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KIF1A-associated neurological disorder (KAND) is a neurodegenerative and often lethal ultrarare disease with a wide phenotypic spectrum associated with largely heterozygous de novo missense variants in KIF1A. Antisense oligonucleotide treatments represent a promising approach for personalized treatments in ultrarare diseases. Here we report the case of one patient with a severe form of KAND characterized by refractory spells of behavioral arrest and carrying a p.Pro305Leu variant in KIF1A, who was treated with intrathecal injections of an allele-specific antisense oligonucleotide specifically designed to degrade the mRNA from the pathogenic allele. The first intrathecal administration was complicated by an epidural cerebrospinal fluid collection, which resolved spontaneously. Otherwise, the antisense oligonucleotide was safe and well tolerated over the 9-month treatment. Most outcome measures, including severity of the spells of behavioral arrest, number of falls and quality of life, improved. There was little change in the 6-min Walk Test distance, but qualitative changes in gait resulting in meaningful reductions in falls and increasing independence were observed. Cognitive performance was stable and did not degenerate over time. Our findings provide preliminary insights on the safety and efficacy of an allele-specific antisense oligonucleotide as a possible treatment for KAND.

Debate remains around the anatomical origins of specific brain cell subtypes and lineage relationships within the human forebrain1–7. Thus, direct observation in the mature human brain is critical for a complete understanding of its structural organization and cellular origins. Here we utilize brain mosaic variation within specific cell types as distinct indicators for clonal dynamics, denoted as cell-type-specific mosaic variant barcode analysis. From four hemispheres and two different human neurotypical donors, we identified 287 and 780 mosaic variants, respectively, that were used to deconvolve clonal dynamics. Clonal spread and allele fractions within the brain reveal that local hippocampal excitatory neurons are more lineage-restricted than resident neocortical excitatory neurons or resident basal ganglia GABAergic inhibitory neurons. Furthermore, simultaneous genome transcriptome analysis at both a cell-type-specific and a single-cell level suggests a dorsal neocortical origin for a subgroup of DLX1+inhibitory neurons that disperse radially from an origin shared with excitatory neurons. Finally, the distribution of mosaic variants across 17 locations within one parietal lobe reveals that restriction of clonal spread in the anterior–posterior axis precedes restriction in the dorsal–ventral axis for both excitatory and inhibitory neurons. Thus, cell-type-resolved somatic mosaicism can uncover lineage relationships governing the development of the human forebrain.

Membrane-shaping proteins characterized by reticulon homology domains play an important part in the dynamic remodelling of the endoplasmic reticulum (ER). An example of such a protein is FAM134B, which can bind LC3 proteins and mediate the degradation of ER sheets through selective autophagy (ER-phagy)1. Mutations in FAM134Bresult in a neurodegenerative disorder in humans that mainly affects sensory and autonomic neurons2. Here we report that ARL6IP1, another ER-shaping protein that contains a reticulon homology domain and is associated with sensory loss3, interacts with FAM134B and participates in the formation of heteromeric multi-protein clusters required for ER-phagy. Moreover, ubiquitination of ARL6IP1 promotes this process. Accordingly, disruption of Arl6ip1in mice causes an expansion of ER sheets in sensory neurons that degenerate over time. Primary cells obtained from Arl6ip1-deficient mice or from patients display incomplete budding of ER membranes and severe impairment of ER-phagy flux. Therefore, we propose that the clustering of ubiquitinated ER-shaping proteins facilitates the dynamic remodelling of the ER during ER-phagy and is important for neuronal maintenance.

Malformations of cortical development (MCD) are neurological conditions involving focal disruptions of cortical architecture and cellular organization that arise during embryogenesis, largely from somatic mosaic mutations, and cause intractable epilepsy. Identifying the genetic causes of MCD has been a challenge, as mutations remain at low allelic fractions in brain tissue resected to treat condition-related epilepsy. Here we report a genetic landscape from 283 brain resections, identifying 69 mutated genes through intensive profiling of somatic mutations, combining whole-exome and targeted-amplicon sequencing with functional validation including in utero electroporation of mice and single-nucleus RNA sequencing. Genotype–phenotype correlation analysis elucidated specific MCD gene sets associated with distinct pathophysiological and clinical phenotypes. The unique single-cell level spatiotemporal expression patterns of mutated genes in control and patient brains indicate critical roles in excitatory neurogenic pools during brain development and in promoting neuronal hyperexcitability after birth.

Mutations in lipid regulator genes are a frequent cause of autism spectrum disorder, including those regulating phosphatidylinositol (PI) and phosphoinositide 3-kinase signaling. MBOAT7encodes a key acyltransferase in PI synthesis and is mutated in an autism-related condition with neurodevelopmental delay and epilepsy. Using liquid chromatography–tandem mass spectrometry, we analyzed the PI-associated glycerolipidome in mice and humans during neurodevelopment and found dynamic regulation at times corresponding to neural apoptosis in the brains of Mboat7knockout mice. Mboat7function was necessary for polyunsaturated lipid synthesis and cortical neural migration, and loss resulted in massive accumulation of the precursor lysophosphatidylinositol and hyperactive mTOR signaling. Inhibiting mTOR signaling rescued migration defects. Our findings demonstrate roles for lipid remodeling during neurodevelopment and implicate lipid regulation in neuronal migration, revealing potential paths to treatment for MBOAT7deficiency.

