scholarly journals Pathogenic DDX3X mutations impair RNA metabolism and neurogenesis during fetal cortical development

2018 ◽  
Author(s):  
Ashley L. Lennox ◽  
Ruiji Jiang ◽  
Lindsey Suit ◽  
Brieana Fregeau ◽  
Charles J. Sheehan ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
De Novo ◽  
Cortical Development ◽  
Rna Helicase ◽  
Mouse Genetics ◽  
Missense Mutations ◽  
Neurodevelopmental Disease ◽  
Brain Malformations ◽  
And Migration ◽  
Human And Mouse

AbstractDe novo germline mutations in the RNA helicase DDX3X account for 1-3% of unexplained intellectual disability (ID) cases in females, and are associated with autism, brain malformations, and epilepsy. Yet, the developmental and molecular mechanisms by which DDX3X mutations impair brain function are unknown. Here we use human and mouse genetics, and cell biological and biochemical approaches to elucidate mechanisms by which pathogenic DDX3X variants disrupt brain development. We report the largest clinical cohort to date with DDX3X mutations (n=78), demonstrating a striking correlation between recurrent dominant missense mutations, polymicrogyria, and the most severe clinical outcomes. We show that Ddx3x controls cortical development by regulating neuronal generation and migration. Severe DDX3X missense mutations profoundly disrupt RNA helicase activity and induce ectopic RNA-protein granules and aberrant translation in neural progenitors and neurons. Together, our study demonstrates novel mechanisms underlying DDX3X syndrome, and highlights roles for RNA-protein aggregates in the pathogenesis of neurodevelopmental disease.

scholarly journals Tubulinopathy mutations in TUBA1A that disrupt neuronal morphogenesis and migration override XMAP215/Stu2 regulation of microtubule dynamics

2021 ◽  
Author(s):  
Katelyn J. Hoff ◽  
Jayne E. Aiken ◽  
Mark A. Gutierrez ◽  
Santos J. Franco ◽  
Jeffrey K. Moore
Keyword(s):  
Molecular Mechanisms ◽  
Ectopic Expression ◽  
Intermediate Phenotype ◽  
Neuronal Cultures ◽  
Missense Mutations ◽  
Primary Neuronal Cultures ◽  
Neuronal Morphogenesis ◽  
Brain Malformations ◽  
Wide Range ◽  
And Migration

ABSTRACTHeterozygous, missense mutations in α- or β-tubulin genes are associated with a wide range of human brain malformations, known as tubulinopathies. We seek to understand whether a mutation’s impact at the molecular and cellular levels scale with the severity of brain malformation. Here we focus on two mutations at the valine 409 residue of TUBA1A, V409I and V409A, identified in patients with pachygyria or lissencephaly, respectively. We find that ectopic expression of TUBA1A-V409I/A mutants disrupt neuronal migration in mice and promote excessive neurite branching and delayed retraction in primary neuronal cultures, accompanied by increased microtubule acetylation. To determine the molecular mechanisms, we modeled the V409I/A mutants in budding yeast and found that they promote intrinsically faster microtubule polymerization rates in cells and in reconstitution experiments with purified tubulin. In addition, V409I/A mutants decrease the recruitment of XMAP215/Stu2 to plus ends and ablate tubulin binding to TOG domains. In each assay tested, the TUBA1A-V409I mutant exhibits an intermediate phenotype between wild type and the more severe TUBA1A-V409A, reflecting the severity observed in brain malformations. Together, our data support a model in which the V409I/A mutations may limit tubulin conformational states and thereby disrupt microtubule regulation during neuronal morphogenesis and migration.

scholarly journals TUBA1A mutations identified in lissencephaly patients dominantly disrupt neuronal migration and impair dynein activity

