RESUMEN
The adult hippocampus generates new granule cells (aGCs) with functional capabilities that convey unique forms of plasticity to the preexisting circuits. While early differentiation of adult radial glia-like cells (RGLs) has been studied extensively, the molecular mechanisms guiding the maturation of postmitotic neurons remain unknown. Here, we used a precise birthdating strategy to study aGC differentiation using single-nuclei RNA sequencing. Transcriptional profiling revealed a continuous trajectory from RGLs to mature aGCs, with multiple immature stages bearing increasing levels of effector genes supporting growth, excitability, and synaptogenesis. Analysis of differential gene expression, pseudo-time trajectory, and transcription factors (TFs) revealed critical transitions defining four cellular states: quiescent RGLs, proliferative progenitors, immature aGCs, and mature aGCs. Becoming mature aGCs involved a transcriptional switch that shuts down pathways promoting cell growth, such SoxC TFs, to activate programs that likely control neuronal homeostasis. aGCs overexpressing Sox4 or Sox11 remained immature. Our results unveil precise molecular mechanisms driving adult RGLs through the pathway of neuronal differentiation.
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Diferenciación Celular , Hipocampo , Neurogénesis , Neuronas , Factores de Transcripción SOXC , Animales , Hipocampo/metabolismo , Hipocampo/citología , Neuronas/metabolismo , Neuronas/citología , Factores de Transcripción SOXC/metabolismo , Factores de Transcripción SOXC/genética , Diferenciación Celular/genética , Neurogénesis/genética , Ratones , Transcripción Genética , Perfilación de la Expresión Génica , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Células Ependimogliales/metabolismo , Células Ependimogliales/citologíaRESUMEN
The mammalian cerebral cortex comprises a complex neuronal network that maintains a delicate balance between excitatory neurons and inhibitory interneurons. Previous studies, including our own research, have shown that specific interneuron subtypes are closely associated with particular pyramidal neuron types, forming stereotyped local inhibitory microcircuits. However, the developmental processes that establish these precise networks are not well understood. Here we show that pyramidal neuron types are instrumental in driving the terminal differentiation and maintaining the survival of specific associated interneuron subtypes. In a wild-type cortex, the relative abundance of different interneuron subtypes aligns precisely with the pyramidal neuron types to which they synaptically target. In Fezf2 mutant cortex, characterized by the absence of layer 5 pyramidal tract neurons and an expansion of layer 6 intratelencephalic neurons, we observed a corresponding decrease in associated layer 5b interneurons and an increase in layer 6 subtypes. Interestingly, these shifts in composition are achieved through mechanisms specific to different interneuron types. While SST interneurons adjust their abundance to the change in pyramidal neuron prevalence through the regulation of programmed cell death, parvalbumin interneurons alter their identity. These findings illustrate two key strategies by which the dynamic interplay between pyramidal neurons and interneurons allows local microcircuits to be sculpted precisely. These insights underscore the precise roles of extrinsic signals from pyramidal cells in the establishment of interneuron diversity and their subsequent integration into local cortical microcircuits.
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Interindividual genetic variation affects the susceptibility to and progression of many diseases1,2. However, efforts to study how individual human brains differ in normal development and disease phenotypes are limited by the paucity of faithful cellular human models, and the difficulty of scaling current systems to represent multiple people. Here we present human brain Chimeroids, a highly reproducible, multidonor human brain cortical organoid model generated by the co-development of cells from a panel of individual donors in a single organoid. By reaggregating cells from multiple single-donor organoids at the neural stem cell or neural progenitor cell stage, we generate Chimeroids in which each donor produces all cell lineages of the cerebral cortex, even when using pluripotent stem cell lines with notable growth biases. We used Chimeroids to investigate interindividual variation in the susceptibility to neurotoxic triggers that exhibit high clinical phenotypic variability: ethanol and the antiepileptic drug valproic acid. Individual donors varied in both the penetrance of the effect on target cell types, and the molecular phenotype within each affected cell type. Our results suggest that human genetic background may be an important mediator of neurotoxin susceptibility and introduce Chimeroids as a scalable system for high-throughput investigation of interindividual variation in processes of brain development and disease.