BRAT1biallelic variants are associated with rigidity and multifocal seizure syndrome, lethal neonatal (RMFSL), and neurodevelopmental disorder associating cerebellar atrophy with or without seizures syndrome (NEDCAS). To date, forty individuals have been reported in the literature. We collected clinical and molecular data from 57 additional cases allowing us to study a large cohort of 97 individuals and draw phenotype-genotype correlations. Fifty-nine individuals presented with BRAT1-related RMFSL phenotype. Most of them had no psychomotor acquisition (100%), epilepsy (100%), microcephaly (91%), limb rigidity (93%), and died prematurely (93%). Thirty-eight individuals presented a non-lethal phenotype of BRAT1-related NEDCAS phenotype. Seventy-six percent of the patients in this group were able to walk and 68% were able to say at least a few words. Most of them had cerebellar ataxia (82%), axial hypotonia (79%) and cerebellar atrophy (100%). Genotype-phenotype correlations in our cohort revealed that biallelic nonsense, frameshift or inframe deletion/insertion variants result in the severe BRAT1-related RMFSL phenotype (46/46; 100%). In contrast, genotypes with at least one missense were more likely associated with NEDCAS (28/34; 82%). The phenotype of patients carrying splice variants was variable: 41% presented with RMFSL (7/17) and 59% with NEDCAS (10/17).

Brain monoamine vesicular transport disease is an infantile-onset movement disorder that mimics cerebral palsy. In 2013, the homozygous SLC18A2variant, p.Pro387Leu, was first reported as a cause of this rare disorder, and dopamine agonists were efficient for treating affected individuals from a single large family. To date, only 6 variants have been reported. In this study, we evaluated genotype–phenotype correlations in individuals with biallelic SLC18A2variants.

Genome sequencing in the clinic often allows patients to receive a molecular diagnosis. However, variants are most often evaluated for pathogenicity, neglecting potential treatability and thus often yielding limited clinical benefit. Antisense oligonucleotides (ASOs), among others, offer attractive programmable and relatively safe platforms for customized therapy based upon the causative genetic variant. The landscape of ASO-treatable variants is largely uncharted, with new developments emerging for loss-of-function, haploinsufficient, and gain-of-function effects. ASOs can access the transcriptome to target splice-gain variants, poison exons, untranslated/regulatory regions, and naturally occurring antisense transcripts. Here we assess public variant databases and find that approximately half of pathogenic variants have one or more viable avenues for ASO therapy. The future might see medical teams considering “treatability” when interpreting genomic sequencing results to fully realize benefits for patients.

Heterozygous mutations in Bicaudal D2 Drosophila homolog 2 (BICD2) gene, encodes a vesicle transport protein involved in dynein-mediated movement along microtubules, are responsible for an exceedingly rare autosomal dominant spinal muscular atrophy type 2A which starts in the childhood and predominantly effects lower extremities. Recently, a more severe form, type 2B, has also been described. Here, we present a patient born to a consanguineous union and who suffered from intellectual disability, speech delay, epilepsy, happy facial expression, truncal obesity with tappering fingers, and joint hypermobility. Whole-exome sequencing analysis revealed a rare, homozygous missense mutation (c.731T>C; p.Leu244Pro) in BICD2gene. This finding presents the first report in the literature for homozygous BICD2mutations and its association with a Cohen-Like syndrome. Patients presenting with Cohen-Like phenotypes should be further interrogated for mutations in BICD2.

Biallelic INPP4Avariants have recently been associated with severe neurodevelopmental disease in single-case reports. Here, we expand and elucidate the clinical-genetic spectrum and provide a pathomechanistic explanation for genotype-phenotype correlations.

The genetic etiology of autism spectrum disorder (ASD) is multifactorial, but how combinations of genetic factors determine risk is unclear. In a large family sample, we show that genetic loads of rare and polygenic risk are inversely correlated in cases and greater in females than in males, consistent with a liability threshold that differs by sex. De novo mutations (DNMs), rare inherited variants and polygenic scores were associated with various dimensions of symptom severity in children and parents. Parental age effects on risk for ASD in offspring were attributable to a combination of genetic mechanisms, including DNMs that accumulate in the paternal germline and inherited risk that influences behavior in parents. Genes implicated by rare variants were enriched in excitatory and inhibitory neurons compared with genes implicated by common variants. Our results suggest that a phenotypic spectrum of ASD is attributable to a spectrum of genetic factors that impact different neurodevelopmental processes.

