Abstract 2131: Plk1 decreases DNA double strand break (DSB) formation during replication- stalling genotoxic stress
Authors: Gebreselassie Daniel , Chun-Kost Gina , Saito Erika , Fernandez Alejandra , O'Brien Travis , Patierno Steven R. , Ceryak Susan M. 
Published: August 2012

Polo-like kinase 1 (Plk1) is a regulator of the mitotic checkpoint, and is involved in adaptation, which is the progression of cells into mitosis with damaged DNA. Deregulation of Plk1 activity is linked to cellular transformation. Recent studies suggest Plk1 activity to be critical for resumption of cell cycle progression during recovery from DNA damage. However, the ability of Plk1 to affect the DNA damage response remains relatively unclear. The aim of the present study was to determine the ability of Plk1 to affect the DNA damage response in the face of different forms of DSB-inducing genotoxic stress. We employed the respiratory carcinogen, hexavalent chromium [Cr(VI)], the anticancer drug and topoisomerase II inhibitor, etoposide, as well as the oxidizer, H2O2. We have previously shown that Plk1 activation was sufficient to bypass the G2/M checkpoint in normal human lung cells (HLFs), in the presence of Cr(VI)-induced acute genotoxic stress. In the present study, we measured DNA DSB induction in HLFs by the respective genotoxins under the condition of Plk1 activation by using the comet assay and γH2AX immunofluorescence staining. Transfection of HLFs with the constitutively active (c/a) Plk1 T210D mutant resulted in increased Plk1 protein expression 24h-48h post transfection. We used equitoxic concentrations of the different genotoxins and found that DNA DSBs increased &gt; 2 fold after exposure to 3 μM Cr(VI), 12.5 μM etoposide, and 150 μM H2O2, respectively, and persisted for at least 4h. Consistent with DNA DSB formation, Cr(VI) exposure was associated with G1 arrest, at least at 4h, as determined by BrdU incorporation. Expression of the c/a T210D mutant increased Plk1 activity in vitro, and abrogated both Cr(VI) and etoposide-induced DNA DSBs as early as 30 min, and up to 4h post-treatment, in comparison to the vector control-transfected cells. In sharp contrast, treatment with 150 μM H2O2 induced a similar level of DNA DSBs in both the c/a T210D mutant- and vector-transfected cells. These data highlight the ability of Plk1 to decrease DNA DSBs uniquely under conditions of replication-stalling genotoxic stress. Given the documented roles of aberrant DNA damage response and Plk1 activation in cellular transformation, there is a critical need to define the molecular mechanism(s) by which Plk1 mediates decreased DNA DSBs after genotoxin exposure and delineate the pathways responsible for Plk1 activation. We postulate that the ability of Plk1 activation to abrogate DNA DSB formation after either Cr(VI) or etoposide exposure may occur at the expense of genomic stability. Supported by NIH grants CA107972 and ES017334 to SC and ES09961 and ES05304 to SRP and PhRMA foundation to GC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2131. doi:1538-7445.AM2012-2131