Fig. 8

Reintroduction of Nrf2 rescues the Mst1/2-null phenotype. a Flow cytometry analyzing of ROS levels in Mst1^fl/flMst2^fl/fl (WT) BMDMs, Mst1^fl/flMst2^fl/fl Lyz-Cre (DKO) BMDMs, or DKO BMDMs infected with adenovirus expressing GFP (Ad-GFP) or Nrf2 (Ad-Nrf2), with the CellRox dye and quantification of the relative fluorescence intensity in the indicated cell samples shown in the right panel. b–e RT-qPCR analysis of the mRNA levels of the antioxidant genes Ho-1 and Nqo-1 (b), fluorescence microscopy of p-H2A.X (red), DAPI-stained nuclei (blue) and GFP⁺ cells (green) (c), Annexin V/DAPI staining (d) and quantification of Annexin V⁺DAPI⁻ cells (e) in WT BMDMs, DKO BMDMs, or DKO BMDMs infected with Ad-GFP or Ad-Nrf2, followed with or without H₂O₂ treatment as indicated. Scale bars, 20 μm. f Relative fluorescence intensities of telomere FISH of GFP⁺ peritoneal macrophages isolated from WT mice transplanted for 6 months with DKO bone marrow cells, which were transduced with retrovirus expressing Nrf2, or control GFP, respectively. g A proposed working model for kinases Mst1/2 sense ROS and maintain cellular redox balance by modulating the stability of Nrf2. Phagosomal or mitochondrial ROS release attracts Mst1/2 to cap around phagosome or mitochondrion from the cytosol and activates Mst1/2; Mst1/2 phosphorylate Keap1 to stabilize Nrf2 and regulate the expression of antioxidant enzymes to protect cell against oxidative damage. Data are from one experiment representative of three independent experiments with similar results. ns, not significant (P > 0.05); *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001, compared with the control sample (Student’s t test). Data are from one experiment representative of three independent experiments with similar results (mean and s.d. of n = 3 (a, b, e); mean and s.e.m. of n = 50 (f); biological replicates (a, e); and experimental replicates (b, f))