2018 ◽  
Vol 28 (8) ◽  
pp. 1227-1243 ◽  
Author(s):  
Jayne Aiken ◽  
Jeffrey K Moore ◽  
Emily A Bates
Keyword(s):  
Motor Activity ◽  
Neuronal Migration ◽  
Ectopic Expression ◽  
Missense Mutations ◽  
Microtubule Cytoskeleton ◽  
Tubulin Genes ◽  
Dynein Motor ◽  
Brain Malformations ◽  
Developing Mouse Brain ◽  
And Migration

Abstract The microtubule cytoskeleton supports diverse cellular morphogenesis and migration processes during brain development. Mutations in tubulin genes are associated with severe human brain malformations known as ‘tubulinopathies’; however, it is not understood how molecular-level changes in microtubule subunits lead to brain malformations. In this study, we demonstrate that missense mutations affecting arginine at position 402 (R402) of TUBA1A α-tubulin selectively impair dynein motor activity and severely and dominantly disrupt cortical neuronal migration. TUBA1A is the most commonly affected tubulin gene in tubulinopathy patients, and mutations altering R402 account for 30% of all reported TUBA1A mutations. We show for the first time that ectopic expression of TUBA1A-R402C and TUBA1A-R402H patient alleles is sufficient to dominantly disrupt cortical neuronal migration in the developing mouse brain, strongly supporting a causal role in the pathology of brain malformation. To isolate the precise molecular impact of R402 mutations, we generated analogous R402C and R402H mutations in budding yeast α-tubulin, which exhibit a simplified microtubule cytoskeleton. We find that R402 mutant tubulins assemble into microtubules that support normal kinesin motor activity but fail to support the activity of dynein motors. Importantly, the level of dynein impairment scales with the expression level of the mutant in the cell, suggesting a ‘poisoning’ mechanism in which R402 mutant α-tubulin acts dominantly by populating microtubules with defective binding sites for dynein. Based on our results, we propose a new model for the molecular pathology of tubulinopathies that may also extend to other tubulin-related neuropathies.

scholarly journals A Novel De Novo TUBB3 Variant Causing Developmental Delay, Epilepsy and Mild Ophthalmological Symptoms in a Chinese Child.

2021 ◽  
Author(s):  
Jiao Xue ◽  
Zhenfeng Song ◽  
Shuyin Ma ◽  
Zhi Yi ◽  
Chengqing Yang ◽  
...  
Keyword(s):  
Developmental Delay ◽  
De Novo ◽  
Febrile Seizures ◽  
Extraocular Muscles ◽  
Global Developmental Delay ◽  
Missense Mutations ◽  
Brain Malformations ◽  
Heterozygous Variant ◽  
Type 3 ◽  
Congenital Fibrosis

Abstract Heterozygous missense mutations in TUBB3 have been implicated in various neurological disorders encompassing either isolated congenital fibrosis of the extraocular muscles type 3 (CFEOM3) or complex cortical dysplasia with other brain malformations 1 (CDCBM1). The description of seizures in patients with TUBB3 mutations is rare. Here, we reported a patient who had febrile seizures before and focal seizure this time, which was diagnosed as epilepsy in combination with an abnormal EEG. MRI showed hypoplastic corpus callosum. Mutation analysis showed a novel de novo heterozygous variant of TUBB3 gene (NM_006086), c.763G>A (p.V255I). He had global developmental delay, photophobia and elliptic pupil, but lacking extraocular muscles involvement and malformations of cortical development, which might be a less severe phenotype of TUBB3 mutations. This was the first report of elliptic pupil in patients with TUBB3 mutations and expanded the spectrum of TUBB3 phenotypes. It indicated that the phenotypic range of TUBB3 mutations might be more continuous than discrete, with a severity ranging from mild to severe. Further studies are needed to elucidate the complete spectrum of TUBB3-related phenotypes.

scholarly journals Neurodevelopmental disease genes implicated by de novo mutation and CNV morbidity

