FBXW11 Activity Regulates Radial Glial Expansion in Human Cerebral Organoids

Cortical organoids from congenital DM1 PSCs reveal MBNL-dependent corticogenesis defects and enable preclinical testing of therapeutic compounds

Cortical organoids are established as a robust human model of CDM-associated neurodevelopmental defects, uncover MBNL-dependent mechanisms underlying early corticogenesis impairment and demonstrate the utility of this platform for translational therapeutic discovery in DM1.

Molecules and Cells

Comparative primate transcriptomics identifies a ZNF90–OVOL2 regulatory axis shaping human neural progenitor cell dynamics

Understanding the behavior of human neural progenitor cells (NPCs) requires comparative characterization of their transcriptomic landscape, particularly through comparisons with closely related primate species. The rhesus macaque represents a key non-hominoid outgroup for such analyses, and several representative transcriptomic datasets are now available. Here, we compare genes enriched in human NPCs with those enriched in rhesus macaque NPCs and found that human NPC-enriched genes are associated with gene programs supporting radial glial identity and proliferative capacity, as well as functions related to sister chromatid segregation. This analysis identified two zinc-finger transcription factors for functional investigation: the ape-specific ZNF90 and the highly conserved OVOL2. By analysing their genome-wide binding, transcriptional output, and cellular effects in cerebral organoids, we uncovered a previously uncharacterized regulatory axis with divergent but partially overlapping effects that converge on apical progenitor (AP) maintenance. Together, our findings support a model in which lineage-specific / evolutionarily young transcription factor become integrated into a conserved developmental gene regulatory network, generating novel NPC dynamics during primate corticogenesis.

Arrayed single-gene perturbations identify drivers of human anterior neural tube closure

Genetic studies of human embryonic morphogenesis are constrained by ethical and practical challenges, restricting insights into developmental mechanisms and disorders. Human pluripotent stem cell (hPSC)-derived organoids provide a powerful alternative for the study of embryonic morphogenesis. However, screening for genetic drivers of morphogenesis in vitro has been infeasible due to organoid variability and the high costs of performing scaled tissue-wide single-gene perturbations. By overcoming both these limitations, we developed a platform that integrates reproducible organoid morphogenesis with uniform single-gene perturbations, enabling high-throughput arrayed CRISPR interference screening in hPSC-derived organoids. To demonstrate the power of this platform, we screened 77 transcription factors in an organoid model of anterior neurulation to identify ZIC2, SOX11, and ZNF521 as essential regulators of neural tube closure. We discovered that ZIC2 and SOX11 are required for closure, while ZNF521 prevents ectopic closure points. Single-cell transcriptomic analysis of perturbed organoids revealed co-regulated gene targets of ZIC2 and SOX11 and an opposing role for ZNF521, suggesting that these transcription factors jointly govern a gene regulatory program driving neural tube closure in the anterior forebrain region. Our single-gene perturbation platform enables high-throughput genetic screening of in vitro models of human embryonic morphogenesis.

A pooled CRISPR-based neuronal activity screen links TMEM50A-dependent MVB function to synaptic integrity and remote memory.

While advances in omics profiling rapidly expand the catalog of genes associated with brain activity in health and disease, functional annotation lags far behind. Here, we establish a high-throughput functional genomics platform that couples the calcium-integrating sensor CaMPARI2 with CRISPRi screening in human iPSC-derived neurons. By converting cumulative neuronal activity into a stable, flow cytometry-readable signal, this approach enables systematic interrogation through pooled screening. Using a focused library of memory-associated genes, we recover known regulators and identify TMEM50A, a previously uncharacterized protein, as an essential regulator of neuronal activity. TMEM50A forms a complex with LEPROTL1 and associates with ESCRT-III machinery on multivesicular bodies (MVBs). TMEM50A loss impairs MVBs function, remodels the neuronal surface proteome, reduces synapse density, and alters behavior in mice. This platform enables systematic discovery of neuronal activity regulators and reveals a critical role for TMEM50A-dependent MVB function in maintaining synaptic integrity and behavior.

Single-cell transcriptomics identifies neural fate disruption and glial reprogramming caused by RARS2 deficiency.

BACKGROUND Pontocerebellar hypoplasia type 6 (PCH6) is caused by biallelic pathogenic variants in RARS2, encoding mitochondrial arginyl-tRNA synthetase. Although mitochondrial dysfunction is a recognised feature, how RARS2 deficiency disrupts neural lineage development remains unclear. METHODS We generated rars2-deficient zebrafish using the clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9) system and performed single-cell RNA sequencing (scRNA-seq) at 48 hours postfertilisation, complemented by immunofluorescence, in situ hybridisation, behavioural assays and ultrastructural analyses. Neural lineage composition, developmental trajectories, intercellular communication and transcriptional programmes were systematically examined. RESULTS rars2 -/- zebrafish displayed impaired survival, locomotor deficits, early mitochondrial ultrastructural damage and marked disruption of neurogenesis. scRNA-seq revealed reduced neuronal populations and expansion of neural progenitor and glial-like cells. Key neurogenic regulators (neurod4, her6 and pou3f1) were downregulated, whereas glial and stress-associated markers (hmgb1a, fabp7a and foxp1b) were upregulated. Developmental pathways including Notch and non-canonical Wnt were attenuated while extracellular matrix (ECM), adhesion and inflammatory programmes were activated. Additional trajectory-based analyses supported dysregulated lineage progression characterised by glial programme activation and impaired maintenance of neurogenic differentiation. CONCLUSION RARS2 deficiency disrupts mitochondrial integrity and reprograms neural lineage development through coordinated suppression of neurogenic transcriptional networks and activation of glial/ECM programmes. These findings provide mechanistic insight into loss-of-function RARS2 deficiency and highlight candidate molecular pathways for future therapeutic investigation.