Extended Data Fig. 8: Cell-type composition, full pseudotime trajectories, and gene modules of ARID1B^+/− and isogenic control organoids.

a, Immunohistochemistry for the postmitotic excitatory neuronal marker TBR1 (magenta) and GABAergic marker DLX2 (green) in Mito210 control and ARID1B^+/− organoids at one month (35 d.i.v.). Scale bars: 200 μm. b, c, scRNA-seq data from Mito210 one month (35 d.i.v. batch I in b, batch II in c) control and ARID1B^+/− organoids. Bar charts show the percentage of cells for all the cell populations in each control and mutant organoid. Adjusted p-values for a difference in cell-type proportions between control and mutant, based on logistic mixed models (see Methods) are shown. d, Immunohistochemistry for TBR1 (magenta) and DLX2 (green) in Mito210 control and ARID1B^+/− organoids at three months (90 d.i.v.). Scale bars: 100 μm. e, scRNA-seq data from Mito210 three months (90 d.i.v.) control and ARID1B^+/− organoids. Left top shows combined t-SNE plots of control and mutant organoids (n = 3 single organoids per genotype). Cells are coloured by cell-type, and the number of cells per plot is indicated. Left bottom, t-SNE plots for control and mutant individual organoids. GABAergic interneurons are highlighted in colour. Left, bar charts show the percentage of cells for all the cell populations in each control and mutant organoid, as shown in b, c. Two out of three mutant organoids show an increase in GABAergic interneurons, vs. zero out of three controls (adjusted p = 0.19, logistic mixed models). f, Immunohistochemistry for the postmitotic excitatory neuronal marker TBR1 (magenta) and GABAergic marker DLX2 (green) in Mito210 control and ARID1B^+/− organoids at three months (90 d.i.v.). Three out of four mutant organoids contain DLX2-positive cells, while no DLX2 expression is detected in the four controls. Scale bars: 500 μm. g, scRNA-seq data from Mito294 one month (35 d.i.v.) ARID1B^+/− and control organoids. Left top shows combined t-SNE plots of control and mutant organoids (n = 3 single organoids per genotype). Cells are coloured by cell type, and the number of cells per plot is indicated. Left bottom, t-SNE plots for control and mutant individual organoids. GABAergic neurons, newborn deep-layer projection neurons and immature deep-layer projection neuron populations are highlighted in colour. Right, bar charts show the percentage of cells for all the cell populations in each control and mutant organoid, as in b, c, e. h, Pseudotime trajectories from the full dataset of Mito210 ARID1B^+/− 35 d.i.v. batch II and control organoids, calculated with Monocle3. The partition highlighted by a box was subsetted and the trajectory is shown in Fig. 2c. i, Module scores (top) and their distribution across mutant and control cells (bottom) for all modules resulting from WGCNA analysis of the partition of interest from Mito210 ARID1B1^+/− and control organoids at 35 d.i.v. Cells were downsampled to have an equal number of cells per organoid. Names were assigned to each module based on the known functions of the genes included in each one. Horizontal bars show median scores, and dots show average score per organoid. Adjusted p-values show differences between control and mutant based on linear mixed models (see Methods). Number of organoids used for each analysis can be found in the Methods under “Statistics and reproducibility”. aRG, apical radial glia; DL, deep layer; UL, upper layer; PN, projection neurons; CP/CH, Choroid Plexus/Cortical Hem; oRG, outer radial glia; IPC, intermediate progenitor cells; CPN, callosal projection neurons; CFuPN, corticofugal projection neurons; GABA NP, GABAergic neuron progenitors; GABA N, GABAergic neurons; GABA INP; GABAergic interneuron progenitors; GABA IN, GABAergic interneurons.