of novel variants, in combination with in vitro and clinical studies. It is currently a considerable challenge to assign pathogenic status to missense variants in these proteins.

scholarly journals A large deletion in the COL2A1 gene expands the spectrum of pathogenic variants causing bulldog calf syndrome in cattle

2020 ◽  
Vol 62 (1) ◽  
Author(s):  
Joana Gonçalves Pontes Jacinto ◽  
Irene Monika Häfliger ◽  
Anna Letko ◽  
Cord Drögemüller ◽  
Jørgen Steen Agerholm
Keyword(s):  
De Novo ◽  
Genetic Disorders ◽  
Polymerase Chain Reaction Analysis ◽  
Large Deletion ◽  
Whole Genome Sequence ◽  
De Novo Mutation ◽  
Loss Of Function ◽  
Single Nucleotide Variants ◽  
Pathogenic Variants ◽  
Base Pair Deletion

Abstract Background Congenital bovine chondrodysplasia, also known as bulldog calf syndrome, is characterized by disproportionate growth of bones resulting in a shortened and compressed body, mainly due to reduced length of the spine and the long bones of the limbs. In addition, severe facial dysmorphisms including palatoschisis and shortening of the viscerocranium are present. Abnormalities in the gene collagen type II alpha 1 chain (COL2A1) have been associated with some cases of the bulldog calf syndrome. Until now, six pathogenic single-nucleotide variants have been found in COL2A1. Here we present a novel variant in COL2A1 of a Holstein calf and provide an overview of the phenotypic and allelic heterogeneity of the COL2A1-related bulldog calf syndrome in cattle. Case presentation The calf was aborted at gestation day 264 and showed generalized disproportionate dwarfism, with a shortened compressed body and limbs, and dysplasia of the viscerocranium; a phenotype resembling bulldog calf syndrome due to an abnormality in COL2A1. Whole-genome sequence (WGS) data was obtained and revealed a heterozygous 3513 base pair deletion encompassing 10 of the 54 coding exons of COL2A1. Polymerase chain reaction analysis and Sanger sequencing confirmed the breakpoints of the deletion and its absence in the genomes of both parents. Conclusions The pathological and genetic findings were consistent with a case of “bulldog calf syndrome”. The identified variant causing the syndrome was the result of a de novo mutation event that either occurred post-zygotically in the developing embryo or was inherited because of low-level mosaicism in one of the parents. The identified loss-of-function variant is pathogenic due to COL2A1 haploinsufficiency and represents the first structural variant causing bulldog calf syndrome in cattle. Furthermore, this case report highlights the utility of WGS-based precise diagnostics for understanding congenital disorders in cattle and the need for continued surveillance for genetic disorders in cattle.

scholarly journals Systematic analysis of PINK1 variants of unknown significance shows intact mitophagy function for most variants

2021 ◽  
Vol 7 (1) ◽  
Author(s):  
Kai Yu Ma ◽  
Michiel R. Fokkens ◽  
Teus van Laar ◽  
Dineke S. Verbeek
Keyword(s):  
Parkinson’S Disease ◽  
Parkinson's Disease ◽  
Missense Variant ◽  
Population Based ◽  
Loss Of Function ◽  
Systematic Analysis ◽  
Missense Variants ◽  
Variants Of Unknown Significance ◽  
Disease Case ◽  
Unknown Significance

AbstractPathogenic variants in PINK1 cause early-onset Parkinson’s disease. Although many PINK1 variants have been reported, the clinical significance is uncertain for the majority of them. To gain insights into the consequences of PINK1 missense variants in a systematic manner, we selected 50 PINK1 missense variants from patient- and population-wide databases and systematically classified them using Sherloc, a comprehensive framework for variant interpretation based on ACMG-AMP guidelines. We then performed functional experiments, including mitophagy and Parkin recruitment assays, to assess the downstream consequences of PINK1 variants. Analysis of PINK1 missense variants based on Sherloc showed that the patient databases over-annotate variants as likely pathogenic. Furthermore, our study shows that pathogenic PINK1 variants are most often linked to a loss-of-function for mitophagy and Parkin recruitment, while this is not observed for variants of unknown significance. In addition to the Sherloc framework, the added layer of evidence of our functional tests suggests a reclassification of 9/50 missense variants. In conclusion, we suggest the assessment of multiple layers of evidence, including functional data on top of available clinical and population-based data, to support the clinical classification of a variant and show that the presence of a missense variant in PINK1 in a Parkinson’s disease case does not automatically imply pathogenicity.

scholarly journals De novo FZR1 loss-of-function variants cause developmental and epileptic encephalopathies including Myoclonic Atonic Epilepsy

2021 ◽  
Author(s):  
Sathiya N. Manivannan ◽  
Jolien Roovers ◽  
Noor Smal ◽  
Candace T. Myers ◽  
Dilsad Turkdogan ◽  
...  
Keyword(s):  
De Novo ◽  
Statistical Tests ◽  
Essential Feature ◽  
Missense Variant ◽  
Anaphase Promoting Complex ◽  
Loss Of Function ◽  
Childhood Onset ◽  
Epileptic Encephalopathies ◽  
Missense Variants ◽  
Generalized Epilepsy

