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We previously reported that the Polo-like Kinase 2 gene (Plk2/Snk) is a direct target for transcriptional regulation by p53 and that silencing Plk2 sensitizes cancer cells to Taxol-induced apoptosis (Burns et al., Mol. Cell. Biol. 23:5556-5571, 2003). Our goals have been to better understand why Plk2 is regulated by p53 and how Plk2 signals protection from cell death through checkpoint activation. Surprisingly, we found that following knockdown of Plk2 in wild-type p53-expressing H460 human non-small cell lung cancer cells there was a significant increase in cell death observed in aphidicolin-treated cells. In plk2-deficient H460 cells there was a further increase in apoptosis observed after release of the cells from aphidicolin block. The highest levels of cell death were observed when plk2-deficient cells were released from both aphidicolin and etoposide treatment. These results suggested that a defective S-phase checkpoint may contribute to enhanced sensitivity of Plk2-deficient cells to the replication stress following Topoisomerase II inhibition. Consistent with this hypothesis we have observed higher levels of Ser139 H2AX phosphorylation in Plk2-deficient as compared to control cells. There were much higher levels of phosphorylated H2AX in the aphidicolin-treated Plk2-deficient cells as compared to control cells, consistent with the idea there is more DNA damage when Plk2 is depleted. We also observed higher levels of Chk1 protein in Plk2-deficient cells that were associated with reduced levels of Ser 317 phosphorylated Chk1. In aphidicolin-treated cells, there were lower levels of Ser 317-phosphorylated Chk1 when Plk2 was knocked down. Thus, the increased cell death we observed after aphidicolin treatment and release in the Plk2-deficient cells may be a result of both higher levels of replication stress-induced DNA damage as well as a dysfunctional S-phase checkpoint.

98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA