Lamprey prohibitin 2 inhibits non-small cell lung carcinoma cell proliferation by down-regulating the expression and phosphorylation levels of cell cycle-associated proteins

Abstract

Prohibitin 2 (PHB2) is an evolutionarily conserved and functionally diverse protein that plays an important role in multiple cellular functions, including cell proliferation, cell migration, and apoptosis, and is also known to participate in the process of tumorigenesis and development. In this study, the lamprey PHB2 (Lm-PHB2) gene was over-expressed in KRAS (kirsten rat sarcoma viral oncogene homolog)-mutated non-small cell lung carcinoma (NSCLC) cells to investigate its effect on cell proliferation. The effects of Lm-PHB2 protein on the proliferation of NSCLC cells were determined by treating cells with the purified recombinant Lm-PHB2 protein (rLm-PHB2) followed by cell counting kit (CCK) assays and flow cytometry. Analysis showed that rLm-PHB2 blocked cells in the G2 phase and inhibited the cell proliferation of A549, Calu-1, and NCI-H226 to various degrees. The effect on Calu-1 cells was the most obvious and was concentration- and time-dependent. Similarly, cells transfected with the pEGFP-N1-Lm-PHB2 plasmid also resulted in the suppression of proliferation in A549 cells and Calu-1 cells. Quantitative real-time polymerase chain reaction (qRT-PCR) showed that Lm-PHB2 inhibited cell proliferation by repressing the transcription of PLK1 (polo-like kinase 1), Wee1 (wee1 kinase), CCNB1 (cyclin B1), and CDC25C (cell division control protein 25C). According to western blot analysis, Lm-PHB2 not only down-regulated the expression of PLK1, Wee1, CCNB1, and CDC25C but also reduced the phosphorylation levels of CCNB1 and CDC25C, thus blocking Calu-1 cells in G2/M phase. Our findings demonstrate a function of lamprey PHB2 that may inhibit the proliferation of some NSCLC cells by down-regulating the expression and phosphorylation of cell cycle-associated proteins.