FZR1, which encodes the Cdh1 subunit of the Anaphase Promoting Complex, plays an important role in neurodevelopment, both through the control of the cell cycle and through its multiple functions in post-mitotic neurons. In this study, the evaluation of 250 unrelated patients with developmental epileptic encephalopathies (DEE) and a connection on GeneMatcher led to the identification of three de novo missense variants in FZR1. Two variants led to the same amino acid change. All individuals had a DEE with childhood-onset generalized epilepsy, intellectual disability, mild ataxia, and normal head circumference. Two individuals were diagnosed with the DEE subtype Myoclonic Atonic Epilepsy (MAE). We provide gene burden testing using two independent statistical tests to support FZR1 association with DEE. Further, we provide functional evidence that the missense variants are loss-of-function (LOF) alleles using Drosophila neurodevelopment assays. Using three fly mutant alleles of the Drosophila homolog fzr and overexpression studies, we show that patient variants do not support proper neurodevelopment. Along with a recent report of a patient with neonatal-onset DEE with microcephaly who also carries a de novo FZR1 missense variant, our study consolidates the relationship between FZR1 and DEE, and expands the associated phenotype. We conclude that heterozygous LOF of FZR1 leads to DEE associated with a spectrum of neonatal to childhood-onset seizure types, developmental delay, and mild ataxia. Microcephaly can be present but is not an essential feature of FZR1-encephalopathy. In summary, our approach of targeted sequencing using novel gene candidates and functional testing in Drosophila will help solve undiagnosed MAE/DEE cases.

scholarly journals Critical Role of E1623 Residue in S3-S4 Loop of Nav1.1 Channel and Correlation Between Nature of Substitution and Functional Alteration

2022 ◽  
Vol 14 ◽  
Author(s):  
Tao Su ◽  
Meng-Long Chen ◽  
Li-Hong Liu ◽  
Hen Meng ◽  
Bin Tang ◽  
...  
Keyword(s):  
Current Density ◽  
Genetic Disorders ◽  
Critical Role ◽  
Missense Variant ◽  
Loss Of Function ◽  
Channel Conductance ◽  
Peak Current Density ◽  
Functional Changes ◽  
Functional Alteration

Objective: An overwhelming majority of the genetic variants associated with genetic disorders are missense. The association between the nature of substitution and the functional alteration, which is critical in determining the pathogenicity of variants, remains largely unknown. With a novel missense variant (E1623A) identified from two epileptic cases, which occurs in the extracellular S3-S4 loop of Nav1.1, we studied functional changes of all latent mutations at residue E1623, aiming to understand the relationship between substitution nature and functional alteration.Methods: Six latent mutants with amino acid substitutions at E1623 were generated, followed by measurements of their electrophysiological alterations. Different computational analyses were used to parameterize the residue alterations.Results: Structural modeling indicated that the E1623 was located in the peripheral region far from the central pore, and contributed to the tight turn of the S3-S4 loop. The E1623 residue exhibited low functional tolerance to the substitutions with the most remarkable loss-of-function found in E1623A, including reduced current density, less steady-state availability of activation and inactivation, and slower recovery from fast inactivation. Correlation analysis between electrophysiological parameters and the parameterized physicochemical properties of different residues suggested that hydrophilicity of side-chain at E1623 might be a crucial contributor for voltage-dependent kinetics. However, none of the established algorithms on the physicochemical variations of residues could well predict changes in the channel conductance property indicated by peak current density.Significance: The results established the important role of the extracellular S3-S4 loop in Nav1.1 channel gating and proposed a possible effect of local conformational loop flexibility on channel conductance and kinetics. Site-specific knowledge of protein will be a fundamental task for future bioinformatics.

scholarly journals Novel variants identified in CKAP2L in two siblings with Filippi Syndrome

2021 ◽  
pp. mcs.a006130
Author(s):  
Ryan J Patrick ◽  
Jill M Weimer ◽  
Laura Davis-Keppen ◽  
Megan L Landsverk
Keyword(s):  
Intellectual Disability ◽  
Autosomal Recessive ◽  
Missense Variant ◽  
Facial Features ◽  
Loss Of Function ◽  
Pathogenic Variants ◽  
Whole Exome ◽  
The Family ◽  
Novel Variants ◽  
In Trans

Pathogenic variants in CKAP2L have previously been reported in Filippi Syndrome (FS), a rare autosomal recessive, craniodigital syndrome characterized by microcephaly, syndactyly, short stature, intellectual disability, and dysmorphic facial features. To date, fewer than ten patients with pathogenic variants in CKAP2L associated with FS have been reported. All of the previously reported probands have presumed loss-of-function variants (frameshift, canonical splice site, starting methionine) and all but one have been homozygous for a pathogenic variant. Here we describe two brothers who presented with microcephaly, micrognathia, syndactyly, dysmorphic features, and intellectual disability. Whole exome sequencing of the family identified a missense variant, c.2066G>A (p.Arg689His), in trans with a frameshift variant, c.1169_1173del (p.Ile390LysfsTer4), in CKAP2L. To our knowledge, these are the first patients with FS to be reported with a missense variant in CKAP2L and only the second family to be reported with two variants in trans.

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