Autism Spectrum Disorder (ASD) is one of the most prevalent neurodevelopmental disorders, affecting an estimated 1 in 59 children. ASD is highly genetically heterogeneous and may be caused by both ...inheritable and
gene variations. In the past decade, hundreds of genes have been identified that contribute to the serious deficits in communication, social cognition, and behavior that patients often experience. However, these only account for 10-20% of ASD cases, and patients with similar pathogenic variants may be diagnosed on very different levels of the spectrum. In this review, we will describe the genetic landscape of ASD and discuss how genetic modifiers such as copy number variation, single nucleotide polymorphisms, and epigenetic alterations likely play a key role in modulating the phenotypic spectrum of ASD patients. We also consider how genetic modifiers can alter convergent signaling pathways and lead to impaired neural circuitry formation. Lastly, we review sex-linked modifiers and clinical implications. Further understanding of these mechanisms is crucial for both comprehending ASD and for developing novel therapies.
Objective
PCDH19‐related epilepsy is characterized by a distinctive pattern of X‐linked inheritance, where heterozygous females exhibit seizures and hemizygous males are asymptomatic. A cellular ...interference mechanism resulting from the presence of both wild‐type and mutant PCDH19 neurons in heterozygous patients or mosaic carriers of PCDH19 variants has been hypothesized. We aim to investigate seizure susceptibility and progression in the Pchd19 mouse model.
Methods
We assessed seizure susceptibility and progression in the Pcdh19 mouse model using three acute seizure induction paradigms. We first induced focal, clonic seizures using the 6‐Hz psychomotor test. Mice were stimulated with increasing current intensities and graded according to a modified Racine scale. We next induced generalized seizures using flurothyl or pentylenetetrazol (PTZ), both γ‐aminobutyric acid type A receptor function inhibitors, and recorded latencies to myoclonic and generalized tonic‐clonic seizures.
Results
Pcdh19 knockout and heterozygous females displayed increased seizure susceptibility across all current intensities in the 6‐Hz psychomotor test, and increased severity overall. They also exhibited shorter latencies to generalized seizures following flurothyl, but not PTZ, seizure induction. Hemizygous males showed comparable seizure incidence and severity to their wild‐type male littermates across all paradigms tested.
Significance
The heightened susceptibility observed in Pcdh19 knockout females suggests additional mechanisms other than cellular interference are at play in PCDH19‐related epilepsy. Further experiments are needed to understand the variability in seizure susceptibility so that this model can be best utilized toward development of future therapeutic strategies for PCDH19‐related epilepsy.
The MAST family of microtubule-associated serine-threonine kinases (STKs) have distinct expression patterns in the developing and mature human and mouse brain. To date, only MAST1 has been ...conclusively associated with neurological disease, with de novo variants in individuals with a neurodevelopmental disorder, including a mega corpus callosum.
Using exome sequencing, we identify MAST3 missense variants in individuals with epilepsy. We also assess the effect of these variants on the ability of MAST3 to phosphorylate the target gene product ARPP-16 in HEK293T cells.
We identify de novo missense variants in the STK domain in 11 individuals, including 2 recurrent variants p.G510S (n = 5) and p.G515S (n = 3). All 11 individuals had developmental and epileptic encephalopathy, with 8 having normal development prior to seizure onset at <2 years of age. All patients developed multiple seizure types, 9 of 11 patients had seizures triggered by fever and 9 of 11 patients had drug-resistant seizures. In vitro analysis of HEK293T cells transfected with MAST3 cDNA carrying a subset of these patient-specific missense variants demonstrated variable but generally lower expression, with concomitant increased phosphorylation of the MAST3 target, ARPP-16, compared to wild-type. These findings suggest the patient-specific variants may confer MAST3 gain-of-function. Moreover, single-nuclei RNA sequencing and immunohistochemistry shows that MAST3 expression is restricted to excitatory neurons in the cortex late in prenatal development and postnatally.
In summary, we describe MAST3 as a novel epilepsy-associated gene with a potential gain-of-function pathogenic mechanism that may be primarily restricted to excitatory neurons in the cortex. ANN NEUROL 2021;90:274-284.
Objective
To identify causes of the autosomal‐recessive malformation, diencephalic‐mesencephalic junction dysplasia (DMJD) syndrome.
Methods
Eight families with DMJD were studied by whole‐exome or ...targeted sequencing, with detailed clinical and radiological characterization. Patient‐derived induced pluripotent stem cells were derived into neural precursor and endothelial cells to study gene expression.
Results
All patients showed biallelic mutations in the nonclustered protocadherin‐12 (PCDH12) gene. The characteristic clinical presentation included progressive microcephaly, craniofacial dysmorphism, psychomotor disability, epilepsy, and axial hypotonia with variable appendicular spasticity. Brain imaging showed brainstem malformations and with frequent thinned corpus callosum with punctate brain calcifications, reflecting expression of PCDH12 in neural and endothelial cells. These cells showed lack of PCDH12 expression and impaired neurite outgrowth.
Interpretation
DMJD patients have biallelic mutations in PCDH12 and lack of protein expression. These patients present with characteristic microcephaly and abnormalities of white matter tracts. Such pathogenic variants predict a poor outcome as a result of brainstem malformation and evidence of white matter tract defects, and should be added to the phenotypic spectrum associated with PCDH12‐related conditions. Ann Neurol 2018;84:646–655
Primary cilia were the largely neglected nonmotile counterparts of their better-known cousin, the motile cilia. For years these nonmotile cilia were considered evolutionary remnants of little ...consequence to cellular function. Fast forward 10 years and we now recognize primary cilia as key integrators of extracellular ligand-based signaling and cellular polarity, which regulate neuronal cell fate, migration, differentiation, as well as a host of adult behaviors. Important future questions will focus on structure-function relationships, their roles in signaling and disease and as areas of target for treatments.
