Abstract
We have previously shown that MBP-1 acts as a general transcriptional repressor, and forced expression of MBP-1 exerts an anti-proliferative effect on a number of human cancer cells. In this report, we have investigated the role of endogenous MBP-1 in cell growth regulation. For this, we generated human prostate cancer cells (PC3) stably transfected with short hairpin RNA targeting MBP-1. We have observed retarded growth and longer doubling time of MBP-1 knockdown PC3 cells as compared with control mock-transfected PC3 cells. Fluorescence-activated cell sorter analysis suggested that PC3 cells expressing MBP-1-specific small interfering RNA accumulated during G2/M phase of the cell cycle. Further analysis suggested that depletion of MBP-1 was associated with reduction of cyclin A and cyclin B1 expression when compared with that of the control cells. A delayed induction of cyclin A and B1 expression was observed in MBP-1-depleted PC3 cells (PC3-4.2) upon serum stimulation, although the level of expression was much lower than that of control PC3 cells. Supplementation of MBP-1 in PC3-4.2 cells restored cyclin A and cyclin B1 expression. Together, these results suggest that knockdown of MBP-1 in prostate cancer cells perturbs cell proliferation by inhibiting cyclin A and cyclin B1 expression.
Highlights
Progression of the cell cycle in eukaryotic cells is controlled by a series of protein complexes composed of cyclins and cyclindependent kinases (CDKs)2 [9]
Inhibition of MBP-1 by RNA Interference—We have identified two potent Small interfering RNA (siRNA) to transiently knock down endogenous MBP-1 in prostate cancer cells [6] and cloned these small hairpin RNAs (shRNAs) targeted to MBP-1 mRNA into a plasmid vector under the control of the H1 promoter (MBPsi-3 and MBPsi-4)
Depletion of MBP-1 Is Associated with Inhibition of Cyclins A and Cyclin B1—To unravel the mechanism of cell growth inhibition induced by MBP-1 depletion, we examined the expression of cyclins specific for G1, S, and G2/M phases of the cell cycle
Summary
Progression of the cell cycle in eukaryotic cells is controlled by a series of protein complexes composed of cyclins and cyclindependent kinases (CDKs)2 [9]. We have observed that depletion of MBP-1 in human prostate cancer cells resulted in inhibition of cyclin A and cyclin B1 expression and delayed cell cycle progression. MBP-1-targeted siRNA stably expressing PC3 clones exhibited a significantly slower rate of proliferation as compared with the control cells suggesting that depletion of MBP-1 inhibited cell proliferation.
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