Regulation of DNA methylation and tumor suppression gene expression by miR-29b in leukemia patients and related mechanisms.

OBJECTIVE

Leukemia is characterized as a kind of malignant clonal disease in hematological stem cells. The study showed an abnormal level of DNA methylation in leukemia cells, which further led to an abnormal expression of hematological genes. This study investigated the role of miR-29b on the modulation of DNA methylation and tumor suppressor gene expression in leukemia patients.

PATIENTS AND

METHODS

A total of 21 leukemia patients were recruited for the collection of monocytes. Methylation levels of promoter sequence of ESR1 and p15 genes were analyzed by methylation assay kit combined with DHPLC. DNA microarray and qRT-PCR were used to measure microRNA expressional profile, and bioinformatics plus luciferase reporter assay confirmed target gene of miR-29b. After transfection with miR-29b, promoter methylation levels of ESR1 and p15 gene were measured. Protein expressions of DNMT1 DNA (cytosine-5)-methyltransferase 1 (DNMT1), DNA (cytosine-5)-methyltransferase 3A (DNMT3A) and DNA (cytosine-5)-methyltransferase 3B (DNMT3B) were quantified.

RESULTS

The methylation levels of the promoter region of ESR1 and p15 genes in monocytes of leukemia patient were significantly elevated (p < 0.05). DNA microarray and qRT-PCR confirmed the down-regulation of miR-29b (p < 0.05). Luciferase reporter assay revealed DNMT1, DNMT3A and DNMT3B as target genes of miR-29b. MiR-29b transfection inhibited the expressions of DNMT3A and DNMT3B in Kasumi-1 cells (p < 0.05), and promoter methylation levels of estrogen Receptor 1 (ESR1) and p15 gene were decreased (p < 0.05).

CONCLUSIONS

In leukemia cells, hyper- methylation existed in the promoter region of tumor suppressor gene. The methylation was enhanced in gene DNMT1, DNMT3A and DNMT3B via the reduction of miR-29b in leukemia tumor cells.