Expression of Cdk4, Cyclin D1 and Rb in exogenous wild type P16 gene transfected bladder cancer cell line

Abstract

Cell cycle control plays a key role in the growth of normal mammalian cells. One of the fundamental abnormalities in human tumors is dysregulated cell cycle control. It is generally accepted that active Cdk4–Cyclin D1 complexes help cells to pass through the R point, a point of no return, after which cells become committed to a new round of replication. Accumulated evidences indicate that Cdk4 is the ‘primary sensor’ for driving cells through the R point and it is widely known that P16INK4a can arrest cells in G1.But how can the expression of exogenous P16INK4 gene affect the activity of Cdk4–Cyclin D1 remains unclear. In this study, using exogenous wild type P16 gene and antibodies for P16, Cdk4, Cyclin D1 and Rb proteins, we examined the expression of exogenous wild type P16 gene and the changes of cell cycle regulatory genes-Cdk4, Cyclin D1 and Rb in human bladder cancer cells. The cell growth analysis revealed that the proliferation of P16 gene transfected cancer cells was inhibited after the transfection of exogenous wild P16 gene. The immunocytochemical results indicated that after the transfection of exogenous wild type P16 gene, the expression of Cdk4, Cyclin D1 and Rb were negative in the nuclei while the expression of P16 significantly increased in the nuclei and the cytoplasm. Our results suggest that the transfection of exogenous wild P16 gene induces the inhibition of proliferation of the bladder cancer cells and the increasing expression P16 inhibits the expression of Cdk4, Cyclin D1 and Rb in nuclei, which results in the cell cycle being inhibited at G0/G1 phase. As a consequence, exogenous P16 has negative effects on the malignant proliferation of bladder cancer cells and it may be considered as target for potential anti-cancer drugs.