Inhibition of prohibitin 2 enhances the sensitivity of non-small cell lung cancer cell line A549 to erlotinib

Abstract

Objective To explore the effects of prohibitin 2(PHB2) on sensitivity of non-small cell lung cancer cell line A549 to erlotinib (Erl) and its potential mechanisms. Methods RACK1-specific small interfering RNA was transfected in A549 cells for knocking-down of PHB2. The effects of PHB2 inhibition on cell proliferation and apoptosis induced by Erl were observed. The colocalization of microtuble-associated protein light chain 3 alpha (LC3) and mitochondria was visualized by MitoTracker staining and green fluorescent protein-microtuble-associated protein light chain 3 alpha (GFP-LC3) transfection. Cell proliferation was detected by 5-ethynyl-2′-deoxyuridine (EdU) staining. Cell colony formation was evaluated by colony forming assay. Apoptotic index of A549 cells was evaluated by TUNEL. Western blot was used to measure the expressions of PHB2 and LC3Ⅱ. Mitochondrial transmembrane potential, cytochrome c and respiratory chain complex Ⅰ/Ⅱ/Ⅴ activity were analyzed by the commercially available kits. Results Compared with the siPHB2 and siCtrl+Erl group, the EdU-positive A549 cells and the number of cell colonies decreased significantly (P＜0.05), while the TUNEL-positive A549 cells increased significantly(P＜0.05) in the siPHB2+Erl group. In addition, compared with the siPHB2 and siCtrl+Erl group, mitochondrial transmembrane potential and respiratory chain complex Ⅰ/Ⅱ/Ⅴ activity decreased significantly (all P＜0.05) and the levels of cytochrome c increased in mitochondrial fractions (P＜0.05) and decreased in cytosolic fractions(P＜0.05) in the siPHB2+Erl group. Conclusions PHB2 inhibition significantly improves sensitivity of A549 cells to Erl,which may be explained by inhibition of PHB2-mediated mitochondrial autophagy.

Keywords

• prohibitin 2|erlotinib|non-small cell lung cancer|proliferation|apoptosis