Supplementary Figure 8: Characterization of iPSC-iNs.

a, Schematic showing the paradigm for direct conversion of iPSCs into induced neurons (iPSC-iNs) by forced expression of an inducible Ngn2 transgene (see Methods). b, Morphological development of iPSC-iNs over the time course of differentiation. Experiments were repeated with 13 independent subject lines with similar results. c, iPSC-iNs express hTau after two weeks of differentiation. The staining was repeated in all 13 subject lines at least once with similar results. d, Regional enrichment analysis for two-week-old iPSC-iNs. Shown are the results of transcriptome correlation analyses between two-week-old iPSC-iNs (ASD: n = 8, control: n = 5) and post-mortem human brain samples from the BrainSpan dataset (n = 524). The x-axis shows agglomerated brain regions of the BrainSpan post-mortem brain samples. The y-axis shows the relative percentage among brain regions that were classified for each iPSC-iN sample. The classification was based on a sample-specific correlation threshold (Supplementary Methods). Values represent means ± s.d.; NCX, neocortex (see e); AMY, amygdala; HIP, hippocampus; CGE, caudal ganglionic eminence; LGE, lateral ganglionic eminence; MGE, medial ganglionic eminence; DTH, dorsal thalamus; CB, cerebellum; STR, striatum. e, Enriched regions of the neocortex (NCX). Shown are percentage values for different cortical regions represented as NCX in f. MFC, medial prefrontal cortex; DFC, dorsolateral prefrontal cortex; OFC, orbital frontal cortex; VFC, ventrolateral prefrontal cortex; Ocx, occipital neocortex; Pcx, parietal neocortex; STC, superior temporal cortex; V1C, primary visual cortex; M1C-S1C, primary motor-sensory cortex. f, Heatmap showing normalized expression values for selected genes across all 14-day-old iPSC-iN samples. IPSC-iNs showed high expression levels of pan-neuronal markers such as RBFOX1, NCAM1 and MAP2 as well as telencephalic markers such as BRN2(POU3F2), CUX1 and SATB2. g, Heatmap showing changes in gene expression between iPSC and iPSC-iNs 14 days after conversion for selected genes and across all lines (see also Supplementary Table 8). Compared with their corresponding iPSC stages, iPSC-iNs showed significant upregulation of other cortical transcription factors including TBR1, whereas other prominent forebrain transcription factors such as CTIP2/BCL11B were lacking. h, Representative immunofluorescence image of two-week-old iPSC-iNs stained with βIII-tubulin (Tuj1) and vGlut1. Scale bar represents 5 μm. The staining was repeated in all 13 subject lines at least once with similar results. i, Electrophysiological characterization over the time course of iPSC-iN maturation. Values represent mean ± s.e.m.; 8 days (iPSC N 8, n = 8 neurons), 15 days (iPSC N 15, n = 9 neurons), 28 days (iPSC N 28, n = 4 neurons).