Effect of eEF-2K down-regulation on breast cancer cell proliferation and clonogenicity.

(A) Knockdown of eEF-2K by siRNA. siRNA specifically targeting eEF-2K inhibits its expression in MDA-MB-231 cells. Cells were transfected with eEF-2K siRNA (5–75 nM) for 48 h, and cell lysates were subjected to Western blot analysis. (B) Effect of knockdown of eEF-2K on cell proliferation. MDA-MB-231 cells were transfected with eEF-2K siRNA, and after 48 h proliferation was detected by an MTS assay. (C) Overexpression of eEF-2K in MDA-MB-231 cells. Cells were transiently transfected with an expression vector encoding GST-tagged wild type eEF-2K, and cell lysates (48 h) were subjected to Western blot analysis. For the effect of overexpression of eEF-2K on cell proliferation, MDA-MB-231 cells were transiently transfected with eEF-2K or a vector control, and cell proliferation was measured using an MTS assay after 96 h. (D–E) Effect of eEF-2K knockdown on colony formation. Knockdown of eEF-2K by siRNA (50 nM) significantly inhibited the number of colonies formed by MDA-MB-231 (D) and MCF-7 (E) cells (*p<0.05). Cells were transfected every 4 days with control or eEF-2K siRNA.