Fig. 2: Knockdown of AATF inhibits DNA damage repair in GSCs and sensitizes GB to chemoradiotherapy. | Nature Communications

Fig. 2: Knockdown of AATF inhibits DNA damage repair in GSCs and sensitizes GB to chemoradiotherapy.

From: Elevated nonhomologous end-joining by AATF enables efficient DNA damage repair and therapeutic resistance in glioblastoma

Fig. 2: Knockdown of AATF inhibits DNA damage repair in GSCs and sensitizes GB to chemoradiotherapy.The alternative text for this image may have been generated using AI.

a–d 4121 GSCs transduced with shNT or shAATFs (a, b), or Flag-vector or Flag-AATF (c, d), were treated with IR (3 Gy) for indicated time points. IB (a, c) and IF (b, d) analyses of γ-H2AX are shown. Nuclei counterstained with Hoechst (blue). Left: representative images; Scale bars (b, d), 10 μm. Right: quantifications of γ-H2AX, each point represents a single cell (n = 3, biologically independent experiments). e 4121 GSCs transduced with shNT or shAATF#1 (red), or Flag-vector or Flag-AATF (green), were treated with increasing IR dose for 48 h or 72 h. Cell viability assessed by CellTiter-Glo assay. (Two-way ANOVA). f–j BALB/c nude mice intracranially implanted with GSCs transduced with Luciferase and Tet-on-inducible shNT or shAATF. Mice treated with control, IR (3 Gy, weekly, 3 times), Dox (1 mg/mL in water), or combined treatment from day 9 post-implantation (f, g, top). GB xenografts tracked by bioluminescence; representative images and quantifications are shown (f, g, n = 6, mean ± SD; Unpaired two-sided Student’s t-test). Kaplan–Meier survival plots of mice are shown (h, Log-rank Mantel–Cox test). Co-IF staining of SOX2 with γ-H2AX (i) or cleaved-caspase3 (CCP3) (j) in GB xenografts are shown. Left: representative images; Scale bars, 20 μm. Right: quantifications of γ-H2AX or CCP3 intensity, each point represents a single cell (n = 4, biologically independent samples). k 4121 GSCs transduced with shNT or shAATFs were treated with TMZ (50 μM) for indicated time points. IB analysis of indicated proteins are shown. l, m BALB/c nude mice intracranially implanted with GSCs (Luciferase) transduced with Tet-on-inducible-shNT or shAATF. Mice treated with control, TMZ (40 mg/kg, every 2 days, 5 times), Dox (1 mg/mL in water), or combined treatment from day 9 post-implantation (l, top). GB xenografts tracked by bioluminescence; representative images and quantifications are shown (l, n = 6 mice for each group; mean ± SD; Unpaired two-sided Student’s t-test). Kaplan–Meier survival plots of mice are shown (m, Log-rank Mantel–Cox test). For IB, protein levels were normalized to GAPDH. Independent experiments were performed three times with similar results. The curves represent the relative expression of γ-H2AX protein (a, c, k). Data represent mean ± SD of three independent experiments (a–e, i–k). Unpaired two-sided Student’s t-test. Source data are provided as a Source Data file.

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