[Effect of AML1-ETO Fusion Protein on Transcriptional Regulation of p14].

OBJECTIVE

To investigate the effect of transcriptional regulation of aberrant transcription factor AML1-ETO on p14.

METHODS

P14 expression both in AML1-ETO-expressing cells or U937 nonexpressing cells and in leukemia cells of AML patients with or without t(8;21) was assessed by quantitative PCR. Methylation-specific polymerase chain reaction (MSP) was used to analyze the methylation status of p14 promoter. The chromatin immunoprecipitation (ChIP)-based PCR was used to investigate the direct interaction between the AML1-ETO and p14 promoter in AML1-ETO positive leukemia cell line. And the p14 mRNA expression level was detected by qRT-PCR after treatment with 5-Aza.

RESULTS

AML1-ETO-expressing cell subclone displayed low level of p14 mRNA in comparison with the non-transfected U937. In primary bone marrow cells of acute myeloid leukemia containing AML1-ETO, level of p14 mRNA was markedly lower when compared with other acute myeloid leukemias lacking this translocation. P14 gene promoter was non-methylated in control group and primary leukemia cells of AML patients without t(8;21) and was hyper-methylated in U937-A/E1-4 and primary leukemia cells of AML patients with t(8;21). The enriched regions in transfected cells were located within p14 promoter. 5-Aza could increase the expression of p14.

CONCLUSION

P14 is a possible target gene of AML1-ETO. The p14 silencing induced by hyper-methlylation may be an important factor for occurrence and development of the M subtype of acute myeloid leukemia.