Methylation silencing of the Apaf-1 gene in acute leukemia.
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.
Division of Stem Cell Regulation, Center for Molecular Medicine, Jichi Medical School, 3311-1 Yakushiji, Minamikawachi-machi, Tochigi 329-0498, Japan.
Yutaka
Y
Furukawa
Krittaya
K
Sutheesophon
Taeko
T
Wada
Miki
M
Nishimura
Yasushi
Y
Saito
Hideshi
H
Ishii
Yusuke
Y
Furukawa
D020121:5' Untranslated Regions; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D053477:Apoptotic Protease-Activating Factor 1; D001374:Azacitidine / Q000031:analogs & derivatives / Q000494:pharmacology; D045744:Cell Line, Tumor; D018899:CpG Islands / Q000235:genetics; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000378:metabolism; D019175:DNA Methylation; D000077209:Decitabine; D015972:Gene Expression Regulation, Neoplastic; D020868:Gene Silencing; D016147:Genes, Tumor Suppressor; D018922:HL-60 Cells; D056572:Histone Deacetylase Inhibitors; D006801:Humans; D019169:Jurkat Cells; D020014:K562 Cells; D015470:Leukemia, Myeloid, Acute / Q000235:genetics* / Q000473:pathology / Q000628:therapy; D011401:Promoter Regions, Genetic; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D020298:U937 Cells
D020121:5' Untranslated Regions; C506592:APAF1 protein, human; D000964:Antimetabolites, Antineoplastic; D053477:Apoptotic Protease-Activating Factor 1; D056572:Histone Deacetylase Inhibitors; D011506:Proteins; D012333:RNA, Messenger; D000077209:Decitabine; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases; C000619380:DNMT1 protein, human; D001374:Azacitidine