2017 ◽  
Author(s):  
Bradley P. Coe ◽  
Holly A.F. Stessman ◽  
Arvis Sulovari ◽  
Madeleine Geisheker ◽  
Fereydoun Hormozdiari ◽  
...  
Keyword(s):  
Developmental Delay ◽  
De Novo ◽  
Gene Dosage ◽  
De Novo Mutation ◽  
Disease Genes ◽  
Missense Mutations ◽  
Genomic Disorder ◽  
Network Analyses ◽  
Neurodevelopmental Disease ◽  
Deletion Syndromes

ABSTRACTWe combined de novo mutation (DNM) data from 10,927 cases of developmental delay and autism to identify 301 candidate neurodevelopmental disease genes showing an excess of missense and/or likely gene-disruptive (LGD) mutations. 164 genes were predicted by two different DNM models, including 116 genes with an excess of LGD mutations. Among the 301 genes, 76% show DNM in both autism and intellectual disability/developmental delay cohorts where they occur in 10.3% and 28.4% of the cases, respectively. Intersecting these results with copy number variation (CNV) morbidity data identifies a significant enrichment for the intersection of our gene set and genomic disorder regions (36/301, LR+ 2.53, p=0.0005). This analysis confirms many recurrent LGD genes and CNV deletion syndromes (e.g., KANSL1, PAFAH1B1, RA1, etc.), consistent with a model of haploinsufficiency. We also identify genes with an excess of missense DNMs overlapping deletion syndromes (e.g., KIF1A and the 2q37 deletion) as well as duplication syndromes, such as recurrent MAPK3 missense mutations within the chromosome 16p11.2 duplication, recurrent CHD4 missense DNMs in the 12p13 duplication region, and recurrent WDFY4 missense DNMs in the 10q11.23 duplication region. Finally, we also identify pathogenic CNVs overlapping more than one recurrently mutated gene (e.g., Sotos and Kleefstra syndromes) raising the possibility that multiple gene-dosage imbalances may contribute to phenotypic complexity of these disorders. Network analyses of genes showing an excess of DNMs confirm previous well-known enrichments but also highlight new functional networks, including cell-specific enrichments in the D1+ and D2+ spiny neurons of the striatum for both recurrently mutated genes and genes where missense mutations cluster.

scholarly journals Proteomic Analysis of KCNK3 Loss of Expression Identified Dysregulated Pathways in Pulmonary Vascular Cells

2020 ◽  
Vol 21 (19) ◽  
pp. 7400 ◽  
Author(s):  
Hélène Le Ribeuz ◽  
Florent Dumont ◽  
Guillaume Ruellou ◽  
Mélanie Lambert ◽  
Thierry Balliau ◽  
...  
Keyword(s):  
Signaling Pathways ◽  
Molecular Mechanisms ◽  
De Novo ◽  
Heme Oxygenase 1 ◽  
Loss Of Function ◽  
Apoptosis Resistance ◽  
Purine Nucleotides ◽  
Proteomic Approach ◽  
Pulmonary Arterial ◽  
And Migration

