Abstract
BackgroundAlterations in the processing of the genetic information in carcinogenesis result from stable genetic mutations or epigenetic modifications. It is becoming clear that nucleosomal histones are central to proper gene expression and that aberrant DNA methylation of genes and histone methylation plays important roles in tumor progression. To date, several histone lysine methyltransferases (HKMTs) have been identified and histone lysine methylation is now considered to be a critical regulator of transcription. However, still relatively little is known about the role of HKMTs in tumorigenesis.ResultsWe observed differential HKMT expression in a lung cancer model in which normal human bronchial epithelial (NHBE) cells expressing telomerase, SV40 large T antigen, and Ras were immortal, formed colonies in soft agar, and expressed specific HKMTs for H3 lysine 9 and 27 residues but not for H3 lysine 4 residue. Modifications in the H3 tails affect the binding of proteins to the histone tails and regulate protein function and the position of lysine methylation marks a gene to be either activated or repressed. In the present study, suppression by siRNA of HKMTs (EZH2, G9A, SETDB1 and SUV39H1) that are over-expressed in immortalized and transformed cells lead to reduced cell proliferation and much less anchorage-independent colony growth. We also found that the suppression of H3-K9, G9A and SUV39H1 induced apoptosis and the suppression of H3-K27, EZH2 caused G1 arrest.ConclusionOur results indicate the potential of these HKMTs in addition to the other targets for epigenetics such as DNMTs and HDACs to be interesting therapeutic targets.
Highlights
We found that the expression levels of H3-K4 MTs, SET9 and SMYD3 remained at similar ranges after introduction of large T antigen (LT) and activated H-Ras into normal human bronchial epithelial (NHBE) cells, while all of the other HKMTs, i.e.; H3-K9, H3-K27, H3-K79 HKMTs, increased after immortalization and transformation (Fig. 1b)
Suppression of H3-K9 or H3-K27 HKMT resulted in reduced cell proliferation in immortalized and transformed NHBE cells To evaluate the function of these HKMTs in the immortalized and the transformed cells, we treated transformed NHBE cells with siRNAs for 7 HKMTs, namely SET9, SMYD3, SETDB1, SUV39H, EZH2, G9A and DOT1L (Fig. 2)
The cell growth rates for both immortalized and transformed NHBE cells were significantly slowed by siRNA treatment of EZH2, G9A and SUV39H compared to the control oligonucleotide treatment, siRNA treatment of SET9, SMYD3, SETDB1 and DOT1L did not alter the cell growth rate (Fig. 2D and 2E)
Summary
Introduction ofSV40 large T antigen in NHBE cells renders over-expression of H3-K27 MT, EZH2. Alterations in the processing of the genetic information in carcinogenesis result from stable genetic mutations involving tumor suppressor genes, oncogenes and DNA stability genes as well as from potentially reversible epigenetic changes leading to modifications in gene function [1,2]. Repressive histone lysine methylation sites at H3-K9 and H3-K27 have been detected at the promoter regions of aberrantly silenced tumor suppressor genes in cancer cells, together with increased DNA methylation and reduced amounts of activating chromatin modifications such as histone acetylation [14,15]. It is apparent that certain histone lysine methylation, along with DNA methylation, establishes the framework for long-term epigenetic maintenance since recent studies have revealed a complex process that controls aspects of short- and long-term transcriptional regulation, in addition to the propagation of bulk chromosome structure and stability [16]
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