CACNA1I gain-of-function mutations differentially affect channel gating and cause neurodevelopmental disorders

Abstract T-type calcium channels (Cav3.1 to Cav3.3) regulate low-threshold calcium spikes, burst firing and rhythmic oscillations of neurons and are involved in sensory processing, sleep, and hormone and neurotransmitter release. Here we examined four heterozygous missense variants in CACNA1I, encoding the Cav3.3 channel, in patients with variable neurodevelopmental phenotypes. The p.(Ile860Met) variant, affecting a residue in the putative channel gate at the cytoplasmic end of the IIS6 segment, was identified in three family members with variable cognitive impairment. The de novo p.(Ile860Asn) variant, changing the same amino acid residue, was detected in a patient with severe developmental delay and seizures. In two additional individuals with global developmental delay, hypotonia, and epilepsy the variants p.(Ile1306Thr) and p.(Met1425Ile), substituting residues at the cytoplasmic ends of IIIS5 and IIIS6, respectively, were found. Because structure modelling indicated that the amino acid substitutions differentially affect the mobility of the channel gate, we analyzed possible effects on CaV3.3 channel function using patch-clamp analysis in HEK293T cells. The mutations resulted in slowed kinetics of current activation, inactivation, and deactivation, and in hyperpolarizing shifts of the voltage-dependence of activation and inactivation, with CaV3.3-I860N showing the strongest and CaV3.3-I860M the weakest effect. Structure modelling suggests that by introducing stabilizing hydrogen bonds the mutations slow the kinetics of the channel gate and cause the gain-of-function effect in CaV3.3 channels. The gating defects left-shifted and increased the window currents, resulting in increased calcium influx during repetitive action potentials and even at resting membrane potentials. Thus, calcium toxicity in neurons expressing the CaV3.3 variants is one likely cause of the neurodevelopmental phenotype. Computer modelling of thalamic reticular nuclei neurons indicated that the altered gating properties of the CaV3.3 disease variants lower the threshold and increase the duration and frequency of action potential firing. Expressing the CaV3.3-I860N/M mutants in mouse chromaffin cells shifted the mode of firing from low-threshold spikes and rebound burst firing with wild-type CaV3.3 to slow oscillations with CaV3.3-I860N and an intermediate firing mode with CaV3.3-I860M, respectively. Such neuronal hyper-excitability could explain seizures in the patient with the p.(Ile860Asn) mutation. Thus, our study implicates CACNA1I gain-of-function mutations in neurodevelopmental disorders, with a phenotypic spectrum ranging from borderline intellectual functioning to a severe neurodevelopmental disorder with epilepsy.

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Clinical Characteristics and Genotype–Phenotype Correlation in Children with KMT2E Gene-Related Neurodevelopmental Disorders: Report of Two New Cases and Review of Published Literature

Neuropediatrics ◽

10.1055/s-0040-1715629 ◽

2020 ◽

Author(s):

Indar Kumar Sharawat ◽

Prateek Kumar Panda ◽

Lesa Dawman

Keyword(s):

Developmental Delay ◽

Neurodevelopmental Disorder ◽

Copy Number Variants ◽

Cerebral White Matter ◽

Global Developmental Delay ◽

Missense Mutations ◽

Search Terms ◽

Pathogenic Factors ◽

The Common ◽

Variable Combination

Abstract Background In recent years, many new candidate genes are being identified as putative pathogenic factors in children with developmental delay and autism. Recently, heterozygous mutations in the KMT2E gene have been identified as a cause of a unique neurodevelopmental disorder with variable combination of global developmental delay or isolated speech delay, intellectual disability, autistic features, and seizures. Methods Here, we present two new cases of KMT2E mutation-associated neurodevelopmental disorder in a 4-year-old girl and 5-year-old boy. We also performed a pooled review of the previously published cases of KMT2E-related neurodevelopmental disorder. Articles were identified through search engines using appropriate search terms. Results Along with the presented 2 cases, 40 cases were analyzed. Out of them, 30, 6, and 4 children had protein-truncating mutations, missense mutations, and copy number variants, respectively. The common features were global developmental delay (97%) followed by macrocephaly (35%), seizures (30%), and autism (25%). Children with missense variants had severe phenotype, with microcephaly, profound developmental delay, and increased frequency of seizures. Neuroimaging revealed nonspecific changes, including cerebral white matter signal abnormalities. Conclusion KMT2E-related neurodevelopmental disorder remains one of the clinical differentials in children with global developmental delay and/or autistic features/seizure. With the reporting of more cases in the future, the already heterogeneous clinical spectrum of this disease is likely to be widened.

