Hypermethylation of XIAP-associated factor 1, a putative tumor suppressor gene from the 17p13.2 locus, in human gastric adenocarcinomas.
X-linked inhibitor of apoptosis (XIAP) is the most potent member of the IAP family that exerts antiapoptotic effects by interfering with the activities of caspases. Recently, XIAP-associated factor 1 (XAF1) and two mitochondrial proteins, Smac/DIABLO and HtrA2, have been identified to negatively regulate the caspase-inhibiting activity of XIAP. To explore the candidacy of XAF1, Smac/DIABLO, and HtrA2 as a tumor suppressor in gastric tumorigenesis, we investigated the expression and mutation status of the genes in 123 gastric tissues and 15 cancer cell lines. Whereas Smac/DIABLO and HtrA2 transcripts were normally expressed in all cancer specimens we examined, XAF1 transcript was not expressed or present at extremely low levels in 40% (6 of 15) of cancer cell lines and in 23% (20 of 87) of primary carcinomas. Abnormal reduction of XAF1 expression showed a strong correlation with stage and grade of tumors, and a tumor-specific down-regulation of XAF1 was observed in 45% (9 of 20) of matched sets. Unlike XAF1, XIAP expression exhibited no detectable alteration in cancers. Whereas loss of heterozygosity within the XAF1 region or somatic mutations of the gene was not detected, expression of XAF1 transcript was reactivated in all nonexpressor cell lines after 5-aza-2-deoxycytidine treatment. The 5' upstream region of the XAF1 gene encompasses no gastric cell-rich region that rigorously satisfies the formal criteria for CpG islands. However, bisulfite DNA sequencing analysis for 34 CpG sites in the promoter region revealed a strong association between hypermethylation and gene silencing. Moreover, transcriptional silencing of XAF1 was tightly associated with hypermethylation of seven CpGs located in the 5' proximal region (nucleotides -23 to -234). Additionally, loss or abnormal reduction of XAF1 expression was found to inversely correlate with p53 mutations, suggesting that epigenetic inactivation of XAF1 and mutational alteration of p53 might be mutually exclusive events in gastric tumorigenesis. Collectively, our study suggests that epigenetic silencing of XAF1 by aberrant promoter methylation may contribute to the malignant progression of human gastric tumors.
Byun
Do-Sun
DS
Department of Pathology, School of Medicine, Kyung Hee University, Seoul 130-701, Korea.
Cho
Kyucheol
K
Ryu
Byung-Kyu
BK
Lee
Min-Goo
MG
Kang
Min-Ju
MJ
Kim
Hak-Ryul
HR
Chi
Sung-Gil
SG
D048868:Adaptor Proteins, Signal Transducing; D000230:Adenocarcinoma / Q000235:genetics* / Q000378:metabolism; D000483:Alleles; D051017:Apoptosis Regulatory Proteins; D002352:Carrier Proteins / Q000096:biosynthesis / Q000235:genetics; D045744:Cell Line, Tumor; D002886:Chromosomes, Human, Pair 17 / Q000235:genetics*; D019175:DNA Methylation; D015536:Down-Regulation; D017353:Gene Deletion; D015972:Gene Expression Regulation, Neoplastic; D020868:Gene Silencing; D016147:Genes, Tumor Suppressor; D000074923:High-Temperature Requirement A Serine Peptidase 2; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D024101:Mitochondrial Proteins / Q000096:biosynthesis / Q000235:genetics; D009154:Mutation; D009363:Neoplasm Proteins / Q000096:biosynthesis / Q000235:genetics*; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D012697:Serine Endopeptidases / Q000096:biosynthesis / Q000235:genetics; D013274:Stomach Neoplasms / Q000235:genetics* / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000235:genetics
D048868:Adaptor Proteins, Signal Transducing; D051017:Apoptosis Regulatory Proteins; D002352:Carrier Proteins; C412403:DIABLO protein, human; D047908:Intracellular Signaling Peptides and Proteins; D024101:Mitochondrial Proteins; D009363:Neoplasm Proteins; D012333:RNA, Messenger; D016159:Tumor Suppressor Protein p53; C417551:XAF1 protein, human; D012697:Serine Endopeptidases; C000615914:HTRA2 protein, human; D000074923:High-Temperature Requirement A Serine Peptidase 2