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Corteza Cerebral , Quimera , Predisposición Genética a la Enfermedad , Neurotoxinas , Organoides , Femenino , Humanos , Masculino , Linaje de la Célula/efectos de los fármacos , Corteza Cerebral/citología , Corteza Cerebral/efectos de los fármacos , Corteza Cerebral/metabolismo , Quimera/genética , Etanol/efectos adversos , Etanol/toxicidad , Variación Genética , Células-Madre Neurales/citología , Células-Madre Neurales/efectos de los fármacos , Células-Madre Neurales/metabolismo , Neurotoxinas/toxicidad , Organoides/citología , Organoides/efectos de los fármacos , Organoides/metabolismo , Fenotipo , Células Madre Pluripotentes/citología , Células Madre Pluripotentes/efectos de los fármacos , Células Madre Pluripotentes/metabolismo , Donantes de Tejidos , Ácido Valproico/efectos adversos , Ácido Valproico/toxicidad , Predisposición Genética a la Enfermedad/genéticaRESUMEN
Since the beautiful images of Santiago Ramón y Cajal provided a first glimpse into the immense diversity and complexity of cell types found in the cerebral cortex, neuroscience has been challenged and inspired to understand how these diverse cells are generated and how they interact with each other to orchestrate the development of this remarkable tissue. Some fundamental questions drive the field's quest to understand cortical development: what are the mechanistic principles that govern the emergence of neuronal diversity? How do extrinsic and intrinsic signals integrate with physical forces and activity to shape cell identity? How do the diverse populations of neurons and glia influence each other during development to guarantee proper integration and function? The advent of powerful new technologies to profile and perturb cortical development at unprecedented resolution and across a variety of modalities has offered a new opportunity to integrate past knowledge with brand new data. Here, we review some of this progress using cortical excitatory projection neurons as a system to draw out general principles of cell diversification and the role of cell-cell interactions during cortical development.
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Corteza Cerebral , Neuronas , Corteza Cerebral/citología , Corteza Cerebral/fisiología , Corteza Cerebral/crecimiento & desarrollo , Animales , Neuronas/fisiología , Neuronas/citología , Humanos , Neurogénesis/fisiologíaRESUMEN
Motivation: Single-cell RNA sequencing (scRNAseq) has transformed our ability to explore biological systems. Nevertheless, proficient expertise is essential for handling and interpreting the data. Results: In this article, we present scX, an R package built on the Shiny framework that streamlines the analysis, exploration, and visualization of single-cell experiments. With an interactive graphic interface, implemented as a web application, scX provides easy access to key scRNAseq analyses, including marker identification, gene expression profiling, and differential gene expression analysis. Additionally, scX seamlessly integrates with commonly used single-cell Seurat and SingleCellExperiment R objects, resulting in efficient processing and visualization of varied datasets. Overall, scX serves as a valuable and user-friendly tool for effortless exploration and sharing of single-cell data, simplifying some of the complexities inherent in scRNAseq analysis. Availability and implementation: Source code can be downloaded from https://github.com/chernolabs/scX. A docker image is available from dockerhub as chernolabs/scx.