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X-linked neonatal-onset epileptic encephalopathy associated with a gain-of-function variant p.R660T in GRIA3

PLoS Genetics ◽

10.1371/journal.pgen.1009608 ◽

2021 ◽

Vol 17 (6) ◽

pp. e1009608

Author(s):

Jia-Hui Sun ◽

Jiang Chen ◽

Fernando Eduardo Ayala Valenzuela ◽

Carolyn Brown ◽

Diane Masser-Frye ◽

...

Keyword(s):

Developmental Delay ◽

Prolonged Exposure ◽

De Novo ◽

Missense Variant ◽

Epileptic Encephalopathy ◽

Global Developmental Delay ◽

Decay Kinetics ◽

Gain Of Function ◽

Ca1 Neurons ◽

Pathogenic Variants

The X-linked GRIA3 gene encodes the GLUA3 subunit of AMPA-type glutamate receptors. Pathogenic variants in this gene were previously reported in neurodevelopmental diseases, mostly in male patients but rarely in females. Here we report a de novo pathogenic missense variant in GRIA3 (c.1979G>C; p. R660T) identified in a 1-year-old female patient with severe epilepsy and global developmental delay. When exogenously expressed in human embryonic kidney (HEK) cells, GLUA3_R660T showed slower desensitization and deactivation kinetics compared to wildtype (wt) GLUA3 receptors. Substantial non-desensitized currents were observed with the mutant but not for wt GLUA3 with prolonged exposure to glutamate. When co-expressed with GLUA2, the decay kinetics were similarly slowed in GLUA2/A3_R660T with non-desensitized steady state currents. In cultured cerebellar granule neurons, miniature excitatory postsynaptic currents (mEPSCs) were significantly slower in R660T transfected cells than those expressing wt GLUA3. When overexpressed in hippocampal CA1 neurons by in utero electroporation, the evoked EPSCs and mEPSCs were slower in neurons expressing R660T mutant compared to those expressing wt GLUA3. Therefore our study provides functional evidence that a gain of function (GoF) variant in GRIA3 may cause epileptic encephalopathy and global developmental delay in a female subject by enhancing synaptic transmission.

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Biophysical classification of a CACNA1D de novo mutation as a high-risk mutation for a severe neurodevelopmental disorder

Molecular Autism ◽

10.1186/s13229-019-0310-4 ◽

2020 ◽

Vol 11 (1) ◽

Cited By ~ 10

Author(s):

Nadja T. Hofer ◽

Petronel Tuluc ◽

Nadine J. Ortner ◽

Yuliia V. Nikonishyna ◽

Monica L. Fernándes-Quintero ◽

...

Keyword(s):

