Regulationsmechanismen des Interferon regulatorischen Faktors IRF-4 in der chronisch myeloischen Leukämie

There is mounting evidence that the deregulation a family of transcription factors, the interferon regulatory factors (IRFs),is implicated in the pathogenesis of CML. The expression of the interferon regulatory factor IRF-4 is downregulated in T cells of CML patients in chronic phase and its increase is associated with a good response to therapy with interferon alpha. Promoter methylation of CpG target sites or direct deletions/insertions of genes are mechanisms of a reversible or permanent deregulation of gene expression, respectively. Therefore, it was examined whether methylation or mutation of the IRF-4 promotor may be involved in the regulation of IRF-4 expression in leukemia cell lines. Whereas promoter mutations or structural rearrangements could be excluded as a cause of altered IRF-4 expression in hematopoietic cells, the IRF-4 promoter methylation status was found to significantly influence IRF-4 transcription. Treatment of IRF-4-negative lymphoid, myeloid and monocytic cell lines with the methylation-inhibitor 5-aza-2-deoxycytidine resulted in a time- and concentration-dependent increase of IRF-4 mRNA and protein levels. Using a restriction-PCR-assay and bisulfit-sequencing we identified specifically methylated CpG sites in IRF-4-negative but not in IRF-4-positive cells. An association of methylational status and mRNA expression of DNA methyltransferases (DNMT) or methyl-CpG-binding proteins (MBP)was not detected. Together, these data suggest CpG site-specific IRF-4 promoter methylation as a putative mechanism of deregulated IRF-4 expression in leukemia.