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Concurrent epigenetic silencing of wnt/β-catenin pathway inhibitor genes in B cell chronic lymphocytic leukaemia
- Source :
- BMC Cancer, BMC Cancer, Vol 12, Iss 1, p 213 (2012)
- Publication Year :
- 2012
-
Abstract
- Background The Wnt/β-catenin signalling is aberrantly activated in primary B cell chronic lymphocytic leukaemia (CLL). Epigenetic silencing of pathway inhibitor genes may be a mechanism for its activation. In this study, we investigated systematically and quantitatively the methylation status of 12 Wnt/β-catenin pathway inhibitor genes – CDH1, DACT1, DKK1, DKK2, DKK3, DKK4, SFRP1, SFRP2, SFRP3, SFRP4, SFRP5 and WIF1 – in the cell lines EHEB and MEC-1 as well as patient samples. Methods Quantification of DNA methylation was performed by means of bisulphite pyrosequencing and confirmed by bisulphite Sanger sequencing. Gene expression was analysed by qPCR using GAPDH as internal control. E-cadherin and β-catenin protein quantification was carried out by microsphere-based immunoassays. Methylation differences observed between the patient and control groups were tested using generalised least squares models. Results For 10 genes, a higher methylation level was observed in tumour material. Only DKK4 exhibited similarly high methylation levels in both tumour and normal specimens, while DACT1 was always essentially unmethylated. However, also for these inhibitors, treatment of cells with the demethylating agent 5-aza-2´-deoxycytidine resulted in an induction of their expression, as shown by quantitative PCR, suggesting an indirect epigenetic control of activity. While the degree of demethylation and its transcriptional consequences differed between the genes, there was an overall high correlation of demethylation and increased activity. Protein expression studies revealed that no constitutive Wnt/β-catenin signalling occurred in the cell lines, which is in discrepancy with results from primary CLL. However, treatment with 5-aza-2´-deoxycytidine caused accumulation of β-catenin. Simultaneously, E-cadherin expression was strongly induced, leading to the formation of a complex with β-catenin and thus demonstrating its epigenetically regulated inhibition effect. Conclusions The results suggest an epigenetic silencing mechanism of the Wnt/β-catenin pathway inhibitor genes in CLL. Hypermethylation and silencing of functionally related genes may not be completely stochastic but result from the tumour epigenome reprogramming orchestrated by Polycomb-group repressive complexes. The data are of interest in the context of epigenetic-based therapy.
- Subjects :
- Cancer Research
Antimetabolites, Antineoplastic
Medizinische Fakultät -ohne weitere Spezifikation
DNA hypermethylation
lcsh:RC254-282
Epigenesis, Genetic
Epigenetic silencing
Cell Line, Tumor
Genetics
Humans
Wnt/β-catenin pathway
ddc:610
Gene Silencing
beta Catenin
B cell chronic lymphocytic leukaemia
Chromosome Mapping
DNA Methylation
β-catenin
lcsh:Neoplasms. Tumors. Oncology. Including cancer and carcinogens
Cadherins
Leukemia, Lymphocytic, Chronic, B-Cell
Inhibitor genes
Gene Expression Regulation, Neoplastic
Wnt Proteins
Oncology
Azacitidine
CpG Islands
Protein Binding
Signal Transduction
Research Article
Subjects
Details
- ISSN :
- 14712407
- Volume :
- 12
- Database :
- OpenAIRE
- Journal :
- BMC cancer
- Accession number :
- edsair.pmid.dedup....08a66e93a8fc1e6763e2e78ce333e85b