High Risk ◽

Neurodevelopmental Disorders ◽

Developmental Disorder ◽

De Novo ◽

Disease Risk ◽

Neurodevelopmental Disorder ◽

Autism Spectrum ◽

De Novo Mutation ◽

Loss Of Function ◽

Gain Of Function

Abstract Background There is increasing evidence that de novo CACNA1D missense mutations inducing increased Cav1.3 L-type Ca2+-channel-function confer a high risk for neurodevelopmental disorders (autism spectrum disorder with and without neurological and endocrine symptoms). Electrophysiological studies demonstrating the presence or absence of typical gain-of-function gating changes could therefore serve as a tool to distinguish likely disease-causing from non-pathogenic de novo CACNA1D variants in affected individuals. We tested this hypothesis for mutation S652L, which has previously been reported in twins with a severe neurodevelopmental disorder in the Deciphering Developmental Disorder Study, but has not been classified as a novel disease mutation. Methods For functional characterization, wild-type and mutant Cav1.3 channel complexes were expressed in tsA-201 cells and tested for typical gain-of-function gating changes using the whole-cell patch-clamp technique. Results Mutation S652L significantly shifted the voltage-dependence of activation and steady-state inactivation to more negative potentials (~ 13–17 mV) and increased window currents at subthreshold voltages. Moreover, it slowed tail currents and increased Ca2+-levels during action potential-like stimulations, characteristic for gain-of-function changes. To provide evidence that only gain-of-function variants confer high disease risk, we also studied missense variant S652W reported in apparently healthy individuals. S652W shifted activation and inactivation to more positive voltages, compatible with a loss-of-function phenotype. Mutation S652L increased the sensitivity of Cav1.3 for inhibition by the dihydropyridine L-type Ca2+-channel blocker isradipine by 3–4-fold. Conclusions and limitations Our data provide evidence that gain-of-function CACNA1D mutations, such as S652L, but not loss-of-function mutations, such as S652W, cause high risk for neurodevelopmental disorders including autism. This adds CACNA1D to the list of novel disease genes identified in the Deciphering Developmental Disorder Study. Although our study does not provide insight into the cellular mechanisms of pathological Cav1.3 signaling in neurons, we provide a unifying mechanism of gain-of-function CACNA1D mutations as a predictor for disease risk, which may allow the establishment of a more reliable diagnosis of affected individuals. Moreover, the increased sensitivity of S652L to isradipine encourages a therapeutic trial in the two affected individuals. This can address the important question to which extent symptoms are responsive to therapy with Ca2+-channel blockers.

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Biallelic and monoallelic variants in PLXNA1 are implicated in a novel neurodevelopmental disorder with variable cerebral and eye anomalies

Genetics in Medicine ◽

10.1038/s41436-021-01196-9 ◽

2021 ◽

Author(s):

Gabriel C. Dworschak ◽

Jaya Punetha ◽

Jeshurun C. Kalanithy ◽

Enrico Mingardo ◽

Haktan B. Erdem ◽

...

Keyword(s):

Developmental Delay ◽

De Novo ◽

Neurodevelopmental Disorder ◽

Structural Modeling ◽

Dominant Negative ◽

Dominant Negative Effect ◽

Global Developmental Delay ◽

Downstream Signaling ◽

Negative Effect

Abstract Purpose To investigate the effect of PLXNA1 variants on the phenotype of patients with autosomal dominant and recessive inheritance patterns and to functionally characterize the zebrafish homologs plxna1a and plxna1b during development. Methods We assembled ten patients from seven families with biallelic or de novo PLXNA1 variants. We describe genotype–phenotype correlations, investigated the variants by structural modeling, and used Morpholino knockdown experiments in zebrafish to characterize the embryonic role of plxna1a and plxna1b. Results Shared phenotypic features among patients include global developmental delay (9/10), brain anomalies (6/10), and eye anomalies (7/10). Notably, seizures were predominantly reported in patients with monoallelic variants. Structural modeling of missense variants in PLXNA1 suggests distortion in the native protein. Our zebrafish studies enforce an embryonic role of plxna1a and plxna1b in the development of the central nervous system and the eye. Conclusion We propose that different biallelic and monoallelic variants in PLXNA1 result in a novel neurodevelopmental syndrome mainly comprising developmental delay, brain, and eye anomalies. We hypothesize that biallelic variants in the extracellular Plexin-A1 domains lead to impaired dimerization or lack of receptor molecules, whereas monoallelic variants in the intracellular Plexin-A1 domains might impair downstream signaling through a dominant-negative effect.

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Delineating the molecular and phenotypic spectrum of the SETD1B-related syndrome

Genetics in Medicine ◽

10.1038/s41436-021-01246-2 ◽

2021 ◽

Author(s):

Marjolein J. A. Weerts ◽

Kristina Lanko ◽

Francisco J. Guzmán-Vega ◽

Adam Jackson ◽

Reshmi Ramakrishnan ◽

...