Primary cilia emerge as key regulators of development, neurogenesis, and signaling in the mammalian brain. Guemez-Gamboa et al. review the status, genetic basis, potential, controversies, and unanswered questions.
PACS1 syndrome is a neurodevelopmental disorder characterized by intellectual disability and distinct craniofacial abnormalities resulting from a de novo p.R203W variant in phosphofurin acidic ...cluster sorting protein 1 (PACS1). PACS1 is known to have functions in the endosomal pathway and nucleus, but how the p.R203W variant affects developing neurons is not fully understood. Here we differentiated stem cells towards neuronal models including cortical organoids to investigate the impact of the PACS1 syndrome-causing variant on neurodevelopment. While few deleterious effects were detected in PACS1
neural precursors, mature PACS1
glutamatergic neurons exhibited impaired expression of genes involved in synaptic signaling processes. Subsequent characterization of neural activity using calcium imaging and multielectrode arrays revealed the p.R203W PACS1 variant leads to a prolonged neuronal network burst duration mediated by an increased interspike interval. These findings demonstrate the impact of the PACS1 p.R203W variant on developing human neural tissue and uncover putative electrophysiological underpinnings of disease.
Docosahexanoic acid (DHA) is the most abundant omega-3 fatty acid in brain, and, although it is considered essential, deficiency has not been linked to disease. Despite the large mass of DHA in ...phospholipids, the brain does not synthesize it. DHA is imported across the blood-brain barrier (BBB) through the major facilitator superfamily domain-containing 2a (MFSD2A) protein. MFSD2A transports DHA as well as other fatty acids in the form of lysophosphatidylcholine (LPC). We identify two families displaying MFSD2A mutations in conserved residues. Affected individuals exhibited a lethal microcephaly syndrome linked to inadequate uptake of LPC lipids. The MFSD2A mutations impaired transport activity in a cell-based assay. Moreover, when expressed in mfsd2aa-morphant zebrafish, mutants failed to rescue microcephaly, BBB breakdown and lethality. Our results establish a link between transport of DHA and LPCs by MFSD2A and human brain growth and function, presenting the first evidence of monogenic disease related to transport of DHA in humans.
Protocadherins (PCDHs) are cell adhesion molecules that regulate many essential neurodevelopmental processes related to neuronal maturation, dendritic arbor formation, axon pathfinding, and synaptic ...plasticity. Biallelic loss-of-function variants in PCDH12 are associated with several neurodevelopmental disorders (NDDs). Despite the highly deleterious outcome resulting from loss of PCDH12, little is known about its role during brain development and disease. Here, we show that PCDH12 loss severely impairs cerebral organoid development, with reduced proliferative areas and disrupted laminar organization. 2D models further show that neural progenitor cells lacking PCDH12 prematurely exit the cell cycle and differentiate earlier when compared with wild type. Furthermore, we show that PCDH12 regulates neuronal migration and suggest that this could be through a mechanism requiring ADAM10-mediated ectodomain shedding and/or membrane recruitment of cytoskeleton regulators. Our results demonstrate a critical involvement of PCDH12 in cortical organoid development, suggesting a potential cause for the pathogenic mechanisms underlying PCDH12-related NDDs.
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•PCDH12 loss of function impairs maturation of cerebral organoids•PCDH12 loss leads to premature neuronal differentiation and defective migration•PCDH12 recruits cytoskeleton dynamics regulators to the cell membrane•PCDH12 facilitates neuronal migration through ADAM10-mediated ectodomain shedding
Loss of PCDH12 function is clinically associated with severe neurodevelopmental disorders, but the pathogenic mechanisms are poorly understood. Rakotomamonjy et al. use stem cell-derived brain organoids to show that the absence of PCDH12 leads to premature neuronal differentiation and defective migration mediated by loss of PCDH12 interaction with cytoskeleton dynamics regulators.
Neurodevelopmental disorders are characterized by complex phenotypes that often result from concomitant dysregulation of cell proliferation, differentiation, or other crucial developmental processes. ...Here, we present a protocol to quantify differentiation of progenitor populations during early stages of neurogenesis in induced pluripotent stem cell (iPSC)-derived cerebral organoids. We describe steps for organoid differentiation and maturation, sample preparation, immunofluorescence, and imaging and analysis using epifluorescence microscopy. This protocol can be used to compare cerebral organoids from control and patient-derived iPSCs.
For complete details on the use and execution of this protocol, please refer to Rakotomamonjy et al. (2023).1
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•Instructions to prepare cerebral organoid samples for immunofluorescence•Detection of neural progenitor and early neuron populations using immunofluorescence•Quantification of progenitor population ratios in cerebral organoids
Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.
Neurodevelopmental disorders are characterized by complex phenotypes that often result from concomitant dysregulation of cell proliferation, differentiation, or other crucial developmental processes. Here, we present a protocol to quantify differentiation of progenitor populations during early stages of neurogenesis in induced pluripotent stem cell (iPSC)-derived cerebral organoids. We describe steps for organoid differentiation and maturation, sample preparation, immunofluorescence, and imaging and analysis using epifluorescence microscopy. This protocol can be used to compare cerebral organoids from control and patient-derived iPSCs.
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