Abstract
The polo-like kinase (PLKs) family, consisting of five known members, are key regulators of important cell cycle processes, which include mitotic entry, centrosome duplication, spindle assembly, and cytokinesis. The PLKs have been implicated in a variety of cancers, such as hepatocellular carcinoma (HCC), with PLK1 typically overexpressed and PLKs 2–5 often downregulated. Altered expression of the PLKs in malignancy is often correlated with aberrant promoter methylation. Epigenetic marks are dynamic and can be modified in response to external environmental stimuli. The aim of our study was to determine if oxidative stress, a common feature of solid tumours, would induce changes to the promoter methylation of the PLKs resulting in changes in expression. We examined the promoter methylation status via MSP and subsequent expression levels of the PLK family members under exposure to hypoxic conditions or reactive oxygen species (ROS). Interestingly, murine embryonic fibroblasts exposed to hypoxia and ROS displayed significant hypermethylation of Plk1 and Plk4 promoter regions post treatment. Corresponding proteins were also depleted by 40% after treatment. We also examined the HCC-derived cell lines HepG2 and Hep3B and found that for PLK1 and PLK4, the increase in hypermethylation was correlated with the presence of functional p53. In p53 wild-type cells, HepG2, both PLK1 and PLK4 were repressed with treatment, while in the p53 null cell line, Hep3B, PLK4 protein was elevated in the presence of hypoxia and ROS. This was also the case for ROS-treated, p53 null, osteosarcoma cells, Saos-2, where the PLK4 promoter became hypomethylated and protein levels were elevated. Our data supports a model in which the PLKs are susceptible to epigenetic changes induced by microenvironmental cues and these modifications may be p53-dependent. This has important implications in HCC and other cancers, where epigenetic alterations of the PLKs could contribute to tumourigenesis and disease progression.
Highlights
The polo-like kinases (PLKs) have been implicated in a variety of solid and hematopoietic tumours, which include B-cell lymphoma, hepatocellular carcinoma (HCC), head and neck squamous carcinoma, colorectal cancers, and most recently gallbladder cancer, just to name a few [1,2,3,4,5]
The primary mediator of the cellular response to hypoxia is hypoxia inducible factor 1a (Hif1a) which is responsible for the transcriptional regulation of several key genes, such as vascular endothelial growth factor (VEGF) [23] and metabolic components such as nitric oxide (NO) which are important for the cellular adaptation to a hypoxic environment [24]
We have previously shown that Plk4 heterozygosity increases the susceptibility of Plk4 promoter methylation in an in vivo murine HCC model [10], we wanted to determine whether Plk4 heterozygosity impacted Plk promoter methylation under oxidative stress
Summary
The polo-like kinases (PLKs) have been implicated in a variety of solid and hematopoietic tumours, which include B-cell lymphoma, hepatocellular carcinoma (HCC), head and neck squamous carcinoma, colorectal cancers, and most recently gallbladder cancer, just to name a few [1,2,3,4,5]. The recently discovered PLK5, has tumour suppressor properties, and it is often hypermethylated in glioblastoma [12] Given that these kinases, which are highly conserved among species, play crucial roles in important cell cycle events such as spindle pole assembly, the DNA damage response, G2/M transitions, and cytokinesis [6,13,14], proper regulation of these proteins is essential for the maintenance of genomic integrity and the prevention of genomic instability. The underlying question is what is prompting the aberrant epigenetic regulation of the polo-like kinases in a variety of cancer types?
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