Fig. 4: P-TEFb activity is required for canonical DNA damage response programs.

Gene ontology network (a) and terms (b) constructed from genes significantly induced by 6 Gy IR but abrogated with concurrent AZD4573 (40 nM) across three cell lines (SU-DIPG4, HSJD-DIPG007, SF8628; n = 3 each condition, LFC + /− 1.2 with p val <0.05). Each node denotes an enriched term, with color density reflecting -log(Pval). Enrichment defined by Metascape using hypergeometric test and Benjamini-Hochberg P value correction algorithm. c. DNA damage as measured by flow cytometry for yH2AX 6 h (left) or 24 h (right) after IR, 8 nM AZD4573, or combination. Comparison reflects p value of two-tailed Student’s t-test, mean ± SEM of n = 3 biologically independent replicates. d. Representative immunofluorescent staining for yH2AX in SU-DIPG4 at same timepoint and conditions as (c) (bar = 10 µm). e. IR-induced G2M arrest in the presence or absence of AZD4573 as measured by increase in G2M fraction from cells treated with 8 nM AZD4573, IR, or combination. Comparison reflects p value of two-tailed Student’s t-test, mean ± SEM of n = 3 biologically independent replicates per cell line. f. Cell cycle distribution of HSJD-DIPG007 cells synchronized to G0/G1 (T0, left) and then treated either while in G0/G1 synchronization (G1 Syn, middle) or after G1 release (G1 Rel, right). n = 3 biologically independent replicates per condition. g. Caspase 3/7 activation measured 24 h after treatment with 4 nM AZD4573, IR, or combination. Comparison reflects p value of two-tailed Student’s t-test, mean ± SEM of n = 4 biologically independent replicates, with HSJD-GBM001 imaged 4 fields per replicate. Representative fluorescent live-cell imaging from HSJD-GBM001 shown on right (bar = 200 µm). Source data are provided as a Source Data file.