title: WIF1 is a frequent target for epigenetic silencing in squamous cell carcinoma of the cervix.
abstract: Aberrant activation of the Wnt/β-catenin signaling axis is a prominent oncogenic mechanism in numerous cancers including cervical cancer. Wnt inhibitory factor-1 (WIF1) is a secreted protein that binds Wnt and antagonizes Wnt activity. While the WIF1 gene is characterized as a target for epigenetic silencing in some tumor types, WIF1 expression has not been examined in human cervical tissue and cervical cancer. Here, we show that WIF1 is unmethylated and its gene product is expressed in normal cervical epithelium and some cultured cervical tumor lines. In contrast, several cervical cancer lines contained dense CpG methylation within the WIF1 gene, and expression of both WIF1 transcript and protein was restored by culturing cells in the presence of the global DNA demethylating agent 5-aza-2'-deoxycytidine. Using single-molecule MAPit methylation footprinting, we observed differences in chromatin structure within the WIF1 promoter region between cell lines that express and those that do not express WIF1, consistent with transcriptional activity and repression, respectively. The WIF1 promoter was aberrantly methylated in ∼60% (10 of 17) high-grade highly undifferentiated squamous cell cervical tumors examined, whereas paired normal tissue showed significantly lower levels of CpG methylation. WIF1 protein was not detectable by immunohistochemistry in tumors with quantitatively high levels of WIF1 methylation. Of note, WIF1 protein was not detectable in two of the seven unmethylated cervical tumors examined, suggesting other mechanisms may contribute WIF1 repression. Our findings establish the WIF1 gene as a frequent target for epigenetic silencing in squamous cell carcinoma of the cervix.
authors.affiliation: Department of Biochemistry and Molecular Biology and UF-Shands Cancer Center, University of Florida, Gainesville, FL 32610, USA.
authors.forename: Amber L
authors.initials: AL
authors.lastname: Delmas
authors.forename: Bridget M
authors.initials: BM
authors.lastname: Riggs
authors.forename: Carolina E
authors.initials: CE
authors.lastname: Pardo
authors.forename: Lisa M
authors.initials: LM
authors.lastname: Dyer
authors.forename: Russell P
authors.initials: RP
authors.lastname: Darst
authors.forename: Eugene G
authors.initials: EG
authors.lastname: Izumchenko
authors.forename: Mänette
authors.initials: M
authors.lastname: Monroe
authors.forename: Ardeshir
authors.initials: A
authors.lastname: Hakam
authors.forename: Michael P
authors.initials: MP
authors.lastname: Kladde
authors.forename: Erin M
authors.initials: EM
authors.lastname: Siegel
authors.forename: Kevin D
authors.initials: KD
authors.lastname: Brown
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics* / Q000378:metabolism; D001374:Azacitidine / Q000031:analogs & derivatives / Q000494:pharmacology; D015153:Blotting, Western; D002294:Carcinoma, Squamous Cell / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D002460:Cell Line; D002584:Cervix Uteri / Q000378:metabolism; D018899:CpG Islands / Q000235:genetics; D019175:DNA Methylation; D000077209:Decitabine; D044127:Epigenesis, Genetic; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D020868:Gene Silencing; D006801:Humans; D007124:Immunoenzyme Techniques; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics; D012097:Repressor Proteins / Q000235:genetics* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D002583:Uterine Cervical Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D012333:RNA, Messenger; D012097:Repressor Proteins; C485569:WIF1 protein, human; D000077209:Decitabine; D001374:Azacitidine