Keyword(s):

Intellectual Disability ◽

Developmental Delay ◽

Neurodevelopmental Disorder ◽

Epileptic Activity ◽

Language Delay ◽

Global Developmental Delay ◽

Sequence Variants ◽

Loss Of Function ◽

Phenotypic Spectrum

Abstract Purpose Pathogenic variants in SETD1B have been associated with a syndromic neurodevelopmental disorder including intellectual disability, language delay, and seizures. To date, clinical features have been described for 11 patients with (likely) pathogenic SETD1B sequence variants. This study aims to further delineate the spectrum of the SETD1B-related syndrome based on characterizing an expanded patient cohort. Methods We perform an in-depth clinical characterization of a cohort of 36 unpublished individuals with SETD1B sequence variants, describing their molecular and phenotypic spectrum. Selected variants were functionally tested using in vitro and genome-wide methylation assays. Results Our data present evidence for a loss-of-function mechanism of SETD1B variants, resulting in a core clinical phenotype of global developmental delay, language delay including regression, intellectual disability, autism and other behavioral issues, and variable epilepsy phenotypes. Developmental delay appeared to precede seizure onset, suggesting SETD1B dysfunction impacts physiological neurodevelopment even in the absence of epileptic activity. Males are significantly overrepresented and more severely affected, and we speculate that sex-linked traits could affect susceptibility to penetrance and the clinical spectrum of SETD1B variants. Conclusion Insights from this extensive cohort will facilitate the counseling regarding the molecular and phenotypic landscape of newly diagnosed patients with the SETD1B-related syndrome.

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Recapitulation of the EEF1A2 D252H neurodevelopmental disorder-causing missense mutation in mice reveals a toxic gain of function

Human Molecular Genetics ◽

10.1093/hmg/ddaa042 ◽

2020 ◽

Vol 29 (10) ◽

pp. 1592-1606 ◽

Cited By ~ 1

Author(s):

Faith C J Davies ◽

Jilly E Hope ◽

Fiona McLachlan ◽

Grant F Marshall ◽

Laura Kaminioti-Dumont ◽

...

Keyword(s):

Missense Mutation ◽

Molecular Modelling ◽

Neurodevelopmental Disorders ◽

De Novo ◽

Neurodevelopmental Disorder ◽

Elongation Factor ◽

Missense Mutations ◽

Loss Of Function ◽

De Novo Mutations ◽

Gain Of Function

Abstract Heterozygous de novo mutations in EEF1A2, encoding the tissue-specific translation elongation factor eEF1A2, have been shown to cause neurodevelopmental disorders including often severe epilepsy and intellectual disability. The mutational profile is unusual; ~50 different missense mutations have been identified but no obvious loss of function mutations, though large heterozygous deletions are known to be compatible with life. A key question is whether the heterozygous missense mutations operate through haploinsufficiency or a gain of function mechanism, an important prerequisite for design of therapeutic strategies. In order both to address this question and to provide a novel model for neurodevelopmental disorders resulting from mutations in EEF1A2, we created a new mouse model of the D252H mutation. This mutation causes the eEF1A2 protein to be expressed at lower levels in brain but higher in muscle in the mice. We compared both heterozygous and homozygous D252H and null mutant mice using behavioural and motor phenotyping alongside molecular modelling and analysis of binding partners. Although the proteomic analysis pointed to a loss of function for the D252H mutant protein, the D252H homozygous mice were more severely affected than null homozygotes on the same genetic background. Mice that are heterozygous for the missense mutation show no behavioural abnormalities but do have sex-specific deficits in body mass and motor function. The phenotyping of our novel mouse lines, together with analysis of molecular modelling and interacting proteins, suggest that the D252H mutation results in a gain of function.

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239-kb Microdeletion Spanning KMT2E in a Child with Developmental Delay: Further Delineation of the Phenotype

Molecular Syndromology ◽

10.1159/000516635 ◽

2021 ◽

pp. 1-6

Author(s):

Konstantina Kosma ◽

Konstantinos Varvagiannis ◽

Anastasios Mitrakos ◽

Maria Tsipi ◽

Joanne Traeger-Synodinos ◽

...

