Influence of CpG island methylation status in O6-metnylguanine- DNA methyltransferase expression of oral cancer cell lines

Jun Murakami, Jun-Ichi Asaumi, Yuu Maki, Hidetsugu Tsujigiwa, Hitoshi Nagatsuka, Susumu Kokeguchi, Tetsuyoshi Inoue, Shoji Kawasaki, Noriaki Tanaka, Donald MacPhee, Nagahide Matsubara, Kanji Kishi

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Abstract

It is known that the O6-methylguanine-DNA methyltransferase (MGMT) gene is susceptible to epigenetic regulation associated with an altered frequency of CpG methylation. To investigate whether epigenetic regulation of the MGMT gene might lead to significant reductions in the expression levels of cancer cells, we sought evidence of a link between the methylation status of the MGMT promoter and the expression levels of seven human oral cancer cell lines. We found two frequently methylated regions: the 5′ region extending from nt 690 to nt 893 in the promoter, and the more 3′ region extending from nt 1060 to nt 1151 in the untranslated first exon. The 3′ region was hypermethylated independently of MGMT expression levels in all cell lines. By contrast, in the three MGMT-downregulated cell lines (SAS, Hep2, HO-1-u-1), the levels of MGMT expression were inversely related to the density of 5′ region of the methylated CpGs in the MGMT promoter. Our results implied that the transcriptional inactivation of MGMT might require methylation of the 5' region, but not that of the 3′ region in oral cancer cell lines. We further explored the role of methylation in MGMT expression by treating cells with 5-Aza-2′-deoxycytidine (5Aza-dC). 5Aza-dC treatment led to the partial or complete cytosine demethylation of two frequently methylated MGMT regions in all cell lines. 5Aza-dC succeeded in upregulating of the MGMT mRNA levels in only 2 of 7 cell lines (HSC3 and HO-1-u-1), and in fact reduced MGMT mRNA in the other 5 cell lines. Furthermore, 5Aza-dC had an inhibitory effect on MGMT protein levels in all cell lines. Our results suggest that MGMT levels may not revert after 5Aza-dC treatment. Based on our findings, the regulation of MGMT expression appears to be more complex than previously thought, although it is at least partially influenced by CpG methylation. Accordingly, care should be taken interpreting the link between MGMT methylation and expression.

Original languageEnglish
Pages (from-to)339-345
Number of pages7
JournalOncology Reports
Volume12
Issue number2
Publication statusPublished - Aug 2004

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CpG Islands
Mouth Neoplasms
Methyltransferases
Methylation
decitabine
Cell Line
DNA
Epigenomics
Protein Methyltransferases
Messenger RNA
Cytosine
DNA Methylation

Keywords

  • CpG
  • Methylation
  • MGMT

ASJC Scopus subject areas

  • Cancer Research
  • Oncology

Cite this

Influence of CpG island methylation status in O6-metnylguanine- DNA methyltransferase expression of oral cancer cell lines. / Murakami, Jun; Asaumi, Jun-Ichi; Maki, Yuu; Tsujigiwa, Hidetsugu; Nagatsuka, Hitoshi; Kokeguchi, Susumu; Inoue, Tetsuyoshi; Kawasaki, Shoji; Tanaka, Noriaki; MacPhee, Donald; Matsubara, Nagahide; Kishi, Kanji.

In: Oncology Reports, Vol. 12, No. 2, 08.2004, p. 339-345.

Research output: Contribution to journalArticle

Murakami, J, Asaumi, J-I, Maki, Y, Tsujigiwa, H, Nagatsuka, H, Kokeguchi, S, Inoue, T, Kawasaki, S, Tanaka, N, MacPhee, D, Matsubara, N & Kishi, K 2004, 'Influence of CpG island methylation status in O6-metnylguanine- DNA methyltransferase expression of oral cancer cell lines', Oncology Reports, vol. 12, no. 2, pp. 339-345.
Murakami, Jun ; Asaumi, Jun-Ichi ; Maki, Yuu ; Tsujigiwa, Hidetsugu ; Nagatsuka, Hitoshi ; Kokeguchi, Susumu ; Inoue, Tetsuyoshi ; Kawasaki, Shoji ; Tanaka, Noriaki ; MacPhee, Donald ; Matsubara, Nagahide ; Kishi, Kanji. / Influence of CpG island methylation status in O6-metnylguanine- DNA methyltransferase expression of oral cancer cell lines. In: Oncology Reports. 2004 ; Vol. 12, No. 2. pp. 339-345.
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