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Epigenetic control of trefoil factor family (TFF) peptide expression in human retinoblastoma cell lines
Claudia Philippeit1, Maike Busch, Nicole Dünker
1University of Duisburg-Essen, Medical Faculty, Institute of Anatomy, Department of Neuroanatomy, Essen, Germany.
Background:
Recent studies demonstrated that epigenetic mechanisms are involved in the regulation of trefoil factor family (TFF) peptide expression in cancer. In human tissues with endogenous TFF1, TFF2 or TFF3 gene expression, the corresponding promoter is unmethylated and in organs without TFF expression, the promoter of the three genes is highly methylated.
Methods:
Retinoblastoma (Rb) cell lines were treated with the DNA methyltransferase inhibitor 5-Aza-2`deoxycytidine (5-Aza-dC), the histone deacetylase inhibitor 4-Phenylbutyric acid (PBA) or both and analyzed for changes (i) in TFF mRNA expression by Real-time PCR and (ii) in the methylation status of the TFF promoters by genomic bisulfite sequencing.
Results:
The degree of promoter methylation correlates with endogenous TFF expression in the retinoblastoma cell lines analyzed. Nearly all Rb cell lines exhibiting high endogenous TFF1 expression displayed low methylation of the CpGs in the corresponding promoter region. Low expression of TFF3 in Rb cell lines is linked with high density methylation of the TFF3 promoter. 5-Aza-dC treatment induced TFF1 and TFF3 expression in nearly all cell lines investigated and combined treatment with PBA further increased this effect. The number of methylated CpG dinucleotides of the TFF promoter is clearly reduced upon treatment with 5-Aza-dC and combined treatment with PBA further extended the degree of demethylation.
Conclusion:
Our data clearly show that the expression of TFF3 in retinoblastoma cell lines is epigenetically regulated, whereas the level of TFF1 and TFF2 seems to be regulated by other or additional mechanisms.
Insights
Epigenetic regulation influences trefoil factor family (TFF) peptide expression in retinoblastoma. DNA methylation changes impact TFF3 expression, while TFF1 and TFF2 may involve additional regulatory mechanisms.
Area of Science:
- Molecular Biology
- Epigenetics
- Cancer Research
Background:
- Epigenetic mechanisms, including DNA methylation, are increasingly recognized for their role in regulating gene expression in cancer.
- Trefoil factor family (TFF) peptides (TFF1, TFF2, TFF3) are implicated in various biological processes, including cancer development and progression.
- Endogenous expression of TFF genes in human tissues correlates with the methylation status of their respective promoters.
Purpose of the Study:
- To investigate the epigenetic regulation of TFF peptide expression in retinoblastoma (Rb) cell lines.
- To determine the impact of DNA methyltransferase and histone deacetylase inhibitors on TFF mRNA expression and promoter methylation in Rb cells.
Main Methods:
- Retinoblastoma cell lines were treated with 5-Aza-2`deoxycytidine (5-Aza-dC), a DNA methyltransferase inhibitor, and/or 4-Phenylbutyric acid (PBA), a histone deacetylase inhibitor.
- Changes in TFF mRNA expression were quantified using Real-time PCR.
- Alterations in TFF promoter methylation status were assessed through genomic bisulfite sequencing.
Main Results:
- A strong correlation was observed between TFF promoter methylation and endogenous TFF expression in Rb cell lines.
- High endogenous TFF1 expression was associated with low promoter methylation, while low TFF3 expression correlated with high promoter methylation.
- Treatment with 5-Aza-dC alone or in combination with PBA significantly induced TFF1 and TFF3 expression.
- 5-Aza-dC treatment, particularly when combined with PBA, led to a marked reduction in TFF promoter CpG methylation.
Conclusions:
- The expression of TFF3 in retinoblastoma cell lines is epigenetically regulated, primarily through DNA methylation.
- The regulation of TFF1 and TFF2 expression in retinoblastoma appears to involve additional or alternative epigenetic mechanisms beyond promoter methylation.
- These findings highlight the potential of epigenetic therapies targeting DNA methylation for modulating TFF peptide expression in retinoblastoma.
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