Keyword(s):

Developmental Delay ◽

De Novo ◽

Neurodevelopmental Disorder ◽

Dominant Negative ◽

Dominant Negative Effect ◽

Chromosomal Microarray ◽

Global Developmental Delay ◽

Common Mechanism ◽

Facial Dysmorphism ◽

Gi Symptoms

Pathogenic KMT2E variants underly O'Donnell-Luria-Rodan syndrome, a recently described neurodevelopmental disorder characterized by global developmental delay, variable degrees of intellectual disability, and subtle facial dysmorphism. Less common findings include autism, seizures, gastrointestinal (GI) problems, and abnormal head circumference. Occurrence of mostly truncating variants as well as the similar phenotype observed in individuals with deletions spanning KMT2E suggest haploinsufficiency of this gene as a common mechanism for the disorder, while a gain-of-function or dominant-negative effect cannot be ruled out for some missense variants. Deletions reported in the literature encompass several additional known or presumed haploinsufficient genes, thus leading to more complex phenotypes. Here, we describe a male with antenatal onset hydronephrosis, hypotonia, global developmental delay, prominent GI symptoms as well as facial dysmorphism. Chromosomal microarray revealed a 239-kb de novo microdeletion spanning KMT2E and LHFPL3. Clinical presentation of our proband, harboring one of the smallest deletions of the region confirms the core features of this disorder, suggests GI symptoms as a prominent finding in affected individuals while expanding the phenotypic spectrum to abnormalities of the urinary tract.

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Delineating the molecular and phenotypic spectrum of the SETD1B-related syndrome

10.1101/2021.02.11.430742 ◽

2021 ◽

Author(s):

Marjolein J.A. Weerts ◽

Kristina Lanko ◽

Francisco J. Guzmán-Vega ◽

Adam Jackson ◽

Reshmi Ramakrishnan ◽

...

Keyword(s):

Intellectual Disability ◽

Developmental Delay ◽

Neurodevelopmental Disorder ◽

Epileptic Activity ◽

Language Delay ◽

Global Developmental Delay ◽

Sequence Variants ◽

Loss Of Function ◽

Phenotypic Spectrum

ABSTRACTPathogenic variants in SETD1B have been associated with a syndromic neurodevelopmental disorder including intellectual disability, language delay and seizures. To date, clinical features have been described for eleven patients with (likely) pathogenic SETD1B sequence variants. We perform an in-depth clinical characterization of a cohort of 36 unpublished individuals with SETD1B sequence variants, describing their molecular and phenotypic spectrum. Selected variants were functionally tested using in vitro and genome-wide methylation assays. Our data present evidence for a loss-of-function mechanism of SETD1B variants, resulting in a core clinical phenotype of global developmental delay, language delay including regression, intellectual disability, autism and other behavioral issues, and variable epilepsy phenotypes. Developmental delay appeared to precede seizure onset, suggesting SETD1B dysfunction impacts physiological neurodevelopment even in the absence of epileptic activity. Interestingly, males are significantly overrepresented and more severely affected, and we speculate that sex-linked traits could affect susceptibility to penetrance and the clinical spectrum of SETD1B variants. Finally, despite the possibility of non-redundant contributions of SETD1B and its paralogue SETD1A to epigenetic control, the clinical phenotypes of the related disorders share many similarities, indicating that elucidating shared and divergent downstream targets of both genes will help to understand the mechanism leading to the neurobehavioral phenotypes. Insights from this extensive cohort will facilitate the counseling regarding the molecular and phenotypic landscape of newly diagnosed patients with the SETD1B-related syndrome.

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Spectrum of Movement Disorders in GNAO1 Encephalopathy: in-depth Phenotyping and Literature Review

10.21203/rs.3.rs-60924/v1 ◽

2020 ◽

Author(s):

Soo Yeon Kim ◽

YoungKyu Shim ◽

Young Joon Ko ◽

Soojin Park ◽

Se Song Jang ◽

...

