Figure 3: LncBRM associates with BRM to initiate the BRG1/BRM switch.

(a) LncBRM intron sequence (Ctrl), lncBRM antisense (Lnc-AS) and lncBRM transcripts were labelled with biotin and incubated with oncosphere lysates, followed by silver staining and mass spectrometry. Black arrow denotes BRM. (b) RNA pulldown was conducted using lncBRM transcript, followed by immunoblotting. (c) Domain mapping of lncBRM transcript. (d) LncBRM was incubated with increased doses of BRM, followed by electrophoretic mobility shift assay (EMSA). The 3 segment of lncBRM was labelled with biotin for probing. (e) Non-spheres and spheres were visualized by fluorescence in situ hybridization (FISH). Scale bar, 10 μm. (f) Antibodies against BRM or BRG1 were used for RNA immunoprecipitation, followed by RT–qPCR. ACTB served as a negative control. (g) Spheres (S) and non-sphere cells (N) were lysed and followed by immunoprecipitation with BAF170 and ARID1A antibodies. BRG1 and BRM enrichment was analysed with western blotting. (h) Different doses of lncBRM transcripts were incubated with oncosphere lysates and followed by co-immunoprecipitation (co-IP). (i) LncBRM-depleted HCC primary spheres were lysed for co-IP as in h. (j,k) The indicated oncosphere lysates were fractionated and followed by size fractionation with glycerol gradient ultracentrifugation. Elute gradients were used for western blotting. Data are shown as means±s.d. Two tailed Student’s t-test was used for statistical analysis, ***P<0.001. Data represent at least three independent experiments.