The Histone Demethylase JMJD2B Plays an Essential Role in Human Carcinogenesis through Positive Regulation of Cyclin-Dependent Kinase 6

Gouji Toyokawa,Tatsuhiko Tsunoda,Yusuke Nakamura,John D Kelly,Shinya Hayami,Yukiko Iwai,Masashi Takawa,Bruce A.J Ponder,Masanori Yoshimatsu,Kazuhiro Maejima,Noriaki Shimizu,Helen I Field,Yoshihiko Maehara,Hyun-Soo Cho,David E Neal,Ryuji Hamamoto,Hirotoshi Tanaka

doi:10.1158/1940-6207.capr-11-0290

Abstract

Histone methyltransferases and demethylases are known to regulate transcription by altering the epigenetic marks on histones, but the pathologic roles of their dysfunction in human diseases, such as cancer, still remain to be elucidated. Herein, we show that the histone demethylase JMJD2B is involved in human carcinogenesis. Quantitative real-time PCR showed notably elevated levels of JMJD2B expression in bladder cancers, compared with corresponding nonneoplastic tissues (P < 0.0001), and elevated protein expression was confirmed by immunohistochemistry. In addition, cDNA microarray analysis revealed transactivation of JMJD2B in lung cancer, and immunohistochemical analysis showed protein overexpression in lung cancer. siRNA-mediated reduction of expression of JMJD2B in bladder and lung cancer cell lines significantly suppressed the proliferation of cancer cells, and suppressing JMJD2B expression lead to a decreased population of cancer cells in S phase, with a concomitant increase of cells in G(1) phase. Furthermore, a clonogenicity assay showed that the demethylase activity of JMJD2B possesses an oncogenic activity. Microarray analysis after knockdown of JMJD2B revealed that JMJD2B could regulate multiple pathways which contribute to carcinogenesis, including the cell-cycle pathway. Of the downstream genes, chromatin immunoprecipitation showed that CDK6 (cyclin-dependent kinase 6), essential in G(1)-S transition, was directly regulated by JMJD2B, via demethylation of histone H3-K9 in its promoter region. Expression levels of JMJD2B and CDK6 were significantly correlated in various types of cell lines. Deregulation of histone demethylation resulting in perturbation of the cell cycle, represents a novel mechanism for human carcinogenesis and JMJD2B is a feasible molecular target for anticancer therapy.

Highlights

Covalent histone modifications, including acetylation, methylation, phosphorylation, ubiquitination, glycosylation, and sumoylation can modulate chromatin dynamics and affect multiple cellular functions [1,2,3]
Functional studies showed that JMJD2B contains hypoxia response elements in its promoter region and, JMJD2B is induced by Hypoxia-inducible factor (HIF; refs. 16, 17), but its significance in human diseases such as cancer remains to be elucidated
We showed significant upregulation of JMJD2B in various types of cancers, including bladder and lung, by quantitative realtime PCR, cDNA microarray, or immunohistochemistry

Summary

Introduction

Covalent histone modifications, including acetylation, methylation, phosphorylation, ubiquitination, glycosylation, and sumoylation can modulate chromatin dynamics and affect multiple cellular functions [1,2,3]. Among these modifications, histone methylation is associated with activated or repressed transcription [3]. The Jumonji C (JmjC) domain containing www.aacrjournals.org protein family, which catalyzes the hydroxylation of a lysine methyl group via a radical-based mechanism, has been identified as histone demethylases which differ from LSD1 [6, 7]. Our knowledge of the physiologic functions of histone demethylases is increasing, it still remains unclear how deregulation of the enzymes is involved in human diseases such as cancer

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Conclusion