The structure of the human neocortex underlies species-specific traits and reflects intricate developmental programs. Here we sought to reconstruct processes that occur during early development by sampling adult human tissues. We analysed neocortical clones in a post-mortem human brain through a comprehensive assessment of brain somatic mosaicism, acting as neutral lineage recorders1,2. We combined the sampling of 25 distinct anatomic locations with deep whole-genome sequencing in a neurotypical deceased individual and confirmed results with 5 samples collected from each of three additional donors. We identified 259 bona fide mosaic variants from the index case, then deconvolved distinct geographical, cell-type and clade organizations across the brain and other organs. We found that clones derived after the accumulation of 90–200 progenitors in the cerebral cortex tended to respect the midline axis, well before the anterior–posterior or ventral–dorsal axes, representing a secondary hierarchy following the overall patterning of forebrain and hindbrain domains. Clones across neocortically derived cells were consistent with a dual origin from both dorsal and ventral cellular populations, similar to rodents, whereas the microglia lineage appeared distinct from other resident brain cells. Our data provide a comprehensive analysis of brain somatic mosaicism across the neocortex and demonstrate cellular origins and progenitor distribution patterns within the human brain.

Pathogenic variants in Lysyl-tRNA synthetase 1 (KARS1) have increasingly been recognized as a cause of early-onset complex neurological phenotypes. To advance the timely diagnosis of KARS1-related disorders, we sought to delineate its phenotype and generate a disease model to understand its function in vivo.

Clinical evidence suggests the central nervous system is frequently impacted by SARS-CoV-2 infection, either directly or indirectly, although the mechanisms are unclear. Pericytes are perivascular cells within the brain that are proposed as SARS-CoV-2 infection points. Here we show that pericyte-like cells (PLCs), when integrated into a cortical organoid, are capable of infection with authentic SARS-CoV-2. Before infection, PLCs elicited astrocytic maturation and production of basement membrane components, features attributed to pericyte functions in vivo. While traditional cortical organoids showed little evidence of infection, PLCs within cortical organoids served as viral ‘replication hubs’, with virus spreading to astrocytes and mediating inflammatory type I interferon transcriptional responses. Therefore, PLC-containing cortical organoids (PCCOs) represent a new ‘assembloid’ model that supports astrocytic maturation as well as SARS-CoV-2 entry and replication in neural tissue; thus, PCCOs serve as an experimental model for neural infection.

HEAT repeats are 37–47 amino acid flexible tandem repeat structural motifs occurring in a wide variety of eukaryotic proteins with diverse functions. Due to their ability to undergo elastic conformational changes, they often serve as scaffolds at sites of protein interactions. Here, we describe four affected children from two families presenting with pontocerebellar hypoplasia manifest clinically with neonatal seizures, severe intellectual disability, and motor delay. Whole exome sequencing identified biallelic variants at predicted splice sites in intron 31 of HEATR5B, encoding the HEAT repeat-containing protein 5B segregating in a recessive fashion. Aberrant splicing was found in patient fibroblasts, which correlated with reduced levels of HEATR5B protein. HEATR5Bis expressed during brain development in human, and we failed to recover live-born homozygous Heatr5bknockout mice. Taken together, our results implicate loss of HEATR5Bin pontocerebellar hypoplasia.

Brain somatic mosaicism is linked to several neurological disorders and is thought to arise post-zygotically. A study suggests that pre-zygotic aneuploidy followed by post-zygotic partial reversion leads to a recurrent form of brain mosaicism-related epilepsy.

Trafficking protein particle (TRAPP) complexes, which include the TRAPPC4 protein, regulate membrane trafficking between lipid organelles in a process termed vesicular tethering. TRAPPC4was recently implicated in a recessive neurodevelopmental condition in four unrelated families due to a shared c.454+3A>G splice variant. Here, we report 23 patients from 17 independent families with an early-infantile-onset neurodegenerative presentation, where we also identified the homozygous variant hg38:11:119020256 A>G (NM_016146.5:c.454+3A>G) in TRAPPC4through exome or genome sequencing. No other clinically relevant TRAPPC4variants were identified among any of over 10,000 patients with neurodevelopmental conditions. We found the carrier frequency of TRAPPC4c.454+3A>G was 2.4–5.4 per 10,000 healthy individuals. Affected individuals with the homozygous TRAPPC4c.454+3A>G variant showed profound psychomotor delay, developmental regression, early-onset epilepsy, microcephaly and progressive spastic tetraplegia. Based upon RNA sequencing, the variant resulted in partial exon 3 skipping and generation of an aberrant transcript owing to use of a downstream cryptic splice donor site, predicting a premature stop codon and nonsense mediated decay. These data confirm the pathogenicity of the TRAPPC4c.454+3A>G variant, and refine the clinical presentation of TRAPPC4-related encephalopathy.

BackgroundPathogenic variants of GNB5encoding the β5subunit of the guanine nucleotide-binding protein cause IDDCA syndrome, an autosomal recessive neurodevelopmental disorder associated with cognitive disability and cardiac arrhythmia, particularly severe bradycardia.MethodsWe used echocardiography and telemetric ECG recordings to investigate consequences of Gnb5loss in mouse.ResultsWe delineated a key role of Gnb5in heart sinus conduction and showed that Gnb5-inhibitory signalling is essential for parasympathetic control of heart rate (HR) and maintenance of the sympathovagal balance. Gnb5−/−mice were smaller and had a smaller heart than Gnb5+/+and Gnb5+/−, but exhibited better cardiac function. Lower autonomic nervous system modulation through diminished parasympathetic control and greater sympathetic regulation resulted in a higher baseline HR in Gnb5−/−mice. In contrast, Gnb5−/−mice exhibited profound bradycardia on treatment with carbachol, while sympathetic modulation of the cardiac stimulation was not altered. Concordantly, transcriptome study pinpointed altered expression of genes involved in cardiac muscle contractility in atria and ventricles of knocked-out mice. Homozygous Gnb5loss resulted in significantly higher frequencies of sinus arrhythmias. Moreover, we described 13 affected individuals, increasing the IDDCA cohort to 44 patients.ConclusionsOur data demonstrate that loss of negative regulation of the inhibitory G-protein signalling causes HR perturbations in Gnb5−/−mice, an effect mainly driven by impaired parasympathetic activity. We anticipate that unravelling the mechanism of Gnb5signalling in the autonomic control of the heart will pave the way for future drug screening.

BackgroundIntellectual disability syndromes (IDSs) with or without developmental delays affect up to 3% of the world population. We sought to clinically and genetically characterise a novel IDS segregating in five unrelated consanguineous families.MethodsClinical analyses were performed for eight patients with intellectual disability (ID). Whole-exome sequencing for selected participants followed by Sanger sequencing for all available family members was completed. Identity-by-descent (IBD) mapping was carried out for patients in two Egyptian families harbouring an identical variant. RNA was extracted from blood cells of Turkish participants, followed by cDNA synthesis and real-time PCR for TTC5.ResultsPhenotype comparisons of patients revealed shared clinical features of moderate-to-severe ID, corpus callosum agenesis, mild ventriculomegaly, simplified gyral pattern, cerebral atrophy, delayed motor and verbal milestones and hypotonia, presenting with an IDS. Four novel homozygous variants in TTC5: c.629A>G;p.(Tyr210Cys), c.692C>T;p.(Ala231Val), c.787C>T;p.(Arg263Ter) and c.1883C>T;p.(Arg395Ter) were identified in the eight patients from participating families. IBD mapping revealed that c.787C>T;p.(Arg263Ter) is a founder variant in Egypt. Missense variants c.629A>G;p.(Tyr210Cys) and c.692C>T;p.(Ala231Val) disrupt highly conserved residues of TTC5 within the fifth and sixth tetratricopeptide repeat motifs which are required for p300 interaction, while the nonsense variants are predicted to decrease TTC5expression. Functional analysis of variant c.1883C>T;p.(Arg395Ter) showed reduced TTC5transcript levels in accordance with nonsense-mediated decay.ConclusionCombining our clinical and molecular data with a recent case report, we identify the core and variable clinical features associated with TTC5loss-of-function variants and reveal the requirement for TTC5 in human brain development and health.

The PIDDosome is a multiprotein complex, composed by the p53-induced death domain protein 1 (PIDD1), the bipartite linker protein CRADD (also known as RAIDD) and the proform of caspase-2 that induces apoptosis in response to DNA damage. In the recent years, biallelic pathogenic variants in CRADDhave been associated with a neurodevelopmental disorder (MRT34; MIM 614499) characterized by pachygyria with a predominant anterior gradient, megalencephaly, epilepsy and intellectual disability. More recently, biallelic pathogenic variants in PIDD1have been described in a few families with apparently nonsydnromic intellectual disability. Here, we aim to delineate the genetic and radio-clinical features of PIDD1-related disorder. Exome sequencing was carried out in six consanguineous families. Thorough clinical and neuroradiological evaluation was performed for all the affected individuals as well as reviewing all the data from previously reported cases. We identified five distinct novelhomozygous variants (c.2584C>T p.(Arg862Trp), c.1340G>A p.(Trp447*), c.2116_2120del p.(Val706Hisfs*30), c.1564_1565delCA p.(Gln522fs*44), and c.1804_1805del p.(Gly602fs*26) in eleven subjects displaying intellectual disability, behaviorial and psychiatric features, and a typical anterior-predominant pachygyria, remarkably resembling the CRADD-related neuroimaging pattern. In summary, we outlin`e the phenotypic and molecular spectrum of PIDD1 biallelic variants supporting the evidence that the PIDD1/CRADD/caspase-2 signaling is crucial for normal gyration of the developing human neocortex as well as cognition and behavior.

