Fig. 3

LINC00470 anchored FUS in the cytoplasm and phosphorylated FUS. a Upper, schematic illustration of substitution mutant constructs of LINC00470; middle and lower, an RNA pulldown assay examined the interaction between FUS and the different mutants of LINC00470. b GST pulldown assays showed that the RRM domain of FUS pulled down LINC00470. c Representative immunofluorescence staining displayed the co-localization of LINC00470 and FUS in the cytoplasm of HEK293 cells after LINC00470 overexpression. Scale bar, 20 μm. d Representative imaging of LINC00470 anchoring FUS in the cytoplasm in HEK293 cells. Scale bar, 20 μm. e Western blotting measured the expression of FUS in whole cell lysis and the cytoplasm and nucleus in U251 cells transfected by pcDNA3.1-LINC00470. f Representative immunoprecipitation analysis detected FUS phosphorylation in U251 cells transfected by pcDNA3.1-LINC00470. FUS immunoprecipitated from U251 cells was immunoblotted with pan-phospho-S/TQ antibodies to assess the phosphorylation level of FUS. g RNA fluorescence in situ hybridization showed the localization of LINC00470 in GBM cells. The nucleus was counterstained with DAPI. Scale bar, 29 μm