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Epigenetic regulations in the IFNγ signalling pathway: IFNγ-mediated MHC class I upregulation on tumour cells is associated with DNA demethylation of antigen-presenting machinery genes.

Vlková V, Štěpánek I, Hrušková V, Šenigl F, Mayerová V, Šrámek M, Šímová J, Bieblová J, Indrová M, Hejhal T, Dérian N, Klatzmann D, Six A, Reiniš M - Oncotarget (2014)

Bottom Line: Our data also documented higher methylation levels in these genes in TC-1/A9 cells, as compared to their parental MHC class I-positive TC-1 cells.IFNγ-mediated DNA demethylation was relatively fast in comparison with demethylation induced by DNA methyltransferase inhibitor 5-azacytidine, and associated with increased histone H3 acetylation in the promoter regions of APM genes.Our data demonstrate that IFNγ acts as an epigenetic modifier when upregulating the expression of antigen-presenting machinery genes.

View Article: PubMed Central - PubMed

Affiliation: Department of Tumour Immunology, Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, v. v. i., Prague.

ABSTRACT
Downregulation of MHC class I expression on tumour cells, a common mechanism by which tumour cells can escape from specific immune responses, can be associated with coordinated silencing of antigen-presenting machinery genes. The expression of these genes can be restored by IFNγ. In this study we documented association of DNA demethylation of selected antigen-presenting machinery genes located in the MHC genomic locus (TAP-1, TAP-2, LMP-2, LMP-7) upon IFNγ treatment with MHC class I upregulation on tumour cells in several MHC class I-deficient murine tumour cell lines (TC-1/A9, TRAMP-C2, MK16 and MC15). Our data also documented higher methylation levels in these genes in TC-1/A9 cells, as compared to their parental MHC class I-positive TC-1 cells. IFNγ-mediated DNA demethylation was relatively fast in comparison with demethylation induced by DNA methyltransferase inhibitor 5-azacytidine, and associated with increased histone H3 acetylation in the promoter regions of APM genes. Comparative transcriptome analysis in distinct MHC class I-deficient cell lines upon their treatment with either IFNγ or epigenetic agents revealed that a set of genes, significantly enriched for the antigen presentation pathway, was regulated in the same manner. Our data demonstrate that IFNγ acts as an epigenetic modifier when upregulating the expression of antigen-presenting machinery genes.

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IFNγ upregulation of the cell-surface MHC class I expression cells is associated with APM gene expression in experimental tumour cellsMHC class I expression (H-2Db and H-2Kb together) was determined by the FACS analysis of control tumour cells and after the treatment with IFNγ. Representative results are presented. (A) Upregulation of APM genes in TC-1/A9 and TRAMP-C2 tumours after treatment with IFNγ. (B) Expression levels of selected APM genes in TC-1/A9 and TRAMP-C2 control tumour cells and after the treatment with IFNγ. As a negative control, MHC class I-deficient RVP-3 cell line that did not respond to the IFNγ treatment was used. TC-1 cells, a MHC class I-positive parental cell line to the TC-1/A9 cells, that also displayed higher APM expression levels compared to TC-1/A9 cells, served as a MHC class I-positive control. *denote significant changes (P<0.05 determined in Student's t-test) as compared to the values for untreated cells. Biological triplicates were used for the analysis. In all experiments, error bars show standard deviations. Relative expression numbers represent the percentage of the β-actin expression levels. The levels of relative gene expression were presented as fold changes compared to the levels found in control samples. Experiments were repeated three times with similar results.
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Figure 1: IFNγ upregulation of the cell-surface MHC class I expression cells is associated with APM gene expression in experimental tumour cellsMHC class I expression (H-2Db and H-2Kb together) was determined by the FACS analysis of control tumour cells and after the treatment with IFNγ. Representative results are presented. (A) Upregulation of APM genes in TC-1/A9 and TRAMP-C2 tumours after treatment with IFNγ. (B) Expression levels of selected APM genes in TC-1/A9 and TRAMP-C2 control tumour cells and after the treatment with IFNγ. As a negative control, MHC class I-deficient RVP-3 cell line that did not respond to the IFNγ treatment was used. TC-1 cells, a MHC class I-positive parental cell line to the TC-1/A9 cells, that also displayed higher APM expression levels compared to TC-1/A9 cells, served as a MHC class I-positive control. *denote significant changes (P<0.05 determined in Student's t-test) as compared to the values for untreated cells. Biological triplicates were used for the analysis. In all experiments, error bars show standard deviations. Relative expression numbers represent the percentage of the β-actin expression levels. The levels of relative gene expression were presented as fold changes compared to the levels found in control samples. Experiments were repeated three times with similar results.

Mentions: First, we assessed the level of the MHC class I and APM molecule expression and its modulation by IFNγ on the selected tumour cells (Fig. 1a). MHC class I expression on tumour cells after IFNγ treatment was upregulated as compared to the tumour cells without treatment. As expected and as has been published previously [19,20], the MHC class I upregulation induced by IFNγ was associated with increased expression of APM genes (Fig. 1b). As a negative control, we used the MHC class I-deficient RVP-3 cell line that did not respond to the IFNγ treatment. TC-1 cells, an MHC class I-positive parental cell line to the TC-1/A9 cells, which also displayed higher APM expression levels, as compared to TC-1/A9 cells, served as an MHC class I-positive control.


