Fig. 2: SC-EC and SC-β cells have unique and common transcriptional and chromatin accessibility signatures.
a, UMAP showing the trajectory of cells from the SC-β, SC-EC1 and SC-EC2 populations (subset of 21,317 cells from 3 independent differentiations; integration of all samples). b, Trajectory heat map showing dynamic changes of gene expression and motif accessibility enriched in β and EC groups. c, Volcano plots showing differential gene expression analysis (left) and differential motif accessibility analysis (right), highlighting relevant genes associated with SC-β and SC-EC cell populations. Statistical significance assessed by two-sided Wilcoxon rank sum test for RNA expression and two-sided logistic regression for motif chromatin accessibility. d, UMAP showing subpopulations by reclustering SC-β or SC-EC cell populations. Violin plots show gene marker expressions of INS and TPH1, highlighting off-target genes. e, Heat map showing DNA-binding motif accessibility associated with SC-β and SC-EC cells in subpopulations. Selected transcription factors plotted to show distribution of cells with target or off-target motif accessibility. f, Schematic of CRISPRa experiment for overexpression of CTCF in differentiating pancreatic progenitor cells. g, qPCR analysis of differentiated SC-islets with CTCF overexpression during endocrine induction, plotting mean ± s.e.m. (n = 4 biologically independent samples), showing expression differences in doxycycline treated compared with untreated control (INS, P = 0.0010; IAPP, P = 1.5 × 10−7; ISL1, P = 3.0 × 10−5) and EC cells (SLC18A1, P = 1.6 × 10−4; FEV, P = 0.0019; DDC, P = 1.2 × 10−4; TPH1, P = 0.0047; LMX1A, P = 4.6 × 10−4). Control represents cells without doxycycline treatment. Statistical significance was assessed by unpaired two-sided t-test. h, ICC quantification of cells expressing C-peptide protein (P = 5.3 × 10−6) and SLC18A1 protein (P = 3.1 × 10−4) with or without CTCF overexpression, plotting mean ± s.e.m. (control; n = 6 biologically independent samples, doxycycline; n = 7 biologically independent samples). Control represents cells without doxycycline treatment. Statistical significance was assessed by unpaired two-sided t-test. i, Volcano plots showing differential motif chromatin accessibility analysis of SC-endocrine population comparing control and CTCF overexpression (12,467 cells from 2 independent biological samples, 1 of each condition from the same differentiation batch; integration of all samples). Statistical significance was assessed by logistic regression. Control represents cells without doxycycline treatment. SC, stem cell derived; EC, enterochromaffin cells; CRISPRa, CRISPR activation.