Keyword(s):

Literature Review ◽

Exome Sequencing ◽

Developmental Delay ◽

Whole Exome Sequencing ◽

De Novo ◽

Neurodevelopmental Disorder ◽

Epileptic Encephalopathy ◽

Global Developmental Delay ◽

Pathogenic Variants ◽

Whole Exome

Abstract Background GNAO1 encephalopathy is a rare neurodevelopmental disorder characterized by distinct movement presentations and early onset epileptic encephalopathy. Here, we report the in-depth phenotyping of genetically confirmed patients with GNAO1 encephalopathy, focusing on movement presentations. Results Six patients who participated in Korean Undiagnosed Disease Program were diagnosed to have pathogenic or likely pathogenic variants in GNAO1 using whole exome sequencing. All medical records and personal video clips were analyzed with a literature review. Three of the 6 patients were male. Mean follow-up duration was 39 months (range, 7–78 months) and age at last examination was 8.0 years (range, 3.3–16.9 years). Initial complaints were hypotonia or developmental delay in 5 and right-hand clumsiness in 1 patient, which were noticed at 20 months of age on average (range, 0–75 months). All patients showed global developmental delay and 4 had severely retarded development. Five patients (5/6, 83.3%) had many different movement symptoms with various onset and progression. The symptoms included stereotyped hands movement, non-epileptic myoclonus, dyskinesia, dystonia and choreoathetosis. Whole exome sequencing identified 6 different variants in GNAO1. Three were novel de novo variants and atypical presentation was noted in a patient. One variant turned out to be inherited from patient’s mother who had mosaic variant. Distinct phenotypes in patients with variant p.Glu246Lys and p.Arg209His were elucidated by in-depth phenotyping and literature review. Conclusions We reported 6 patients with GNAO1 encephalopathy showing an extremely diverse clinical spectrum on video. Some characteristic movement features identified by careful inspection may also provide important diagnostic insight and practice guidelines.

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Spectrum of Movement Disorders in GNAO1 Encephalopathy: In-Depth Phenotyping and Case-by-Case Analysis

10.21203/rs.3.rs-60924/v2 ◽

2020 ◽

Author(s):

Soo Yeon Kim ◽

YoungKyu Shim ◽

Young Joon Ko ◽

Soojin Park ◽

Se Song Jang ◽

...

Keyword(s):

Literature Review ◽

Exome Sequencing ◽

Developmental Delay ◽

Whole Exome Sequencing ◽

De Novo ◽

Neurodevelopmental Disorder ◽

Epileptic Encephalopathy ◽

Global Developmental Delay ◽

Pathogenic Variants ◽

Whole Exome

Abstract Background: GNAO1 encephalopathy is a rare neurodevelopmental disorder characterized by distinct movement presentations and early onset epileptic encephalopathy. Here, we report the in-depth phenotyping of genetically confirmed patients with GNAO1 encephalopathy, focusing on movement presentations.Results: Six patients who participated in Korean Undiagnosed Disease Program were diagnosed to have pathogenic or likely pathogenic variants in GNAO1 using whole exome sequencing. All medical records and personal video clips were analyzed with a literature review. Three of the 6 patients were male. Median follow-up duration was 41 months (range, 7–78 months) and age at last examination was 7.4 years (range, 3.3–16.9 years). Initial complaints were hypotonia or developmental delay in 5 and right-hand clumsiness in 1 patient, which were noticed at median age of 3 months (range, 0–75 months). All patients showed global developmental delay and 4 had severely retarded development. Five patients (5/6, 83.3%) had many different movement symptoms with various onset and progression. The symptoms included stereotyped hands movement, non-epileptic myoclonus, dyskinesia, dystonia and choreoathetosis. Whole exome sequencing identified 6 different variants in GNAO1. Three were novel de novo variants and atypical presentation was noted in a patient. One variant turned out to be inherited from patient’s mother who had mosaic variant. Distinct and characteristics movement phenotypes in patients with variant p.Glu246Lys and p.Arg209His were elucidated by in-depth phenotyping and literature review. Conclusions: We reported 6 patients with GNAO1 encephalopathy showing an extremely diverse clinical spectrum on video. Some characteristic movement features identified by careful inspection may also provide important diagnostic insight and practice guidelines.

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