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Methylation silencing of the Apaf-1 gene in acute leukemia
Yutaka Furukawa1, Krittaya Sutheesophon, Taeko Wada
1Division of Stem Cell Regulation, Center for Molecular Medicine, Jichi Medical School, 3311-1 Yakushiji, Minamikawachi-machi, Tochigi 329-0498, Japan.
Abstract:
Apaf-1 is important for tumor suppression and drug resistance because it plays a central role in DNA damage-induced apoptosis. Inactivation of the Apaf-1 gene is implicated in disease progression and chemoresistance of some malignancies. In this study, we attempted to clarify the role of Apaf-1 in leukemogenesis. Apaf-1 mRNA levels were below the detection limit or very low in 5 of 20 human leukemia cell lines (25%) and 5 of 12 primary acute myeloblastic leukemia cells (42%). There were no gross structural abnormalities in the Apaf-1 gene in these samples. Expression of factors regulating Apaf-1 transcription, such as E2F-1, p53, and Sp-1, did not differ between Apaf-1-positive and Apaf-1-negative cells. Methylation of CpG in the region between +87 and +128 of the Apaf-1 gene was almost exclusively observed in Apaf-1-defective cell lines. Treatment of these cells with 5-aza-2'-deoxycytidine, a specific inhibitor of DNA methylation, restored the expression of Apaf-1. Furthermore, we showed that the region between +87 and +128 could act as a repressor element by recruiting corepressors such as methylated DNA-binding domain 2 and histone deacetylase 1 upon methylation. Overexpression of Dnmt1, a mammalian maintenance DNA methyltransferase, was associated with Apaf-1 gene methylation. DNAs from Dnmt1-overexpressing cells were more resistant to digestion with methylation-sensitive enzyme HpaII than those from cells with low Dnmt1 expression, suggesting that Dnmt1 mediates aberrant methylation of multiple genes. In conclusion, methylation silencing is a mechanism of the inactivation of Apaf-1 in acute leukemia, and Dnmt1 overexpression may underlie hypermethylation of the Apaf-1 gene.
Insights
Apoptotic factor Apaf-1 (Apoptotic protease activating factor 1) is silenced in leukemia via DNA methylation. This mechanism, potentially driven by Dnmt1 overexpression, contributes to cancer progression and chemoresistance.
Area of Science:
- Molecular Biology
- Cancer Biology
- Epigenetics
Background:
- Apoptotic protease activating factor 1 (Apaf-1) is crucial for tumor suppression and drug resistance through its role in DNA damage-induced apoptosis.
- Inactivation of the Apaf-1 gene is linked to disease progression and chemoresistance in certain cancers.
Purpose of the Study:
- To investigate the role of Apaf-1 in leukemogenesis.
- To elucidate the mechanisms underlying Apaf-1 gene inactivation in acute leukemia.
Main Methods:
- Analysis of Apaf-1 mRNA levels in leukemia cell lines and primary acute myeloid leukemia cells.
- Investigation of Apaf-1 gene structure and regulatory factor expression (E2F-1, p53, Sp-1).
- Assessment of DNA methylation in the Apaf-1 gene promoter region and the effect of DNA methylation inhibitors (5-aza-2'-deoxycytidine).
- Evaluation of the role of DNA methyltransferase 1 (Dnmt1) in Apaf-1 gene methylation.
Main Results:
- Reduced Apaf-1 mRNA levels were observed in 25% of leukemia cell lines and 42% of primary acute myeloid leukemia samples.
- No gross structural abnormalities were found in the Apaf-1 gene.
- CpG methylation in a specific region (+87 to +128) of the Apaf-1 gene correlated with Apaf-1 deficiency.
- DNA methylation inhibitor treatment restored Apaf-1 expression.
- The +87 to +128 region acted as a methylation-dependent repressor element, recruiting corepressors like MBD2 and HDAC1.
- Overexpression of Dnmt1 was associated with Apaf-1 gene hypermethylation and resistance to methylation-sensitive enzyme digestion.
Conclusions:
- Methylation-induced silencing is a key mechanism for Apaf-1 inactivation in acute leukemia.
- Overexpression of Dnmt1 may drive the aberrant hypermethylation of the Apaf-1 gene, contributing to leukemogenesis and chemoresistance.
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