Fig. 4: Cell-cycle arrest and senescence in STING competent and deficient (KO) B16-GM cells following C′ dot exposure.
a, Schematic depicting particle-induced cell-cycle arrest and senescence in B16-GM cells. b, Percentage (%) of p-H2AX + STING-competent B16-GM cells detected by flow cytometry after particle exposure (15 μM, 72 h) relative to vehicle. c, Cell-cycle analysis using propidium iodide and phosphorylated histone 3 (pHH3) staining after C′ dot or vehicle treatment. d, Proportion of beta-galactosidase (β-Gal+)-expressing B16-GM cells with or without C′ dots (15 μM, 72 h) measured by flow cytometry, and e, in a concentration-dependent manner by colorimetric analysis with microscopy. f,g, RT-qPCR of p21 expression (f) and cytosolic DNA quantification in vehicle- and 15 μM C′ dot-treated cells. Genomic-to-mitochondrial DNA ratios (g) were determined using nuclear DNA (Tert) and mitochondrial DNA (Mt-DNA; D-loop) markers. h, Mean fluorescence intensity (MFI) of autophagosome formation during autophagy in particle- and vehicle-treated cells labelled with DALGreen dye by flow cytometry. i, Western blot analysis of B16-GM (WT), B16-GM STING-KO and B16-GM STING scrambled (scr) cells (right) confirming STING-KO efficiency, with β-actin (left) as a loading control. j, Cell-cycle profiling of STING-KO cells, as in c. k, β-Gal+ staining in B16-GM STING-KO cells exposed to vehicle or C′ dots (15 μM, 72 h). Data reflect analyses of three biological replicates for particle- and vehicle-treated (control) groups computed as the mean ± s.e.m. Unpaired Student’s t-test and non-parametric one-way ANOVA with Tukey’s test for multiple comparisons were performed. All statistical tests were two-sided. *P < 0.05, **P < 0.01, ***P < 0.005 and ****P < 0.001. Schematic in a created with BioRender.com.
