Replication stress, defective S-phase checkpoint and increased death in Plk2-deficient human cancer cells.
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. Our goals have been to better understand why Plk2 is regulated by p53 and how Plk2 signals protection from cell death through checkpoint activation. We found that following knock-down 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 and a further increase after release 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 replication stress. Consistent with this hypothesis, we observed higher levels of Serine 139 H2AX phosphorylation in Plk2-deficient as compared to control cells before and after aphidicolin treatment indicating that 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 Serine 317-phosphorylated Chk1. In aphidicolin-treated cells, there were lower levels of Serine 317-phosphorylated Chk1 when Plk2 was knocked-down. Plk2 was demonstrated to interact with Chk2, Chk1, Serine 317-phosphorylated Chk1 and p53. Thus, increased cell death observed after aphidicolin treatment and release in Plk2-deficient cells may result from both higher levels of replication stress-induced DNA damage and a dysfunctional S-phase checkpoint.
Matthew
Elizabeth M
EM
Department of Medicine, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104, USA.
Yen
Timothy J
TJ
Dicker
David T
DT
Dorsey
Jay F
JF
Yang
Wensheng
W
Navaraj
Arunasalam
A
El-Deiry
Wafik S
WS
D045744:Cell Line, Tumor; D000071877:Checkpoint Kinase 1; D004247:DNA / Q000235:genetics*; D004261:DNA Replication / Q000235:genetics*; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D009369:Neoplasms / Q000201:enzymology* / Q000235:genetics / Q000473:pathology*; D010768:Phosphoserine / Q000378:metabolism; D011485:Protein Binding; D011494:Protein Kinases / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000172:deficiency* / Q000235:genetics / Q000378:metabolism*; D016196:S Phase; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism
D016159:Tumor Suppressor Protein p53; D010768:Phosphoserine; D004247:DNA; D011494:Protein Kinases; C496126:PLK2 protein, human; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; D017346:Protein Serine-Threonine Kinases