De novo mutations arising on the paternal chromosome make the largest known contribution to autism risk, and correlate with paternal age at the time of conception. The recurrence risk for autism spectrum disorders is substantial, leading many families to decline future pregnancies, but the potential impact of assessing parental gonadal mosaicism has not been considered. We measured sperm mosaicism using deep-whole-genome sequencing, for variants both present in an offspring and evident only in father’s sperm, and identified single-nucleotide, structural and short tandem-repeat variants. We found that mosaicism quantification can stratify autism spectrum disorders recurrence risk due to de novo mutations into a vast majority with near 0% recurrence and a small fraction with a substantially higher and quantifiable risk, and we identify novel mosaic variants at risk for transmission to a future offspring. This suggests, therefore, that genetic counseling would benefit from the addition of sperm mosaicism assessment.

BackgroundProtein disulfide isomerase (PDI) proteins are part of the thioredoxin protein superfamily. PDIs are involved in the formation and rearrangement of disulfide bonds between cysteine residues during protein folding in the endoplasmic reticulum and are implicated in stress response pathways.MethodsEight children from four consanguineous families residing in distinct geographies within the Middle East and Central Asia were recruited for study. All probands showed structurally similar microcephaly with lissencephaly (microlissencephaly) brain malformations. DNA samples from each family underwent whole exome sequencing, assessment for repeat expansions and confirmatory segregation analysis.ResultsAn identical homozygous variant in TMX2(c.500G>A), encoding thioredoxin-related transmembrane protein 2, segregated with disease in all four families. This variant changed the last coding base of exon 6, and impacted mRNA stability. All patients presented with microlissencephaly, global developmental delay, intellectual disability and epilepsy. While TMX2is an activator of cellular C9ORF72repeat expansion toxicity, patients showed no evidence of C9ORF72repeat expansions.ConclusionThe TMX2c.500G>A allele associates with recessive microlissencephaly, and patients show no evidence of C9ORF72expansions. TMX2is the first PDI implicated in a recessive disease, suggesting a protein isomerisation defect in microlissencephaly.

BackgroundPutative nucleotidyltransferase MAB21L1 is a member of an evolutionarily well-conserved family of the male abnormal 21 (MAB21)-like proteins. Little is known about the biochemical function of the protein; however, prior studies have shown essential roles for several aspects of embryonic development including the eye, midbrain, neural tube and reproductive organs.ObjectiveA homozygous truncating variant in MAB21L1has recently been described in a male affected by intellectual disability, scrotal agenesis, ophthalmological anomalies, cerebellar hypoplasia and facial dysmorphism. We employed a combination of exome sequencing and homozygosity mapping to identify the underlying genetic cause in subjects with similar phenotypic features descending from five unrelated consanguineous families.ResultsWe identified four homozygous MAB21L1loss of function variants (p.Glu281fs*20, p.Arg287Glufs*14 p.Tyr280* and p.Ser93Serfs*48) and one missense variant (p.Gln233Pro) in 10 affected individuals from 5 consanguineous families with a distinctive autosomal recessive neurodevelopmental syndrome. Cardinal features of this syndrome include a characteristic facial gestalt, corneal dystrophy, hairy nipples, underdeveloped labioscrotal folds and scrotum/scrotal agenesis as well as cerebellar hypoplasia with ataxia and variable microcephaly.ConclusionThis report defines an ultrarare but clinically recognisable Cerebello-Oculo-Facio-Genital syndrome associated with recessive MAB21L1variants. Additionally, our findings further support the critical role of MAB21L1 in cerebellum, lens, genitalia and as craniofacial morphogenesis.

Neuronal migration defects, including pachygyria, are among the most severe developmental brain defects in humans. Here, we identify biallelic truncating mutations in CTNNA2, encoding αN-catenin, in patients with a distinct recessive form of pachygyria. CTNNA2was expressed in human cerebral cortex, and its loss in neurons led to defects in neurite stability and migration. The αN-catenin paralog, αE-catenin, acts as a switch regulating the balance between β-catenin and Arp2/3 actin filament activities1. Loss of αN-catenin did not affect β-catenin signaling, but recombinant αN-catenin interacted with purified actin and repressed ARP2/3 actin-branching activity. The actin-binding domain of αN-catenin or ARP2/3 inhibitors rescued the neuronal phenotype associated with CTNNA2loss, suggesting ARP2/3 de-repression as a potential disease mechanism. Our findings identify CTNNA2as the first catenin family member with biallelic mutations in humans, causing a new pachygyria syndrome linked to actin regulation, and uncover a key factor involved in ARP2/3 repression in neurons.

