Fig. 1: TRF2 forms a complex with INM proteins and directly interacts with Emerin.

A Protein-protein interaction map (STRING) to explore TRF2-nuclear inner nuclear membrane proteins network. B Co-immunoprecipitation experiment with anti-TRF2 antibody in MDA-MB-231 cells. Mouse immunoglobulines (IgG M) were used as negative control. Immunoblots were probed with the indicated antibodies. C ChIP qPCR analysis shows the binding of TRF2 and Lamin B1 on the indicated LADs. SCN10A region, was used as positive control of LADs. D Upper panel, schematic representation of TRF2 and its deletion mutants used in this study. Bottom panel, the various GST-TRF2 proteins or GST alone were affinity-purified and incubated with lysates prepared from MDA-MB-231 cells, followed by Western blotting with indicated antibodies. E Proximity ligation assay to visualize interaction of TRF2 with Lamin B1 in MDA-MB-231 cells stable overexpressing or silenced for TRF2. Left panel: representative images. Right panel: quantification of TRF2-LaminB1 PLA spots per nucleus (n = 60 cells), red lines represent mean values. F Proximity ligation assay to visualize interaction of TRF2 with Emerin in MDA-MB-231 cells stable overexpressing or silenced for TRF2. Left panel: representative images. Right panel: quantification of TRF2-Emerin PLA spots per nucleus (n = 60 cells), red lines represent mean values. G In vitro GST pull-down assay with GST alone or GST-TRF2 in the presence of myc-Emerin recombinant protein. Following the pull-down, the samples were analyzed by western blotting using specific anti-Emerin antibody. H GST-Emerin protein or GST alone was affinity-purified and incubated with lysates prepared from MDA-MB-231 cells, followed by Western blotting with indicated antibodies. For (D) (bottom panel), (G, H), purified GST fusion proteins (indicated by the asterisks) were visualized by ponceau staining. For (E, F), Mann Whitney test was used to calculate statistical significance. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.