RESUMEN
The adult hippocampus generates new granule cells (aGCs) that exhibit distinct functional capabilities along development, conveying a unique form of plasticity to the preexisting circuits. While early differentiation of adult radial glia-like neural stem cells (RGL) has been studied extensively, the molecular mechanisms guiding the maturation of postmitotic neurons remain unknown. Here, we used a precise birthdating strategy to follow newborn aGCs along differentiation using single-nuclei RNA sequencing (snRNA-seq). Transcriptional profiling revealed a continuous trajectory from RGLs to mature aGCs, with multiple sequential immature stages bearing increasing levels of effector genes supporting growth, excitability and synaptogenesis. Remarkably, four discrete cellular states were defined by the expression of distinct sets of transcription factors (TFs): quiescent neural stem cells, proliferative progenitors, postmitotic immature aGCs, and mature aGCs. The transition from immature to mature aCGs involved a transcriptional switch that shutdown molecular cascades promoting cell growth, such as the SoxC family of TFs, to activate programs controlling neuronal homeostasis. Indeed, aGCs overexpressing Sox4 or Sox11 remained stalled at the immature state. Our results unveil precise molecular mechanisms driving adult neural stem cells through the pathway of neuronal differentiation.
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Single-cell RNA sequencing (scRNA-seq) has transformed our ability to explore biological systems. Nevertheless, proficient expertise is essential for handling and interpreting the data. In this paper, we present scX, an R package built on the Shiny framework that streamlines the analysis, exploration, and visualization of single-cell experiments. With an interactive graphic interface, implemented as a web application, scX provides easy access to key scRNAseq analyses, including marker identification, gene expression profiling, and differential gene expression analysis. Additionally, scX seamlessly integrates with commonly used single-cell Seurat and Single-CellExperiment R objects, resulting in efficient processing and visualization of varied datasets. Overall, scX serves as a valuable and user-friendly tool for effortless exploration and sharing of single-cell data, simplifying some of the complexities inherent in scRNAseq analysis.
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Cerebellar ataxia, neuropathy and vestibular areflexia syndrome (CANVAS) is an autosomal recessive neurodegenerative disease, usually caused by biallelic AAGGG repeat expansions in RFC1. In this study, we leveraged whole genome sequencing data from nearly 10 000 individuals recruited within the Genomics England sequencing project to investigate the normal and pathogenic variation of the RFC1 repeat. We identified three novel repeat motifs, AGGGC (n = 6 from five families), AAGGC (n = 2 from one family) and AGAGG (n = 1), associated with CANVAS in the homozygous or compound heterozygous state with the common pathogenic AAGGG expansion. While AAAAG, AAAGGG and AAGAG expansions appear to be benign, we revealed a pathogenic role for large AAAGG repeat configuration expansions (n = 5). Long-read sequencing was used to characterize the entire repeat sequence, and six patients exhibited a pure AGGGC expansion, while the other patients presented complex motifs with AAGGG or AAAGG interruptions. All pathogenic motifs appeared to have arisen from a common haplotype and were predicted to form highly stable G quadruplexes, which have previously been demonstrated to affect gene transcription in other conditions. The assessment of these novel configurations is warranted in CANVAS patients with negative or inconclusive genetic testing. Particular attention should be paid to carriers of compound AAGGG/AAAGG expansions when the AAAGG motif is very large (>500 repeats) or the AAGGG motif is interrupted. Accurate sizing and full sequencing of the satellite repeat with long-read sequencing is recommended in clinically selected cases to enable accurate molecular diagnosis and counsel patients and their families.