BackgroundTransport protein particle (TRAPP) is a multisubunit complex that regulates membrane trafficking through the Golgi apparatus. The clinical phenotype associated with mutations in various TRAPP subunits has allowed elucidation of their functions in specific tissues. The role of some subunits in human disease, however, has not been fully established, and their functions remain uncertain.ObjectiveWe aimed to expand the range of neurodevelopmental disorders associated with mutations in TRAPP subunits by exome sequencing of consanguineous families.MethodsLinkage and homozygosity mapping and candidate gene analysis were used to identify homozygous mutations in families. Patient fibroblasts were used to study splicing defect and zebrafish to model the disease.ResultsWe identified six individuals from three unrelated families with a founder homozygous splice mutation in TRAPPC6B, encoding a core subunit of the complex TRAPP I. Patients manifested a neurodevelopmental disorder characterised by microcephaly, epilepsy and autistic features, and showed splicing defect. Zebrafish trappc6bmorphants replicated the human phenotype, displaying decreased head size and neuronal hyperexcitability, leading to a lower seizure threshold.ConclusionThis study provides clinical and functional evidence of the role of TRAPPC6Bin brain development and function.

Deadenylases are best known for degrading the poly(A) tail during mRNA decay. The deadenylase family has expanded throughout evolution and, in mammals, consists of 12 Mg2+-dependent 3′-end RNases with substrate specificity that is mostly unknown. Pontocerebellar hypoplasia type 7 (PCH7) is a unique recessive syndrome characterized by neurodegeneration and ambiguous genitalia. We studied 12 human families with PCH7, uncovering biallelic, loss-of-function mutations in TOE1, which encodes an unconventional deadenylase. toe1-morphant zebrafish displayed midbrain and hindbrain degeneration, modeling PCH-like structural defects in vivo. Surprisingly, we found that TOE1 associated with small nuclear RNAs (snRNAs) incompletely processed spliceosomal. These pre-snRNAs contained 3′ genome-encoded tails often followed by post-transcriptionally added adenosines. Human cells with reduced levels of TOE1 accumulated 3′-end-extended pre-snRNAs, and the immunoisolated TOE1 complex was sufficient for 3′-end maturation of snRNAs. Our findings identify the cause of a neurodegenerative syndrome linked to snRNA maturation and uncover a key factor involved in the processing of snRNA 3′ ends.

BackgroundMicrocephaly with nephrotic syndrome is a rare co-occurrence, constituting the Galloway-Mowat syndrome (GAMOS), caused by mutations in WDR73(OMIM: 616144). However, not all patients harbour demonstrable WDR73deleterious variants, suggesting that there are other yet unidentified factors contributing to GAMOS aetiology.MethodsAutozygosity mapping and candidate analysis was used to identify deleterious variants in consanguineous families. Analysis of patient fibroblasts was used to study splicing and alterations in cellular function.ResultsIn two consanguineous families with five affected individuals from Turkey with a GAMOS-like presentation, we identified a shared homozygous variant leading to partial exon 4 skipping in nucleoporin, 107-KD(NUP107). The founder mutation was associated with concomitant reduction in NUP107 protein and in the obligate binding partner NUP133 protein, as well as density of nuclear pores in patient cells.ConclusionRecently, NUP107was suggested as a candidate in a family with nephrotic syndrome and developmental delay. Other NUP107-reported cases had isolated renal phenotypes. With the addition of these individuals, we implicate an allele-specific critical role for NUP107in the regulation of brain growth and a GAMOS-like presentation.

Zika virus (ZIKV)-related neuropathology is an important global health concern. Several studies have shown that ZIKV can infect neural stem cells in the developing brain, but infection in the adult brain has not been examined. Two areas in the adult mouse brain contain neural stem cells: the subventricular zone of the anterior forebrain and the subgranular zone of the hippocampus. Here, using 6-week-old mice triply deficient in interferon regulatory factor (IRF) as a model, we show that blood-borne ZIKV administration can lead to pronounced evidence of ZIKV infection in these adult neural stem cells, leading to cell death and reduced proliferation. Our data therefore suggest that adult as well as fetal neural stem cells are vulnerable to ZIKV neuropathology. Thus, although ZIKV is considered a transient infection in adult humans without marked long-term effects, there may in fact be consequences of exposure in the adult brain.

The Greater Middle East (GME) has been a central hub of human migration and population admixture. The tradition of consanguinity, variably practiced in the Persian Gulf region, North Africa, and Central Asia, has resulted in an elevated burden of recessive disease. Here we generated a whole-exome GME variome from 1,111 unrelated subjects. We detected substantial diversity and admixture in continental and subregional populations, corresponding to several ancient founder populations with little evidence of bottlenecks. Measured consanguinity rates were an order of magnitude above those in other sampled populations, and the GME population exhibited an increased burden of runs of homozygosity (ROHs) but showed no evidence for reduced burden of deleterious variation due to classically theorized 'genetic purging'. Applying this database to unsolved recessive conditions in the GME population reduced the number of potential disease-causing variants by four- to sevenfold. These results show variegated genetic architecture in GME populations and support future human genetic discoveries in Mendelian and population genetics.

