RACK1 promotes the proliferation of lung cancer cells by targeting phosphorylation of MCM7 protein / 肿瘤

ABSTRACT

Objective: To investigate the effects of gene silencing and overexpression of RACK 1 (receptor for activated C kinase 1) on the proliferation of large-cell lung cancer H460 cells and lung adenocarcinoma A549 cells, and to explore the possible mechanism. Methods: The RACK1 siRNA (small interfering RNA) targeting RACK 1 gene and recombinant vector pCMV-sport6-RACK1 were transfected into both of H460 cells and A549 cells, respectively. MTT method and colony formation assay were used to detect the effect of RACK 1 gene expression on the proliferation of lung cancer cells. Flow cytometry was used to examine the change of cell cycle. The association and interaction of RACK 1 gene expression with the proliferation of lung cancer cells were analyzed by yeast two-hybrid system, co-immunoprecipitation, laser scanning confocal microscopy and co-immunoprecipitation of phosphoproteins. Results: The expression levels of RACK1 protein in the H460 cells and A549 cells were both decreased after transfection with RACK1 siRNA, and the abilities of proliferation and colony-formation were also weakened. The proportion of lung cancer cells arrested at phase S was significantly declined (P <0.01). Meanwhile, the expression level of RACK1 protein was increased after transfection with pCMV-sport6-RACK1, and the abilities of proliferation and colony-formation of lung cancer cells were both strengthened with a prolonged doubling time. The proportion of lung cancer cells arrested at phase S was significantly increased (P <0.01). The results of yeast two-hybrid system and co-immunoprecipitation revealed that RACK1 could directly interact with MCM7 (minichromosome maintenance protein 7). The phosphorylation of MCM7 protein was strengthened through binding to RACK1 which translocated into the cell nucleus. Conclusion: RACK1 promotes the proliferation of lung cancer cells through activating the phosphorylation of MCM7 binding to RACK1. Copyright© 2012 by TUMOR.