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Ataxia Cerebelosa , Enfermedades del Sistema Nervioso Periférico , Síndrome , Enfermedades Vestibulares , Humanos , Vestibulopatía Bilateral , Ataxia Cerebelosa/genética , Ataxia Cerebelosa/diagnóstico , Enfermedades Neurodegenerativas , Enfermedades del Sistema Nervioso Periférico/diagnóstico , Enfermedades del Sistema Nervioso Periférico/genética , Enfermedades Vestibulares/diagnóstico , Enfermedades Vestibulares/genéticaRESUMEN
BACKGROUND: Leukodystrophy with vanishing white matter (LVWM) is an autosomal recessive disease with typical pediatric-onset caused by mutations in one of the five EIF2B genes. Adult-onset (AO) cases are rare. METHODS: In this observational study, we reviewed clinical and laboratory information of the patients with AO-LVWM assessed at two referral centers in Italy and Portugal from Jan-2007 to Dec-2019. RESULTS: We identified 18 patients (13 females) with AO-LVWM caused by EIF2B5 or EIF2B3 mutations. Age of neurological onset ranged from 16 to 60 years, with follow-ups occurring from 2 to 37 years. Crucial symptoms were cognitive and motor decline. In three patients, stroke-like events were the first manifestation; in another, bladder dysfunction remained the main complaint across decades. Brain MRI showed white matter (WM) rarefaction in all cases, except two. Diffusion-weighted imaging documented focal hyperintensity in the acute stage of stroke-like events. 1H-spectroscopy primarily showed N-acetyl-aspartate reduction; 18fluorodeoxyglucose-PET revealed predominant frontoparietal hypometabolism; evoked potential studies demonstrated normal-to-reduced amplitudes; neuro-ophthalmological assessment showed neuroretinal thinning, and b-wave reduction on full-field electroretinogram. Interestingly, we found an additional patient with LVWM-compatible phenotype and monoallelic variants in two distinct eIF2B genes, EIF2B1 and EIF2B2. CONCLUSIONS: AO-LVWM presents varying clinical manifestations at onset, including stroke-like events. WM rarefaction is the most consistent diagnostic clue even in the latest onset cases. Spectroscopy and electrophysiological features are compatible with axon, rather than myelin, damage. Cerebral glucose metabolic abnormalities and retinal alterations can be present. LVWM might also be caused by a digenic inheritance affecting the eIF2B complex.
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Enfermedades Desmielinizantes , Leucoencefalopatías , Enfermedades por Almacenamiento Lisosomal , Enfermedades Neurodegenerativas , Accidente Cerebrovascular , Sustancia Blanca , Adolescente , Adulto , Femenino , Humanos , Masculino , Persona de Mediana Edad , Adulto Joven , Factor 2B Eucariótico de Iniciación/genética , Leucoencefalopatías/diagnóstico por imagen , Leucoencefalopatías/genética , Imagen por Resonancia Magnética , Mutación/genética , Estudios Observacionales como Asunto , Sustancia Blanca/diagnóstico por imagenRESUMEN
BACKGROUND AND OBJECTIVES: Spinocerebellar ataxias (SCAs) are autosomal dominant disorders with extensive clinical and genetic heterogeneity. We recently identified a form of SCA transmitted with a digenic pattern of inheritance caused by the concomitant presence of an intermediate-length expansion in TATA-box binding protein gene (TBP40-46 ) and a heterozygous pathogenic variant in the Stip1-homologous and U-Box containing protein 1 gene (STUB1). This SCATBP/STUB1 represents the first example of a cerebellar disorder in which digenic inheritance has been identified. OBJECTIVES: We studied a large cohort of patients with SCATBP/STUB1 with the aim of describing specific clinical and neuroimaging features of this distinctive genotype. METHODS: In this observational study, we recruited 65 affected and unaffected family members from 21 SCATBP/STUB1 families and from eight families with monogenic SCA17. Their characteristics and phenotypes were compared with those of 33 age-matched controls. RESULTS: SCATBP/STUB1 patients had multi-domain dementia with a more severe impairment in respect to patient carrying only fully expanded SCA17 alleles. Cerebellar volume and thickness of cerebellar cortex were reduced in SCATBP/STUB1 compared with SCA17 patients (P = 0.03; P = 0.008). Basal ganglia volumes were reduced in both patient groups, as compared with controls, whereas brainstem volumes were significantly reduced in SCATBP/STUB1 , but not in SCA17 patients. CONCLUSIONS: The identification of the complex SCATBP/STUB1 phenotype may impact on diagnosis and genetic counseling in the families with both hereditary and sporadic ataxia. The independent segregation of TBP and STUB1 alleles needs to be considered for recurrence risk and predictive genetic tests. © 2023 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.