Linear nevus sebaceous syndrome (LNSS) is a neurocutaneous disorder caused by somatic gain-of-function mutations in KRASor HRAS. LNSS brains have neurodevelopmental defects, including cerebral defects and epilepsy; however, its pathological mechanism and potentials for treatment are largely unclear. We show that introduction of KRASG12Vin the developing mouse cortex results in subcortical nodular heterotopia and enhanced excitability, recapitulating major pathological manifestations of LNSS. Moreover, we show that decreased firing frequency of inhibitory neurons without KRASG12Vexpression leads to disrupted excitation and inhibition balance. Transcriptional profiling after destabilization domain-mediated clearance of KRASG12Vin human neural progenitors and differentiating neurons identifies reversible functional networks underlying LNSS. Neurons expressing KRASG12Vshow molecular changes associated with delayed neuronal maturation, most of which are restored by KRASG12Vclearance. These findings provide insights into the molecular networks underlying the reversibility of some of the neuropathologies observed in LNSS caused by dysregulation of the RAS pathway.

In human FMCD tissue, a small fraction of pS6+neurons are enriched for somatic activating mutations of the PI3K-AKT-mTOR pathway. Sparse electroporation of the AKT3mutation into the developing mouse brain causes a reelin-dependent, non–cell autonomous disruption of neuronal migration, leading to impaired cortical lamination and seizure-like epileptiform EEG activity.

The dysfunction of the primary cilium, a complex, evolutionarily conserved, organelle playing an important role in sensing and transducing cell signals, is the unifying pathogenetic mechanism of a growing number of diseases collectively termed “ciliopathies”, typically characterized by multiorgan involvement. Developmental defects of the central nervous system (CNS) characterize a subset of ciliopathies showing clinical and genetic overlap, such as Joubert syndrome (JS) and Meckel syndrome (MS). Although several knock-out mice lacking a variety of ciliary proteins have shown the importance of primary cilia in the development of the brain and CNS-derived structures, developmental in vitrostudies, extremely useful to unravel the role of primary cilia along the course of neural differentiation, are still missing.

Primary cilia are generally solitary organelles that emanate from the surface of almost all vertebrate cell types. Until recently, details regarding the function of these structures were lacking; however, extensive evidence now suggests that primary cilia have critical roles in sensing the extracellular environment, and in coordinating developmental and homeostatic signalling pathways. Furthermore, disruption of these functions seems to underlie a diverse spectrum of disorders, known as primary ciliopathies. These disorders are characterized by wide-ranging clinical and genetic heterogeneity, but with substantial overlap among distinct conditions. Indeed, ciliopathies are associated with a large variety of manifestations that often include distinctive neurological findings. Herein, we review neurological features associated with primary ciliopathies, highlight genotype–phenotype correlations, and discuss potential mechanisms underlying these findings.

Autosomal-recessive loss of the NSUN2gene has been identified as a causative link to intellectual disability disorders in humans. NSun2 is an RNA methyltransferase modifying cytosine-5 in transfer RNAs (tRNAs), yet the identification of cytosine methylation in other RNA species has been hampered by the lack of sensitive and reliable molecular techniques. Here, we describe miCLIP as an additional approach for identifying RNA methylation sites in transcriptomes. miCLIP is a customized version of the individual-nucleotide-resolution crosslinking and immunoprecipitation (iCLIP) method. We confirm site-specific methylation in tRNAs and additional messenger and noncoding RNAs (ncRNAs). Among these, vault ncRNAs contained six NSun2-methylated cytosines, three of which were confirmed by RNA bisulfite sequencing. Using patient cells lacking the NSun2 protein, we further show that loss of cytosine-5 methylation in vault RNAs causes aberrant processing into Argonaute-associated small RNA fragments that can function as microRNAs. Thus, impaired processing of vault ncRNA may contribute to the etiology of NSun2-deficiency human disorders.

Combining human genomics and molecular biology, recent studies have made pivotal progress toward understanding the cause of hemimegalencephaly (HME) and other cerebral megalencephaly syndromes. The present article highlights recent advances of the genetic cause of these conditions, and considers the role of somatic postzygotic genetic lesions in brain maldevelopment.

Ciliopathies are genetic disorders caused by defects of primary ciliary structure and/or function and are characterized by pleiotropic clinical features. The ciliopathies include several partially overlapping syndromes such as Joubert syndrome, Bardet–Biedl syndrome and Meckel–Gruber syndrome, all of which have pronounced neurodevelopmental features. Here we focus on potential roles of cilia in central nervous system function, to explore how impairments may cause brain malformation and neurodevelopmental disease.

