Fig. 8

FAM49B positively regulates Rab10/TLR4 pathway by stabilizing ELAVL1. A Exogenous interaction between FAM49B and ELAVL1 in 293 cells, observed via co-immunoprecipitation. B pLNCX2-FAM49B or pLNCX2-vector were transfected into FAM49B-shRNA-expressing MCF-7 or MDA-MB-231 cells. ELAVL1 expression was quantified via western blotting (n = 3). C Level of ELAVL1 mRNA was determined using quantitative real-time PCR in FAM49B-shRNA-expressing MCF-7 or MDA-MB-231 cells. D FAM49B mediated ELAVL1 stabilization in BC cells. scr- or FAM49B-shRNA-expressing MDA-MB-231 cells were treated with CHX (50 μM) for the indicated time points and analyzed for endogenous ELAVL1 expression via western blotting. E FAM49B inhibition increased ELAVL1 ubiquitination in MDA-MB-231 cells. HA-Ubiquitin was electroporated into scr- or FAM49B-shRNA-expressing MDA-MB-231 cells. Cells were treated with MG132 (20 μM) for 2 h. ELAVL1 complex in resulting lysates was examined using an antibody against Ubiquitin (Ub). F, G scr- or ELAVL1-shRNA were transfected into FAM49B-overexpressing MCF-7 and MDA-MB-231 cells. ELAVL1, Rab10, and TLR4 expression was quantified via western blotting (n = 3). Results are presented as the mean ± SD. The statistical significance was assessed by Student’s t-test; *p < 0.05, **p < 0.01