Abstract LB-179: The effects of hypoxia and reactive oxygen species on the methylation status of the polo-like kinases

Abstract

Abstract The family of polo-like kinases (Plks) consisting of five known members are key regulators of important cell cycle processes, such as mitotic entry, centrosome duplication, spindle assembly, and cytokinesis. Plk1 is typically overexpressed in many cancers, while Plks 2–4 deregulation is often associated with tumorigenesis and malignancy. Plk2 has been shown to be downregulated via promoter DNA hypermethylation in blood neoplasms. Furthermore, in hepatocellular carcinoma (HCC), we and others have shown Plks 2–4 are downregulated through epigenetic modifications, while concurrent decreased Plk 1 promoter methylation was correlated with increased expression. As changes in epigenetic modifications have been associated with external environmental stimuli, we hypothesized that the Plks may be important targets whose disregulation could contribute to malignancy. We therefore examined the status of promoter methylation and expression levels of the Plk family members under hypoxic conditions, and upon exposure to reactive oxygen species (ROS). Interestingly both wild-type and Plk4 heterozygous murine embryonic fibroblasts (MEFs) exposed to ROS displayed significant promoter methylation post treatment. Correspondingly, Plk4 transcript levels were reduced to 90% with no detectable Plk4 protein after exposure to ROS. Since ROS and hypoxic conditions are associated with liver function and normal metabolism, we next examined the promoter methylation status for the individual Plks and other markers in HepG2 and Hep3B liver cell lines. We found increased hypermethylation of the promoter region as well as decreased transcripts levels for both Plk1 and Plk4 post ROS treatment in both cell lines. No detectable methylation changes were detected in SAOS (bone sarcoma, p53 null) cells, while U2OS (p53 wild type) displayed increased methylation and decreased transcript levels. This suggests that the response to ROS might be dependent upon p53 status and origin of cell line. Similar to the case with ROS, hypoxia treatment also induced hypermethylation of Plk4 and decreased in transcripts in both HepG2 and Hep3B cells, while Plk1 levels were only reduced in HepG2 cells. Our results suggest that changes in the cellular environment may be a key regulator of Plk methylation status that in turn leads to deregulation. This has important implications in HCC and other cancers, where these epigenetic alterations could be contributing factors in tumourigenesis and disease progression. 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 LB-179. doi:10.1158/1538-7445.AM2011-LB-179