Ciliopathies are an expanding group of rare conditions characterized by multiorgan involvement, that are caused by mutations in genes encoding for proteins of the primary cilium or its apparatus. Among these genes, CEP290bears an intriguing allelic spectrum, being commonly mutated in Joubert syndrome and related disorders JSRD, Meckel syndrome MKS, SeniorLoken syndrome and isolated Leber congenital amaurosis LCA. Although these conditions are recessively inherited, in a subset of patients only one CEP290mutation could be detected. To assess whether genomic rearrangements involving the CEP290gene could represent a possible mutational mechanism in these cases, exon dosage analysis on genomic DNA was performed in two groups of CEP290heterozygous patients, including five JSRDMKS cases and four LCA, respectively. In one JSRD patient, we identified a large heterozygous deletion encompassing CEP290Cterminus that resulted in marked reduction of mRNA expression. No copy number alterations were identified in the remaining probands. The present work expands the CEP290genotypic spectrum to include multiexon deletions. Although this mechanism does not appear to be frequent, screening for genomic rearrangements should be considered in patients in whom a single CEP290mutated allele was identified. © 2009 WileyLiss, Inc.

We identified two female siblings, derived from healthy first cousin parents, with congenital unilateral cerebral ventriculomegaly detected prenatally. Patient 1 underwent ventriculoperitoneal shunt operation at 1 week old, while Patient 2 was followed without surgical intervention. Both patients presented with mild developmental delay and hemiparesis contralateral to the involved hemisphere. Focal seizures were observed in Patient 1, whose neuroimaging revealed posterior insular polymicrogyria in the normal sized ventricle hemisphere and retrocerebellar cyst. Both siblings displayed near absence of white matter with marked thinning of the overlying cortex in the affected hemisphere and very thin corpus callosum. Investigations revealed no other system involvement and karyotyping was normal. Normal TORCH screening in subsequent pregnancies, normal parental coagulation profile and undetectable maternal autoantibodies suggested against the possible role of extrinsic factors as an etiological factor for unilateral ventriculomegaly. Parents had normal brain imaging findings. We suggest delineation of a distinct developmental brain defect, most likely of autosomal recessive inheritance. © 2009 Wiley‐Liss, Inc.

Zika virus (ZIKV) and dengue virus (DENV) are arthropod-borne pathogenic flaviviruses that co-circulate in many countries. To understand some of the pressures that influence ZIKV evolution, we mimic the natural transmission cycle by repeating serial passaging of ZIKV through cultured mosquito cells and either DENV-naive or DENV-immune mice. Compared with wild-type ZIKV, the strains passaged under both conditions exhibit increased pathogenesis in DENV-immune mice. Application of reverse genetics identifies an isoleucine-to-valine mutation (I39V) in the NS2B proteins of both passaged strains that confers enhanced fitness and escape from pre-existing DENV immunity. Introduction of I39V or I39T, a naturally occurring homologous mutation detected in recent ZIKV isolates, increases the replication of wild-type ZIKV in human neuronal precursor cells and laboratory-raised mosquitoes. Our data indicate that ZIKV strains with enhanced transmissibility and pathogenicity can emerge in DENV-naive or -immune settings, and that NS2B-I39 mutants may represent ZIKV variants of interest.

Two siblings from a consanguineous Egyptian marriage showed an identical phenotype of cortical lissencephaly with cerebellar hypoplasia, severe epilepsy, and mental retardation. Examination of karyotype revealed 46, t(7;12)(q22;p13)mat (7;12)(q22;p13)pat in both affected children, suggesting a homozygous reciprocal balanced translocation. Each healthy parent was a carrier of the balanced translocation in the heterozygous state, suggesting homozygous disruption of a gene involved in brain development. There were early spontaneous abortions in this family, as would be expected from transmission of an unbalanced chromosome. A disruption of RELN at 7q22.1 with absence of encoded protein was identified. This is the first demonstration that such rare homozygous translocations can be used to identify recessive disease gene mutations. © 2007 Wiley‐Liss, Inc.

Jouberts syndrome–related disorders are a group of recessively inherited conditions showing cerebellar vermis hypoplasia and the molar tooth sign of the midbrain–hindbrain junction. Recent analyses have suggested at least three loci, JBTS1 (9q34.3), ‐2 (11p11.2‐q12.3), and ‐3 (6q23), but the phenotypic spectrum associated with each locus has not been delineated. In addition, deletions of the NPHP1gene, usually responsible for isolated juvenile nephronophthisis, are occasionally encountered among Jouberts syndrome–related disorder patients. Here, we describe four novel families showing evidence of linkage to two of these loci, provide a 3.6Mb refinement of the JBTS2 locus, and perform a detailed comparison of all linked families identified so far, to define the clinical and radiographical hallmarks for each genetic condition. We find that JBTS1 and ‐3 primarily show features restricted to the central nervous system, with JBTS1 showing largely pure cerebellar and midbrain–hindbrain junction involvement, and JBTS3 displaying cerebellar, midbrain–hindbrain junction, and cerebral cortical features, most notably polymicrogyria. Conversely, JBTS2 is associated with multiorgan involvement of kidney, retina, and liver, in addition to the central nervous system features, and results in extreme phenotypic variability. This provides a useful framework for genetic testing strategies and prediction of which patients are most likely to experience development of systemic complications. Ann Neurol 2005;57:513–519