Fig. 5

ATRX loss induces a global loss of CTCF, correlated gene dysregulation, and protects glioma cells from therapy-induced senescence. A Anti-CTCF single-cell CUT&Tag in SB28+IDH1R132H cells with ATRX KO or ATRX-wildtype scrambled control. Reads per 10 bp are shown as heatmaps (bottom) and summarized as averages (top) in a neighborhood of JASPAR CTCF motif sites. B CTCF motif frequencies in scCUT&Tag peaks, represented as standardized deviances from a data-driven null distribution via ChromVAR, for ATRX KO vs. wildtype cells (top), ***t test p < 1e−16. Numbers of, and overlap between, peaks in ATRX KO and wildtype datasets. C Browser shots of CTCF scCUT&Tag read pile-ups with significant peaks and genes annotated. D The percentages of NSC chromatin-loops affected, directly or indirectly, by CTCF loss upon ATRX KO (top). A Q-Q plot of the average ATRX-KO/WT cell-averaged log fold-changes in gene expression for each chromatin loop determined from NSC Hi-C data. Values on the y-axis are represented as percentages of a corresponding null distribution (Methods). The x-axis quantiles are provided by a Beta distribution fit to the inter-quartile range of y-axis percentiles. Loops containing specific genes are annotated. E Browser shots of a chromatin loop containing VEGFA and closeup of one boundary domain (left). A null distribution of average ATRX-KO/WT cell-averaged log fold-changes in gene expression measured via snRNA-seq in vivo in neoplastic cells, taken across all consecutive windows of four adjacent genes in chromosome 17. The loop containing VEGFA and the percentile of its average log fold-change are annotated (right-top). Violin plots of Vegfa expression in ATRX KO and WT neoplastic cells, in vivo, *MAST q < 0.05 (right-bottom). F As in E, except for a loop containing TNC. G Fluorometric assay for β-galactosidase activity in ATRX-KO and scrambled-control SB28 cells, both expressing IDH1R132H, post-treatment with one of Doxorubicin or CDKi. Error bars indicate standard error. *t test p < 0.05. B As in A, but for patient-derived ATRX/IDH1 double-mutant cells (SF10602) compared with an ATRX-wildtype, IDH1-mutant control (SF10417). Cells were treated with one of Temozolomide, Imatinib, Doxorubicin, or CDKi. C Visualization of β-galactosidase expression in ATRX-KO and scrambled-control SB28 cells, both expressing IDH1R132H, before and after treatment with CDKi. H Model of gene dysregulation due to CTCF disruption in chromatin-loop boundaries