Epigenetic regulations in the IFNγ signalling pathway: IFNγ-mediated MHC class I upregulation on tumour cells is associated with DNA demethylation of antigen-presenting machinery genes.

Vlková V, Štěpánek I, Hrušková V, Šenigl F, Mayerová V, Šrámek M, Šímová J, Bieblová J, Indrová M, Hejhal T, Dérian N, Klatzmann D, Six A, Reiniš M - Oncotarget (2014)

IFNγ upregulation of the cell-surface MHC class I expression cells is associated with APM gene expression in experimental tumour cellsMHC class I expression (H-2Db and H-2Kb together) was determined by the FACS analysis of control tumour cells and after the treatment with IFNγ. Representative results are presented. (A) Upregulation of APM genes in TC-1/A9 and TRAMP-C2 tumours after treatment with IFNγ. (B) Expression levels of selected APM genes in TC-1/A9 and TRAMP-C2 control tumour cells and after the treatment with IFNγ. As a negative control, MHC class I-deficient RVP-3 cell line that did not respond to the IFNγ treatment was used. TC-1 cells, a MHC class I-positive parental cell line to the TC-1/A9 cells, that also displayed higher APM expression levels compared to TC-1/A9 cells, served as a MHC class I-positive control. *denote significant changes (P<0.05 determined in Student's t-test) as compared to the values for untreated cells. Biological triplicates were used for the analysis. In all experiments, error bars show standard deviations. Relative expression numbers represent the percentage of the β-actin expression levels. The levels of relative gene expression were presented as fold changes compared to the levels found in control samples. Experiments were repeated three times with similar results.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4196173&req=5

Figure 1: IFNγ upregulation of the cell-surface MHC class I expression cells is associated with APM gene expression in experimental tumour cellsMHC class I expression (H-2Db and H-2Kb together) was determined by the FACS analysis of control tumour cells and after the treatment with IFNγ. Representative results are presented. (A) Upregulation of APM genes in TC-1/A9 and TRAMP-C2 tumours after treatment with IFNγ. (B) Expression levels of selected APM genes in TC-1/A9 and TRAMP-C2 control tumour cells and after the treatment with IFNγ. As a negative control, MHC class I-deficient RVP-3 cell line that did not respond to the IFNγ treatment was used. TC-1 cells, a MHC class I-positive parental cell line to the TC-1/A9 cells, that also displayed higher APM expression levels compared to TC-1/A9 cells, served as a MHC class I-positive control. *denote significant changes (P<0.05 determined in Student's t-test) as compared to the values for untreated cells. Biological triplicates were used for the analysis. In all experiments, error bars show standard deviations. Relative expression numbers represent the percentage of the β-actin expression levels. The levels of relative gene expression were presented as fold changes compared to the levels found in control samples. Experiments were repeated three times with similar results.
Mentions: First, we assessed the level of the MHC class I and APM molecule expression and its modulation by IFNγ on the selected tumour cells (Fig. 1a). MHC class I expression on tumour cells after IFNγ treatment was upregulated as compared to the tumour cells without treatment. As expected and as has been published previously [19,20], the MHC class I upregulation induced by IFNγ was associated with increased expression of APM genes (Fig. 1b). As a negative control, we used the MHC class I-deficient RVP-3 cell line that did not respond to the IFNγ treatment. TC-1 cells, an MHC class I-positive parental cell line to the TC-1/A9 cells, which also displayed higher APM expression levels, as compared to TC-1/A9 cells, served as an MHC class I-positive control.

Bottom Line: Our data also documented higher methylation levels in these genes in TC-1/A9 cells, as compared to their parental MHC class I-positive TC-1 cells.IFNγ-mediated DNA demethylation was relatively fast in comparison with demethylation induced by DNA methyltransferase inhibitor 5-azacytidine, and associated with increased histone H3 acetylation in the promoter regions of APM genes.Our data demonstrate that IFNγ acts as an epigenetic modifier when upregulating the expression of antigen-presenting machinery genes.

View Article: PubMed Central - PubMed

Affiliation: Department of Tumour Immunology, Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, v. v. i., Prague.

ABSTRACT
Downregulation of MHC class I expression on tumour cells, a common mechanism by which tumour cells can escape from specific immune responses, can be associated with coordinated silencing of antigen-presenting machinery genes. The expression of these genes can be restored by IFNγ. In this study we documented association of DNA demethylation of selected antigen-presenting machinery genes located in the MHC genomic locus (TAP-1, TAP-2, LMP-2, LMP-7) upon IFNγ treatment with MHC class I upregulation on tumour cells in several MHC class I-deficient murine tumour cell lines (TC-1/A9, TRAMP-C2, MK16 and MC15). Our data also documented higher methylation levels in these genes in TC-1/A9 cells, as compared to their parental MHC class I-positive TC-1 cells. IFNγ-mediated DNA demethylation was relatively fast in comparison with demethylation induced by DNA methyltransferase inhibitor 5-azacytidine, and associated with increased histone H3 acetylation in the promoter regions of APM genes. Comparative transcriptome analysis in distinct MHC class I-deficient cell lines upon their treatment with either IFNγ or epigenetic agents revealed that a set of genes, significantly enriched for the antigen presentation pathway, was regulated in the same manner. Our data demonstrate that IFNγ acts as an epigenetic modifier when upregulating the expression of antigen-presenting machinery genes.

Show MeSH
Related in: MedlinePlus