Figure 1. Expression of wild‐type and ALS/FTD‐mutant FUS reduces ER–mitochondria associations in NSC34 cells.

• A
  Expression of FUS does not alter expression of VAPB, PTPIP51 or mitofusin‐2 (MFN2) in transfected NSC34 cells. Immunoblots of NSC34 cells transfected with EGFP as a control (CTRL), or wild‐type or mutant EGFP‐FUS. Transfected cells were purified via EGFP using a cell sorter and the samples probed on immunoblots as indicated. On the FUS immunoblot, samples were probed with FUS antibody to show endogenous and transfected proteins; tubulin is shown as a loading control.
• B
  Representative electron micrographs of ER–mitochondria associations in NSC34 cells transfected with control EGFP vector (CTRL), EGFP‐FUS, EGFP‐FUSR521C or EGFP‐FUSR518K as indicated; arrowheads with loops show regions of association. Scale bar = 200 nm. Bar chart shows % of the mitochondrial surface closely apposed to ER in the different samples. Data were analysed by one‐way analysis of variance (ANOVA) followed by Tukey's multiple comparison test. N = 27–30 cells and 247–424 mitochondria, error bars are s.e.m.; ***P < 0.001.
• C, D
  siRNA loss of FUS does not affect ER–mitochondria associations or alter expression of VAPB, PTPIP51 or mitofusin‐2 (MFN2) in NSC34 cells. (C) Immunoblots of cells either mock transfected or treated with control (CTRL) or FUS siRNAs; GAPDH is shown as a loading control. (D) Representative electron micrographs of ER–mitochondria associations in control (CTRL) and FUS siRNA‐treated cells. Arrowheads with loops show regions of association. Scale bar = 200 nm. Data analysed by unpaired t‐test. N = 27–28 cells and 193–202 mitochondria, error bars are s.e.m.