Human cerebral organoids reveal progenitor pathology in EML1-linked cortical malformation.

Malformations of human cortical development (MCD) can cause severe disabilities. The lack of human-specific models hampers our understanding of the molecular underpinnings of the intricate processes leading to MCD. Here, we use cerebral organoids derived from patients and genome edited-induced pluripotent stem cells to address pathophysiological changes associated with a complex MCD caused by mutations in the echinoderm microtubule-associated protein-like 1 (EML1) gene. EML1-deficient organoids display ectopic neural rosettes at the basal side of the ventricular zone areas and clusters of heterotopic neurons. Single-cell RNA sequencing shows an upregulation of basal radial glial (RG) markers and human-specific extracellular matrix components in the ectopic cell population. Gene ontology and molecular analyses suggest that ectopic progenitor cells originate from perturbed apical RG cell behavior and yes-associated protein 1 (YAP1)-triggered expansion. Our data highlight a progenitor origin of EML1 mutation-induced MCD and provide new mechanistic insight into the human disease pathology.

Natural variation in gene expression and viral susceptibility revealed by neural progenitor cell villages.

Human genome variation contributes to diversity in neurodevelopmental outcomes and vulnerabilities; recognizing the underlying molecular and cellular mechanisms will require scalable approaches. Here, we describe a "cell village" experimental platform we used to analyze genetic, molecular, and phenotypic heterogeneity across neural progenitor cells from 44 human donors cultured in a shared in vitro environment using algorithms (Dropulation and Census-seq) to assign cells and phenotypes to individual donors. Through rapid induction of human stem cell-derived neural progenitor cells, measurements of natural genetic variation, and CRISPR-Cas9 genetic perturbations, we identified a common variant that regulates antiviral IFITM3 expression and explains most inter-individual variation in susceptibility to the Zika virus. We also detected expression QTLs corresponding to GWAS loci for brain traits and discovered novel disease-relevant regulators of progenitor proliferation and differentiation such as CACHD1. This approach provides scalable ways to elucidate the effects of genes and genetic variation on cellular phenotypes.

Human amyotrophic lateral sclerosis excitability phenotype screen: Target discovery and validation.

Drug development is hampered by poor target selection. Phenotypic screens using neurons differentiated from patient stem cells offer the possibility to validate known and discover novel disease targets in an unbiased fashion. To identify targets for managing hyperexcitability, a pathological feature of amyotrophic lateral sclerosis (ALS), we design a multi-step screening funnel using patient-derived motor neurons. High-content live cell imaging is used to evaluate neuronal excitability, and from a screen against a chemogenomic library of 2,899 target-annotated compounds, 67 reduce the hyperexcitability of ALS motor neurons carrying the SOD1(A4V) mutation, without cytotoxicity. Bioinformatic deconvolution identifies 13 targets that modulate motor neuron excitability, including two known ALS excitability modulators, AMPA receptors and Kv7.2/3 ion channels, constituting target validation. We also identify D2 dopamine receptors as modulators of ALS motor neuron excitability. This screen demonstrates the power of human disease cell-based phenotypic screens for identifying clinically relevant targets for neurological disorders.