Fig. 5.

Minority MOMP stimulates cGAS/STING in mMKs post IR. a Relative mtDNA amounts in cytosols of mouse BM mMKs at 1 dpi were assessed by quantitative polymerase chain reaction (n = 3). b Colocalization of cytosolic DNA with cGAS in BM mMKs of mice at 1 dpi was assessed by immunofluorescence staining with antibodies specific for DNA and cGAS. Scale bar = 20 μm. The arrow indicates cGAS and DNA colocalization. c Relative enrichment of DNA fragments as indicated from FLAG-cGAS immunoprecipitants in Meg-01 cells at 1 dpi (n = 3). d p-TBK1 and p-IRF3 expression in BM mMKs of mice at 1 dpi were assessed by immunofluorescence staining. Scale bar = 20 μm. e Western blotting analysis and quantification of p-TBK1 and p-IRF3 expression in BM mMKs of mice at 1 dpi (n = 3). f Flow cytometric analysis and quantification of p-TBK1 and p-IRF3 expression in BM mMKs of mice at indicated dpi (n = 5). g MPV in peripheral blood of Sting^fl and Sting^cko mice at indicated dpi (n = 5). h CD62P expression in response to thrombin on washed platelets from Sting^fl and Sting^cko mice at indicated dpi (n = 4–5). Data represent mean ± standard deviation. ^*P < 0.05, ^**P < 0.01, compared to Ctrl or IR/Sting^fl as indicated. Two-tailed unpaired student’s t-test. ns non-significance, MOMP mitochondrial outer membrane permeabilization, mMK mature megakaryocyte, cGAS cyclic GMP-AMP synthase, STING stimulator of interferon genes, Ctrl control, IR ionizing radiation, mtDNA mitochondrial DNA, BM bone marrow, dpi day post IR, TBK1 TANK-binding kinase 1, IRF3 interferon regulatory factor 3, MPV mean platelet volume, Cox1 cytochrome c oxidase subunit 1, ND1-4 NADH dehydrogenase subunit 1–4, LINE1 long interspersed nuclear element 1, RNA18S 18S ribosomal RNA, nDNA nuclear DNA, DAPI 4′,6-diamidino-2-phenylindole, GAPDH glyceraldehyde-3-phosphate dehydrogenase