The physiopathology of pulmonary arterial hypertension (PAH) is characterized by pulmonary artery smooth muscle cell (PASMC) and endothelial cell (PAEC) dysfunction, contributing to pulmonary arterial obstruction and PAH progression. KCNK3 loss of function mutations are responsible for the first channelopathy identified in PAH. Loss of KCNK3 function/expression is a hallmark of PAH. However, the molecular mechanisms involved in KCNK3 dysfunction are mostly unknown. To identify the pathological molecular mechanisms downstream of KCNK3 in human PASMCs (hPASMCs) and human PAECs (hPAECs), we used a Liquid Chromatography-Tandem Mass Spectrometry-based proteomic approach to identify the molecular pathways regulated by KCNK3. KCNK3 loss of expression was induced in control hPASMCs or hPAECs by specific siRNA targeting KCNK3. We found that the loss of KCNK3 expression in hPAECs and hPASMCs leads to 326 and 222 proteins differentially expressed, respectively. Among them, 53 proteins were common to hPAECs and hPASMCs. The specific proteome remodeling in hPAECs in absence of KCNK3 was mostly related to the activation of glycolysis, the superpathway of methionine degradation, and the mTOR signaling pathways, and to a reduction in EIF2 signaling pathways. In hPASMCs, we found an activation of the PI3K/AKT signaling pathways and a reduction in EIF2 signaling and the Purine Nucleotides De Novo Biosynthesis II and IL-8 signaling pathways. Common to hPAECs and hPASMCs, we found that the loss of KCNK3 expression leads to the activation of the NRF2-mediated oxidative stress response and a reduction in the interferon pathway. In the hPAECs and hPASMCs, we found an increased expression of HO-1 (heme oxygenase-1) and a decreased IFIT3 (interferon-induced proteins with tetratricopeptide repeats 3) (confirmed by Western blotting), allowing us to identify these axes to understand the consequences of KCNK3 dysfunction. Our experiments, based on the loss of KCNK3 expression by a specific siRNA strategy in control hPAECs and hPASMCs, allow us to identify differences in the activation of several signaling pathways, indicating the key role played by KCNK3 dysfunction in the development of PAH. Altogether, these results allow us to better understand the consequences of KCNK3 dysfunction and suggest that KCNK3 loss of expression acts in favor of the proliferation and migration of hPASMCs and promotes the metabolic shift and apoptosis resistance of hPAECs.

scholarly journals Genetic heterogeneity of polymicrogyria: study of 123 patients using deep sequencing

2020 ◽  
Author(s):  
Chloe A Stutterd ◽  
Stefanie Brock ◽  
Katrien Stouffs ◽  
Miriam Fanjul-Fernandez ◽  
Paul J Lockhart ◽  
...  
Keyword(s):  
Deep Sequencing ◽  
De Novo ◽  
Diagnostic Yield ◽  
Cortical Development ◽  
Copy Number Variants ◽  
Additional Patient ◽  
Cognitive Disability ◽  
Pathogenic Variants ◽  
Brain Malformations ◽  
Gene Panels

Abstract Polymicrogyria is a malformation of cortical development characterized by overfolding and abnormal lamination of the cerebral cortex. Manifestations include epilepsy, speech disturbance and motor and cognitive disability. Causes include acquired prenatal insults and inherited and de novo genetic variants. The proportion of patients with polymicrogyria and a causative germline or mosaic variant is not known. The aim of this study was to identify the monogenic causes of polymicrogyria in a heterogeneous cohort of patients reflective of specialized referral services. Patients with polymicrogyria were recruited from two clinical centres in Australia and Belgium. Patients with evidence of congenital cytomegalovirus infection or causative chromosomal copy number variants were excluded. One hundred and twenty-three patients were tested using deep sequencing gene panels including known and candidate genes for malformations of cortical development. Causative and potentially causative variants were identified and correlated with phenotypic features. Pathogenic or likely pathogenic variants were identified in 25/123 (20.3%) patients. A candidate variant was identified for an additional patient but could not be confirmed as de novo, and therefore it was classified as being of uncertain significance with high clinical relevance. Of the 22 dominant variants identified, 5 were mosaic with allele fractions less than 0.33 and the lowest allele fraction 0.09. The most common causative genes were TUBA1A and PIK3R2. The other eleven causative genes were PIK3CA, NEDD4L, COL4A1, COL4A2, GPSM2, GRIN2B, WDR62, TUBB3, TUBB2B, ACTG1 and FH. A genetic cause was more likely to be identified in the presence of an abnormal head size or additional brain malformations suggestive of a tubulinopathy, such as dysmorphic basal ganglia. A gene panel test provides greater sequencing depth and sensitivity for mosaic variants than whole exome or genome sequencing but is limited to the genes included, potentially missing variants in newly discovered genes. The diagnostic yield of 20.3% indicates that polymicrogyria may be associated with genes not yet known to be associated with brain malformations, brain-specific somatic mutations or non-genetic causes.

