Induction of apoptosis by the p53-related p73 and partial inhibition by the baculovirus-encoded p35 in neuronal cells.

To better understand whether the p53-related p73 gene could induce neuronal apoptosis, we tested whether p73 induced cell killing in three neuronal cell lines and whether apoptosis could be inhibited by p35, a baculovirus-encoded protein that blocks caspase 3. Recombinant adenoviruses carrying the hemagglutinin (HA)-tagged p73beta or p35, or the green fluorescent protein gene driven by the cytomegalovirus immediate-early promoter were constructed, and used to infect human SK-N-AS and SK-N-SH neuroblastoma, and rat PC12 pheochromocytoma cells. Infection with Adp73beta virus resulted in p73beta over-expression and substantial reduction of cell viability due to apoptosis in all three neuronal cell lines as compared with the control AdGFP virus. These results indicate that p73beta over-expression in neuronal cells could induce apoptotic cell death regardless of the endogenous expression of p73. The p73 effect was partially blocked by co-expression of the wild-type p35, suggesting caspase-mediated cell killing. Insertion of a hemagglutinin (HA) tag at the N-terminus of p35 markedly reduced its ability to inhibit the p73 effect compared with the wild-type p35, while insertion of an HA tag to the C-terminus of p35 had no appreciable effect. Taken together, our results suggest that the N-terminal structure of p35 is critical for its anti-apoptotic activity on p73-induced apoptosis in neuronal cells.