Figure 5: Constitutive DNA damage and type I interferon activation in TREX1-deficient cells.

(a) Alkaline single-cell electrophoresis depicting DNA damage in patient fibroblasts (LE, AGS) as shown by formation of longer comets with deformed nuclei compared with wild type cells (WT). Scale bar, 100 μm (left). The Olive tail moment (OTM) corresponds to the amount of DNA fragmentation (right). Means and s.e.m. of two independent experiments for each patient (LE1, LE2, AGS1, AGS2) and WT control cell lines (WT, n=2). ***P<0.001. (b) Increased pan-nuclear γH2AX-staining in TREX1-deficient fibroblasts determined by flow cytometry. Means and s.d. of at least three independent experiments for each patient (LE1, LE2, AGS1, AGS2) and WT controls (WT, n=5). **P<0.01; ***P<0.001. (c) Growth curves of TREX1-deficient fibroblasts (LE1, LE2, AGS1, AGS2) and WT cells (n=5). *P<0.05 (LE1) and **P<0.01 (LE2, AGS1, AGS2) versus WT at day 7. (d) Immunoblot analysis of unstressed patient fibroblasts (LE1, LE2, AGS1, AGS2) showing activation of p53 (p-p53; Ser15), p16, Chk1 (p-Chk1; Ser345) as well as IRF3 (p-IRF3). β-actin was probed as a loading control. (e) Senescent phenotype of patient fibroblasts (LE1, LE2, AGS1, AGS2) as shown by an increased percentage of β-galactosidase-positive cells (WT, n=4; left) and representative images of senescent cells (right). Scale bar, 100 μm. Means and s.d. of at least three independent experiments. *P<0.05; **P<0.01; ***P<0.001. (f) IFN-β secretion over 24 h. (WT, n=4). Means and s.e.m. of five independent experiments. *P<0.05; **P<0.01; ***P<0.001 versus WT.