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Ataxia , Demencia , Ataxias Espinocerebelosas , Humanos , Ataxia/genética , Demencia/genética , Genotipo , Fenotipo , Ataxias Espinocerebelosas/diagnóstico por imagen , Ataxias Espinocerebelosas/genética , Ataxias Espinocerebelosas/metabolismo , Proteína de Unión a TATA-Box/genética , Proteína de Unión a TATA-Box/metabolismo , Expansión de Repetición de Trinucleótido , Ubiquitina-Proteína Ligasas/genéticaRESUMEN
We generated an iPSC line from a patient with spastic paraplegia type 10 (SPG10) carrying the novel missense variant c.50G > A (p.R17Q) in the N-terminal motor domain of the kinesin family member 5A (KIF5A) gene. This patient-derived in vitro cell model will help to investigate the role of different KIF5A mutations in inducing neurodegeneration in spastic paraplegia and in other KIF5A-related disorders, including Charcot-Marie-Tooth type 2 (CMT2) and amyotrophic lateral sclerosis (ALS).
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Células Madre Pluripotentes Inducidas , Paraplejía Espástica Hereditaria , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , Paraplejía Espástica Hereditaria/genética , Cinesinas/genética , Mutación/genética , ParaplejíaRESUMEN
INTRODUCTION: Biallelic intronic AAGGG repeat expansion in the replication factor C subunit 1 (RFC1) gene was recently identified in two/third of patients with cerebellar ataxia, sensory neuropathy, and bilateral vestibular areflexia syndrome (CANVAS). The phenotypic spectrum has expanded since (i.e., parkinsonism, motor neuron involvement, cognitive decline); no behavioral symptoms have been reported yet. CASE REPORT: We report an Italian family that met the diagnostic criteria for CANVAS, and RFC1-expansion was detected in five of seven. All the affected members presented behavioral-psychiatric symptoms (anxiety, panic attacks, alcohol abuse) before the multisystemic RFC1-expansion manifestation. The disease course was progressive, with ataxia and behavioral-cognitive aspects as the most disabling symptoms. CONCLUSION: These behavioral-cognitive observations may broaden the RFC1-expansion phenotypic spectrum and highlight the importance of investigating the whole non-motor symptoms in ataxic patients.
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Vestibulopatía Bilateral , Ataxia Cerebelosa , Enfermedades Vestibulares , Ataxia , Vestibulopatía Bilateral/diagnóstico , Ataxia Cerebelosa/diagnóstico , Humanos , Reflejo AnormalRESUMEN
PURPOSE: This study aimed to unravel the genetic factors underlying missing heritability in spinocerebellar ataxia type 17 (SCA17) caused by polyglutamine-encoding CAG/CAA repeat expansions in the TBP gene. Alleles with >49 CAG/CAA repeats are fully penetrant. Most patients, however, carry intermediate TBP41-49 alleles that show incomplete penetrance. METHODS: Using next-generation sequencing approaches, we investigated 40 SCA17/TBP41-54 index patients, their affected (n = 55) and unaffected (n = 51) relatives, and a cohort of patients with ataxia (n = 292). RESULTS: All except 1 (30/31) of the index cases with TBP41-46 alleles carried a heterozygous pathogenic variant in the STUB1 gene associated with spinocerebellar ataxias SCAR16 (autosomal recessive) and SCA48 (autosomal dominant). No STUB1 variant was found in patients carrying TBP47-54 alleles. TBP41-46 expansions and STUB1 variants cosegregate in all affected family members, whereas the presence of either TBP41-46 expansions or STUB1 variants individually was never associated with the disease. CONCLUSION: Our data reveal an unexpected genetic interaction between STUB1 and TBP in the pathogenesis of SCA17 and raise questions on the existence of SCA48 as a monogenic disease with crucial implications for diagnosis and counseling. They provide a convincing explanation for the incomplete penetrance of intermediate TBP alleles and demonstrate a dual inheritance pattern for SCA17, which is a monogenic dominant disorder for TBP≥47 alleles and a digenic TBP/STUB1 disease (SCA17-DI) for intermediate expansions.
