Fig. 5

Endogenous FUS is mislocalized to the cytoplasm and is incorporated into cytoplasmic stress granules in response to arsenite in patient iNeurons. a A representative control shows intense staining for FUS (green) in the nuclei (DAPI) in TUJ1-positive (red) iNeurons at day 10 of neuronal induction, whereas the patients show a majority of FUS protein in the cytoplasm. Cells were counter stained with the nuclear marker DAPI (blue). Scale bars = 50 μm. b Confocal images of vehicle treated iNeurons (left panel) as compared to cells treated with 0.5 mM arsenite for 30 min (right panel) at day 10 are shown. A representative control shows FUS protein predominantly localized to the nuclei. ALS-FUS patient with Q519E mutation recapitulated the FUS neuropathology only in iNeurons: iNeurons from the patient show a majority of FUS protein (green) in the cytoplasm. In response to oxidative stress conditions, cytoplasmic FUS-positive inclusion bodies (green) in iNeurons were co-localized with G3BP stress granules (red). Cells were fixed and probed by immunofluorescence for DAPI (blue). Scale bars = 25 μm. Bar graphs represent (c) the numbers of stress granules and (d) the numbers of FUS-positive stress granules (SGs). Data are from three experiments (the mean ± SEM, n = 20). One-way ANOVA followed by Tukey multiple comparisons test; **p < 0.001; N.S., not significant