scholarly journals Neuronal Modeling of Alternating Hemiplegia of Childhood Reveals Transcriptional Compensation and Replicates a Trigger-Induced Phenotype

2020 ◽  
Author(s):  
John P. Snow ◽  
Grant Westlake ◽  
Lindsay K. Klofas ◽  
Soyoun Jeon ◽  
Laura C. Armstrong ◽  
...  
Keyword(s):  
Microelectrode Array ◽  
De Novo ◽  
Control Cell ◽  
Disease Model ◽  
Patient Specific ◽  
Neuronal Cultures ◽  
Missense Mutations ◽  
Unrelated Control ◽  
Neurodevelopmental Disease ◽  
Alternating Hemiplegia Of Childhood

ABSTRACTAlternating hemiplegia of childhood (AHC) is a rare neurodevelopmental disease caused by heterozygous de novo missense mutations in the ATP1A3 gene that encodes the neuronal specific α3 subunit of the Na,K-ATPase (NKA) pump. Mechanisms underlying patient episodes including environmental triggers remain poorly understood, and there are no empirically proven treatments for AHC. In this study, we generated patient-specific induced pluripotent stem cells (iPSCs) and isogenic controls for the E815K ATP1A3 mutation that causes the most phenotypically severe form of AHC. Using an in vitro iPSC-derived cortical neuron disease model, we found elevated levels of ATP1A3 mRNA in AHC lines compared to controls, without significant perturbations in protein expression. Microelectrode array analyses demonstrated that in cortical neuronal cultures, ATP1A3+/E815K iPSC-derived neurons displayed a non-significant trend toward less overall activity than neurons differentiated from isogenic mutation-corrected and unrelated control cell lines. However, induction of cellular stress by elevated temperature revealed a hyperactivity phenotype following heat stress in ATP1A3+/E815K lines compared to control lines. Treatment with flunarizine, a drug commonly used to prevent AHC episodes, did not impact this stress-triggered phenotype. These findings support the use of iPSC-derived neuronal cultures for studying complex neurodevelopmental conditions such as AHC and provide a potential route toward future therapeutic screening and mechanistic discovery in a human disease model.

scholarly journals β-Catenin Signalling in Glioblastoma Multiforme and Glioma-Initiating Cells

2012 ◽  
Vol 2012 ◽  
pp. 1-7 ◽  
Author(s):  
Mireia Nager ◽  
Deepshikha Bhardwaj ◽  
Carles Cantí ◽  
Loreta Medina ◽  
Pere Nogués ◽  
...  
Keyword(s):  
Glioblastoma Multiforme ◽  
Molecular Mechanisms ◽  
De Novo ◽  
Cell Types ◽  
Surrounding Tissue ◽  
Glioma Initiating Cells ◽  
And Migration ◽  
Canonical Wnt ◽  
Different Cell Types

Glioblastoma multiforme (GBM) is a commonly occurring brain tumor with a poor prognosis. GBM can develop both “de novo” or evolve from a previous astrocytoma and is characterized by high proliferation and infiltration into the surrounding tissue. Following treatment (surgery, radiotherapy, and chemotherapy), tumors often reappear. Glioma-initiating cells (GICs) have been identified in GBM and are thought to be responsible for tumors initiation, their continued growth, and recurrence. β-catenin, a component of the cell-cell adhesion complex and of the canonical Wnt pathway, regulates proliferation, adhesion, and migration in different cell types. β-catenin and components of the Wnt canonical pathway are commonly overexpressed in GBM. Here, we review previous work on the role of Wnt/β-catenin signalling in glioma initiation, proliferation, and invasion. Understanding the molecular mechanisms regulating GIC biology and glioma progression may help in identifying novel therapeutic targets for GBM treatment.

scholarly journals DRP1-mediated mitochondrial fission is essential to maintain cristae morphology and bioenergetics