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Péptidos , Ataxias Espinocerebelosas , Proteína de Unión a TATA-Box , Ubiquitina-Proteína Ligasas , Humanos , Penetrancia , Péptidos/genética , Ataxias Espinocerebelosas/genética , Ataxias Espinocerebelosas/patología , Proteína de Unión a TATA-Box/genética , Expansión de Repetición de Trinucleótido/genética , Ubiquitina-Proteína Ligasas/genéticaRESUMEN
Diverse types of glutamatergic pyramidal neurons mediate the myriad processing streams and output channels of the cerebral cortex1,2, yet all derive from neural progenitors of the embryonic dorsal telencephalon3,4. Here we establish genetic strategies and tools for dissecting and fate-mapping subpopulations of pyramidal neurons on the basis of their developmental and molecular programs. We leverage key transcription factors and effector genes to systematically target temporal patterning programs in progenitors and differentiation programs in postmitotic neurons. We generated over a dozen temporally inducible mouse Cre and Flp knock-in driver lines to enable the combinatorial targeting of major progenitor types and projection classes. Combinatorial strategies confer viral access to subsets of pyramidal neurons defined by developmental origin, marker expression, anatomical location and projection targets. These strategies establish an experimental framework for understanding the hierarchical organization and developmental trajectory of subpopulations of pyramidal neurons that assemble cortical processing networks and output channels.
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Corteza Cerebral/citología , Regulación de la Expresión Génica/genética , Ácido Glutámico/metabolismo , Células Piramidales/citología , Células Piramidales/metabolismo , Animales , Linaje de la Célula/genética , Corteza Cerebral/metabolismo , Masculino , Ratones , Células Piramidales/clasificación , Factores de Transcripción/metabolismoRESUMEN
INTRODUCTION: Spastic paraplegia type 46 (SPG46) is a rare autosomal recessive hereditary spastic paraplegia, caused by mutations in the non-lysosomal glucosylceramidase ß2 (GBA2) gene. Worldwide, approximately twenty SPG46 families have been identified so far. CASE REPORT: We describe a compound heterozygous Italian patient carrying a novel (p.Arg879Gln) and a recurrent (p.Arg399 *) GBA2 gene variant. The patient presented unsteady gait at age 2, and progressively manifested spastic-ataxia, scoliosis, mild intellectual decline, and bilateral cataract. DISCUSSION: Clinical manifestations associated with GBA2 gene variants encompass a spectrum of overlapping phenotypes including cerebellar ataxia, spastic paraplegia, and Marinesco-Sjogren-like syndrome. We review previously reported cases of SPG46 and discuss possible genetic differential diagnosis.
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Espasticidad Muscular , Paraplejía Espástica Hereditaria , Preescolar , Glucosilceramidasa/genética , Humanos , Discapacidad Intelectual , Italia , Mutación/genética , Atrofia Óptica , Paraplejía/genética , Linaje , Fenotipo , Paraplejía Espástica Hereditaria/genética , Ataxias EspinocerebelosasRESUMEN
The mammalian cerebral cortex has an unparalleled diversity of cell types, which are generated during development through a series of temporally orchestrated events that are under tight evolutionary constraint and are critical for proper cortical assembly and function1,2. However, the molecular logic that governs the establishment and organization of cortical cell types remains unknown, largely due to the large number of cell classes that undergo dynamic cell-state transitions over extended developmental timelines. Here we generate a comprehensive atlas of the developing mouse neocortex, using single-cell RNA sequencing and single-cell assay for transposase-accessible chromatin using sequencing. We sampled the neocortex every day throughout embryonic corticogenesis and at early postnatal ages, and complemented the sequencing data with a spatial transcriptomics time course. We computationally reconstruct developmental trajectories across the diversity of cortical cell classes, and infer their spatial organization and the gene regulatory programs that accompany their lineage bifurcation decisions and differentiation trajectories. Finally, we demonstrate how this developmental map pinpoints the origin of lineage-specific developmental abnormalities that are linked to aberrant corticogenesis in mutant mice. The data provide a global picture of the regulatory mechanisms that govern cellular diversification in the neocortex.
