RNA interference-mediated silencing of the PAR gene inhibits the growth of PC3 cells via the induction of G2/M cell cycle arrest and apoptosis.

BACKGROUND

The prostate androgen-regulated (PAR) gene is ubiquitously overexpressed in prostate cancer (PCa) cells and is involved in proliferation of PCa. However, the mechanism by which the modulation of PAR gene expression elicits its biological effects on PCa cells is not well documented. Here, we investigate the mechanism of PAR depletion inhibiting PCa cell growth.

METHODS

PAR expression was depleted by small interfering RNA (siRNA) and its subsequent effects on proliferation of PC3 cells were determined by the trypan blue exclusion assay. Flow cytometric analysis provided the evidence for the progression of cell cycle and the induction of apoptosis which was further confirmed by the observation of cleavage of poly(ADP-ribose) polymerase. Western blot analysis was performed to investigate the involvement of critical molecular events known to regulate the cell cycle and the apoptotic machinery.

RESULTS

siRNA transfection results in a dose-dependent inhibition of cell growth in PC3 cells by causing G2/M phase cell cycle arrest and apoptosis. The G2/M arrest by PAR depletion was associated with decreased levels of cyclin B1, pCdc2 (Tyr15), Cdc2 and Cdc25C. PAR depletion also was found to result in inhibition of procaspases 9, 8, 6 and 3 with significant increase in the ratio of Bax : Bcl-2.

CONCLUSIONS

Our data indicate that PAR depletion induces G2/M arrest via the Cdc25C-Cdc2/cyclin B1 pathway. Furthermore, the results of the present study point toward involvement of pathways mediated by both caspase 8 and caspase 9 in apoptosis induction by PAR depletion.