2022 ◽  
Author(s):  
Gabriella L. Robertson ◽  
Stellan Riffle ◽  
Mira Patel ◽  
Andrea Marshall ◽  
Heather Beasley ◽  
...  
Keyword(s):  
Cell Fate ◽  
De Novo ◽  
Mitochondrial Dynamics ◽  
Mitochondrial Fission ◽  
Coupling Efficiency ◽  
Acid Oxidation ◽  
Mitochondrial Morphology ◽  
Missense Mutations ◽  
Neurodevelopmental Disease ◽  
Signaling Organelles

Mitochondria and peroxisomes are both dynamic signaling organelles that constantly undergo fission. While mitochondrial fission is known to coordinate cellular metabolism, proliferation, and apoptosis, the physiological relevance of peroxisome dynamics and the implications for cell fate are not fully understood. DRP1 (dynamin-related protein 1) is an essential GTPase that executes both mitochondrial and peroxisomal fission. Patients with de novo heterozygous missense mutations in the gene that encodes DRP1, DNM1L, present with encephalopathy due to defective mitochondrial and peroxisomal fission (EMPF1). EMPF1 is a devastating neurodevelopmental disease with no effective treatment. To interrogate the mechanisms by which DRP1 mutations cause cellular dysfunction, we utilized human-derived fibroblasts from patients with mutations in DRP1 who present with EMPF1. As expected, patient cells display elongated mitochondrial morphology and lack of fission. Patient cells display a lower coupling efficiency of the electron transport chain, increased proton leak, and upregulation of glycolysis. In addition to these metabolic abnormalities, mitochondrial hyperfusion results in aberrant cristae structure and hyperpolarized mitochondrial membrane potential, both of which are tightly linked to the changes in metabolism. Peroxisome structure is also severely elongated in patient cells and results in a potential functional compensation of fatty acid oxidation. Understanding the mechanism by which DRP1 mutations cause these metabolic changes will give insight into the role of mitochondrial dynamics in cristae maintenance and the metabolic capacity of the cell, as well as the disease mechanism underlying EMPF1.

Wnt/beta-catenin and PI3K/Akt/mTOR Signaling Pathways in Glioblastoma: Two Main Targets for Drug Design: A Review

2020 ◽  
Vol 26 (15) ◽  
pp. 1729-1741 ◽  
Author(s):  
Seyed H. Shahcheraghi ◽  
Venant Tchokonte-Nana ◽  
Marzieh Lotfi ◽  
Malihe Lotfi ◽  
Ahmad Ghorbani ◽  
...  
Keyword(s):  
Signaling Pathways ◽  
Molecular Mechanisms ◽  
Cellular Proliferation ◽  
Chemotherapy Resistance ◽  
Wnt Signaling Pathway ◽  
Therapeutic Interventions ◽  
Beta Catenin ◽  
Clinical Prognosis ◽  
Invasion And Migration ◽  
And Migration

: Glioblastoma (GBM) is the most common and malignant astrocytic glioma, accounting for about 90% of all brain tumors with poor prognosis. Despite recent advances in understanding molecular mechanisms of oncogenesis and the improved neuroimaging technologies, surgery, and adjuvant treatments, the clinical prognosis of patients with GBM remains persistently unfavorable. The signaling pathways and the regulation of growth factors of glioblastoma cells are very abnormal. The various signaling pathways have been suggested to be involved in cellular proliferation, invasion, and glioma metastasis. The Wnt signaling pathway with its pleiotropic functions in neurogenesis and stem cell proliferation is implicated in various human cancers, including glioma. In addition, the PI3K/Akt/mTOR pathway is closely related to growth, metabolism, survival, angiogenesis, autophagy, and chemotherapy resistance of GBM. Understanding the mechanisms of GBM’s invasion, represented by invasion and migration, is an important tool in designing effective therapeutic interventions. This review will investigate two main signaling pathways in GBM: PI3K/Akt/mTOR and Wnt/beta-catenin signaling pathways.

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