Lineage-specific silencing of human IL-10 gene expression by promoter methylation in cervical cancer cells

Anita Szalmás, Ferenc Bánáti, Anita Koroknai, Brigitta László, Eniko Fehér, D. Salamon, L. Gergely, J. Mináróvits, József Kónya

Research output: Contribution to journalArticle

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Abstract

Epigenetic analysis was performed to demonstrate that the normal and neoplastic epithelial cells do not serve as the source of the locally elevated IL-10 production during cervical carcinogenesis. Bisulfite sequencing was used to correlate promoter CpG methylation with the transcription of the gene. Lack of IL-10 transcription in HeLa, SiHa, Caski, HT-3, C33-A, HaCaT cell lines and in primary human keratinocytes correlated consistently with the methylated state of the proximal CpG residues, particularly with the two most proximal CpGs at positions -185 and -110. These two sites were also highly methylated in normal and malignant cervical cells directly isolated from patient material. On the other hand, IL-10 producing peripheral blood mononuclear cells had unmethylated CpG residues in the proximal promoter associated with acetylated H3 and H4 histones as determined by chromatin immunoprecipitation. In HeLa carrying epigenetically silenced endogeneous IL-10 promoter, the transfected non-CpG methylated 1 kb and 0.6 kb proximal promoter fragments could drive reporter gene expression, which was reversed by cassette methylation of these promoter fragments. In conclusion, the CpG methylation pattern of the proximal promoter is implicated as a major determinant of transcriptional silencing of human IL-10 expression in cells of cervical epithelial origin.

Original languageEnglish
Pages (from-to)1030-1038
Number of pages9
JournalEuropean Journal of Cancer
Volume44
Issue number7
DOIs
Publication statusPublished - May 2008

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Uterine Cervical Neoplasms
Interleukin-10
Methylation
Gene Expression
Histones
Epithelial Cells
Chromatin Immunoprecipitation
Keratinocytes
Reporter Genes
Epigenomics
Blood Cells
Carcinogenesis
Cell Line
Genes

Keywords

  • Cervical cancer
  • Epithelial cell
  • Gene silencing
  • Methylation cassette assay
  • Promoter methylation

ASJC Scopus subject areas

  • Cancer Research
  • Hematology
  • Oncology

Cite this

Lineage-specific silencing of human IL-10 gene expression by promoter methylation in cervical cancer cells. / Szalmás, Anita; Bánáti, Ferenc; Koroknai, Anita; László, Brigitta; Fehér, Eniko; Salamon, D.; Gergely, L.; Mináróvits, J.; Kónya, József.

In: European Journal of Cancer, Vol. 44, No. 7, 05.2008, p. 1030-1038.

Research output: Contribution to journalArticle

Szalmás, Anita ; Bánáti, Ferenc ; Koroknai, Anita ; László, Brigitta ; Fehér, Eniko ; Salamon, D. ; Gergely, L. ; Mináróvits, J. ; Kónya, József. / Lineage-specific silencing of human IL-10 gene expression by promoter methylation in cervical cancer cells. In: European Journal of Cancer. 2008 ; Vol. 44, No. 7. pp. 1030-1038.
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AB - Epigenetic analysis was performed to demonstrate that the normal and neoplastic epithelial cells do not serve as the source of the locally elevated IL-10 production during cervical carcinogenesis. Bisulfite sequencing was used to correlate promoter CpG methylation with the transcription of the gene. Lack of IL-10 transcription in HeLa, SiHa, Caski, HT-3, C33-A, HaCaT cell lines and in primary human keratinocytes correlated consistently with the methylated state of the proximal CpG residues, particularly with the two most proximal CpGs at positions -185 and -110. These two sites were also highly methylated in normal and malignant cervical cells directly isolated from patient material. On the other hand, IL-10 producing peripheral blood mononuclear cells had unmethylated CpG residues in the proximal promoter associated with acetylated H3 and H4 histones as determined by chromatin immunoprecipitation. In HeLa carrying epigenetically silenced endogeneous IL-10 promoter, the transfected non-CpG methylated 1 kb and 0.6 kb proximal promoter fragments could drive reporter gene expression, which was reversed by cassette methylation of these promoter fragments. In conclusion, the CpG methylation pattern of the proximal promoter is implicated as a major determinant of transcriptional silencing of human IL-10 expression in cells of cervical epithelial origin.

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