Abstract 3914: Plk1 enhances DNA double strand break (DSB) repair during acute genotoxin exposure

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL 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 in cancer cells during recovery from DNA damage. However, the ability of Plk1 to affect the DNA damage response remains relatively unclear. In this study we employed the respiratory carcinogen, hexavalent chromium [Cr(VI)], which is a well-documented genotoxin of occupational and environmental concern. 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 by Cr(VI) in HLFs under the condition of Plk1 activation by using the comet assay and gamma H2AX 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 found that DNA DSBs increased > 2 fold 15 min after 3 μM Cr(VI) exposure, 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 Cr(VI)-induced DNA DSBs as early as 30 min, and up to 4h post-treatment, in comparison to the vector control transfected cells. Since Cr(VI)-induced DNA DSBs were equally formed by 15 min in both vector- and Plk1 T210D-transfected cells, these data highlight the heretofore unreported ability of Plk1 to enhance DNA repair. Our previous report found that Plk1 expression increased survival and mutagenesis of wt S. cerevisiae treated with Cr(VI). Therefore, we also determined the ability of Plk1 to “rescue” repair deficient rad52 yeast from Cr(VI)-induced lethality. S. cerevisae transformed with Plk1-Gal-HA-EGFP plasmid grown under inducible conditions overexpress Plk1. We found that induction of Plk1 in rad52 yeast significantly protected them from Cr(VI) clonogenic lethality. 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 DNA damage repair after genotoxin exposure and delineate the pathways responsible for Plk1 activation. We postulate that the ability of Plk1 activation to provoke DNA DSB repair after Cr(VI) exposure occurs at the expense of genomic stability. Supported by NIH grants CA107972 to SC and ES09961 and ES05304 to SRP and PhRMA foundation to GC. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3914. doi:10.1158/1538-7445.AM2011-3914