p53 dephosphorylation and p21^Cip1/Waf1 translocation correlate with caspase-3 activation in TGF-β1-induced apoptosis of HuH-7 cells

The p53 tumor suppressor gene product plays an important role in the regulation of apoptosis. Transforming growth factor β1 (TGF-β1)- induced apoptosis in hepatic cells is associated with reduced expression of the retinoblastoma protein (pRb) and subsequent E2F-1-activated expression of apoptosis-related genes. In this study, we explored the potential role of p53 in TGF-β1-induced apoptosis. HuH-7 human hepatoma cells were either synchronized in G₁, S and G₂/M phases, or treated with 1 nM TGF-β1. The results indicated that greater than 90% of the TGF-β1 -treated cells were arrested in G1 phase of the cell cycle. This was associated with enhanced p53 dephosphorylation and p21^Cip1/Waf1 expression, which coincided with decreased Cdk2, Cdk4, and cyclin E expression, compared with synchronized G₁ cells. In addition, p53 dephosphorylation coincided with caspase-3 activation, and translocation of p21 ^Cip1/Waf1 and p27^Kip1 into the cytoplasm, all of which were suppressed by caspase inhibition of TGF-β1-induced apoptosis. Finally, phosphatase inhibition and pRb overexpression partially inhibited p53-mediated apoptosis. In conclusion, the results demonstrated that TGF-β1-induced p53 dephosphorylation is associated with caspase-3 activation, and cytosolic translocation of p21^Cip1/Waf1 and p27^Kip1, resulting in decreased expression of Cdks and cyclins. Further, p53 appears to mediate TGF-β1-induced apoptosis downstream of the pRb/E2F-1 pathway.