Ornithine Decarboxylase Antizyme Induces Hypomethylation of Genome DNA and Histone H3 Lysine 9 Dimethylation (H3K9me2) in Human Oral Cancer Cell Line

Daisuke Yamamoto,Kaori Shima,Guo-Fu Hu,Akira Sasaki,Takanori Tsuji,Chang Yan Chen,Takashi Nishioka,Kou Matsuo,Shuang-Yong Xu

doi:10.1371/journal.pone.0012554

Abstract

BackgroundMethylation of CpG islands of genome DNA and lysine residues of histone H3 and H4 tails regulates gene transcription. Inhibition of polyamine synthesis by ornithine decarboxylase antizyme-1 (OAZ) in human oral cancer cell line resulted in accumulation of decarboxylated S-adenosylmethionine (dcSAM), which acts as a competitive inhibitor of methylation reactions. We anticipated that accumulation of dcSAM impaired methylation reactions and resulted in hypomethylation of genome DNA and histone tails.Methodology/Principal FindingsGlobal methylation state of genome DNA and lysine residues of histone H3 and H4 tails were assayed by Methylation by Isoschizomers (MIAMI) method and western blotting, respectively, in the presence or absence of OAZ expression. Ectopic expression of OAZ mediated hypomethylation of CpG islands of genome DNA and histone H3 lysine 9 dimethylation (H3K9me2). Protein level of DNA methyltransferase 3B (DNMT3B) and histone H3K9me specific methyltransferase G9a were down-regulated in OAZ transfectant.Conclusions/SignificanceOAZ induced hypomethylation of CpG islands of global genome DNA and H3K9me2 by down-regulating DNMT3B and G9a protein level. Hypomethylation of CpG islands of genome DNA and histone H3K9me2 is a potent mechanism of induction of the genes related to tumor suppression and DNA double strand break repair.

Highlights

Ornithine decarboxylase (ODC; EC4.1.1.17) is the first and rate-limiting key enzyme for polyamine biosynthesis by catalyzing from L-ornithine to putrescine [1] and is implicated in cell proliferation and differentiation [2]
We examined the global methylation state of genome DNA and lysine residues from histone tails in human oral cancer cells in the absence or presence of ornithine decarboxylase antizyme-1 (OAZ) expression
Inhibition of polyamine synthesis by OAZ accumulated abnormally high level of decarboxylated S-adenosylmethionine (dcSAM), which acts as a competitive inhibitor of S-adenosylmethionine virtually in all methylation reactions

Summary

Introduction

Ornithine decarboxylase (ODC; EC4.1.1.17) is the first and rate-limiting key enzyme for polyamine biosynthesis by catalyzing from L-ornithine to putrescine [1] and is implicated in cell proliferation and differentiation [2]. OAZ is known to bind various proteins and mediate degradation or stabilization of the target proteins These proteins include ODC [5], antizyme inhibitor [6], cyclin D1 [7,8] and HPV16 E2 [9]. Inhibition of ODC activity and the subsequent polyamine synthesis by OAZ [10] or chemical compound, a-difluoromethylornithine (DFMO) [5] resulted in accumulation of dcSAM. DcSAM serves as an aminopropyl donor for polyamine synthesis but it acts as a competitive inhibitor of Sadenosylmethionine, virtually in all methylation reactions including DNA, RNA, lipid and protein [5,11]. Inhibition of polyamine synthesis by ornithine decarboxylase antizyme-1 (OAZ) in human oral cancer cell line resulted in accumulation of decarboxylated S-adenosylmethionine (dcSAM), which acts as a competitive inhibitor of methylation reactions. We anticipated that accumulation of dcSAM impaired methylation reactions and resulted in hypomethylation of genome DNA and histone tails

Methods

Results

Conclusion