Fig. 4

Traf3 gene silencing abolishes the antiapoptotic function of GMEB1. a Assessment of viability of HeLa cells, treated by CHX and TNF-α, using MTT assay. A shRNA-expression vector targeting TRAF3 (shRNATRAF3) (0.5 μg), a control shRNA-expressing vector (shRNA luc) (0.5 μg) and GMEB1-His (1 μg) were cotransfected to HeLa cells. 20 h postransfection, HeLa cells were treated with CHX and TNF-α for 24 h, in order to induce apoptosis. Viability is calculated as the ratio of O.D. sample/O.D. control untreated * 100. The data are presented as the mean ± standard error (± SEM) of three independent repetitions. Statistical evaluation of the differences between the values was performed by Student’s t-test. Statistically significant differences are indicated by brackets and an asterisk (p < 0.05). b, c Immunoblot analysis. Total cell lysates from HeLa cells transfected with shRNATRAF3 or control shRNAluc, and GMEB1-His were analyzed by SDS-PAGE on an 8% gel. The detection of the indicated proteins was performed using the antibodies reported in “Methods” section. β-actin was used as a loading control