The effects of brefeldin A (BFA) on cell cycle progression involving the modulation of the retinoblastoma protein (pRB) in PC-3 prostate cancer cells.
OBJECTIVE
To investigate the effects of brefelding A (BFA) on the growth of the androgen-independent human prostate cancer PC-3 cells, focusing on cell cycle regulation.


METHODS
BFA is a fungal macrocyclic lactone with an antiviral activity. PC-3 cells were cultured with various concentrations of BFA for indicated times and cell growth was monitored at each time point. Cell cycle analysis was performed to explore the mechanism of BFA-induced growth inhibition. To further investigate the cell cycle regulation, cell cycle-controlling factors, such as the retinoblastoma gene product (pRB) and its regulatory components cdk2, cdk4, and cyclin D1, were analyzed by Western immunoblots.


RESULTS
BFA was a potent growth inhibitor at a concentration of 30 ng./ml., resulting in a > 70% reduction in cell number at 3 days. Cell cycle analysis revealed a cell arrest in the G1 to S phase transition. Western blots further showed that BFA induced dephosphorylation of pRB accompanied by down regulation of cdk2, cdk4, and cyclin D1 expression. The extended pRB dephosphorylation in control cell lysates was also observed by the addition of BFA-treated lysates, but was prevented by the inclusion of phosphatase inhibitors in assay mixtures.


CONCLUSIONS
These results suggest that BFA may be a potent cell cycle modulator, which post-translationally regulates pRB phosphorylation possibly by down-regulating cdk2, cdk4, and cyclin D1 and/or by up-regulating a phosphatase(s) capable of dephosphorylating pRB. Thus, BFA-induced growth inhibition in PC-3 cells appears to be at least partially due to the modulation of a pRB-mediated growth pathway.
Mordente
J A
JA
Department of Urology, New York Medical College, Valhalla 10595, USA.
Konno
S
S
Chen
Y
Y
Wu
J M
JM
Tazaki
H
H
Mallouh
C
C
D020126:Brefeldin A; D002455:Cell Division / Q000187:drug effects; D018844:Cyclin-Dependent Kinases / Q000187:drug effects* / Q000378:metabolism; D003517:Cyclopentanes / Q000494:pharmacology*; D016193:G1 Phase / Q000187:drug effects*; D006801:Humans; D008297:Male; D010766:Phosphorylation; D011471:Prostatic Neoplasms / Q000378:metabolism / Q000473:pathology*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology*; D016160:Retinoblastoma Protein / Q000378:metabolism*; D016196:S Phase / Q000187:drug effects*; D013997:Time Factors; D014407:Tumor Cells, Cultured / Q000187:drug effects
D003517:Cyclopentanes; D011500:Protein Synthesis Inhibitors; D016160:Retinoblastoma Protein; D020126:Brefeldin A; D018844:Cyclin-Dependent Kinases