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Neocórtex/citología , Neurogénesis , Animales , Diferenciación Celular , Proteínas de Unión al ADN/genética , Embrión de Mamíferos , Regulación del Desarrollo de la Expresión Génica , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Neocórtex/embriología , Proteínas del Tejido Nervioso/genética , Análisis de Secuencia de ARN , Análisis de la Célula Individual , TranscriptomaRESUMEN
A wide spectrum of neurodegenerative diseases has been associated with pathogenic variants in the PNPLA6 (patatin-like phospholipase domain-containing protein 6) gene, including spastic paraplegia type 39, Gordon-Holmes, Boucher-Neuhauser, Oliver-Mc Farlane, and Laurence-Moon syndromes. These syndromes present variable and overlapping clinical symptoms, encompassing cerebellar ataxia, hypogonadotropic hypogonadism, chorioretinal dystrophy, spastic paraplegia, muscle wasting, peripheral neuropathy, and cognitive impairment. In the present study, we performed a wide genetic screening in 292 patients presenting with ataxia or spastic paraplegia using a probe-based customized gene panel, covering >200 genes associated with spinocerebellar diseases. We identified six novel and four recurrent PNPLA6 gene variants in eight patients (2.7%). Six patients presented an infantile or juvenile onset (age <18), and two patients had an adult onset. Cerebellar ataxia was observed in seven patients and spastic paraplegia in one patient. Progression of cerebellar symptoms was slow in all patients, who retained ambulation even after a mean disease duration of 15 years. Brain MRI showed cerebellar atrophy in 6/8 patients, more pronounced in superior and dorsal vermis lobules (I to VII). Additional clinical features included hypogonadotropic hypogonadism (5/8), growth hormone deficiency (2/8), peripheral axonal neuropathy (4/8), cognitive impairment (3/8), chorioretinal dystrophy (2/8), and bilateral vestibular areflexia with a reduced visual vestibule-ocular reflex (1/8). In accordance with previous studies, chorioretinal dystrophy was the most frequent presenting symptom in early onset patients, hypogonadotropic hypogonadism in juvenile onset cases, and cerebellar ataxia in adult patients. One patient had an initial clinical presentation compatible with Cerebellar Ataxia with Neuropathy and Vestibular Areflexia Syndrome (CANVAS), but no pathological expansions in the RFC1 gene. In conclusion, patients with PNPLA6 variants present a variable age of onset spanning from infancy to adulthood, and each clinical symptom has an age-dependent manifestation thus requiring a multi-systemic diagnostic approach. The description of patients presenting very late-onset cerebellar ataxia suggests that PNPLA6 genetic screening should also be considered in the diagnostic workout of adult cerebellar ataxia.
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Measurement of the location of molecules in tissues is essential for understanding tissue formation and function. Previously, we developed Slide-seq, a technology that enables transcriptome-wide detection of RNAs with a spatial resolution of 10 µm. Here we report Slide-seqV2, which combines improvements in library generation, bead synthesis and array indexing to reach an RNA capture efficiency ~50% that of single-cell RNA-seq data (~10-fold greater than Slide-seq), approaching the detection efficiency of droplet-based single-cell RNA-seq techniques. First, we leverage the detection efficiency of Slide-seqV2 to identify dendritically localized mRNAs in neurons of the mouse hippocampus. Second, we integrate the spatial information of Slide-seqV2 data with single-cell trajectory analysis tools to characterize the spatiotemporal development of the mouse neocortex, identifying underlying genetic programs that were poorly sampled with Slide-seq. The combination of near-cellular resolution and high transcript detection efficiency makes Slide-seqV2 useful across many experimental contexts.