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Expression of the RET proto-oncogene is regulated by TFAP2C in breast cancer independent of the estrogen receptor.

ABSTRACT: BACKGROUND:The RET proto-oncogene is expressed as part of the estrogen receptor (ER) cluster in breast cancer. We sought to determine if TFAP2C regulates Ret expression directly or indirectly through ER. METHODS:Chromatin immunoprecipitation sequencing (ChIP-Seq) and gel-shift assay were used to identify TFAP2C binding sites in the RET promoter in four breast cancer cell lines. Ret mRNA and protein levels were evaluated in ER-positive and ER-negative breast cancer cell lines after knockdown of TFAP2C. Luciferase expression assay was performed to assess expression from two of the identified sites. RESULTS:ChIP-Seq identified five main binding peaks for TFAP2C in the RET promoter at -101.5 kb, -50.7 kb, -32.5 kb, +5.0 kb, and +33.6 from the RET transcriptional start site. Binding at three of the AP-2 sites was conserved across all four cell lines, whereas the RET -101.5 and RET +33.6 sites were each found to be unbound by TFAP2C in one cell line. A TFAP2C consensus element was confirmed for all five sites. Knockdown of TFAP2C by siRNA in ER-positive MCF-7 cells resulted in significant down regulation of Ret mRNA compared to nontargeting (NT) siRNA (0.09 vs. 1.0, P < 0.001). Knockdown of TFAP2C in ER-negative MDA-MB-453 cells also led to a significant reduction in Ret mRNA compared to NT siRNA (0.16 vs. 1.0, P < 0.001). In MCF-7 cells, knockdown of TFAP2C abrogated Ret protein expression (0.02 vs. 1.0, P < 0.001) before reduction in ER. CONCLUSIONS:TFAP2C regulates expression of the RET proto-oncogene through five AP-2 regulatory sites in the RET promoter. Regulation of Ret by TFAP2C occurs independently of ER expression in breast carcinoma.

SUBMITTER: Spanheimer PM

PROVIDER: S-EPMC3697477 | biostudies-literature | 2013 Jul

REPOSITORIES: biostudies-literature

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Expression of the RET proto-oncogene is regulated by TFAP2C in breast cancer independent of the estrogen receptor.

Spanheimer Philip M PM Woodfield George W GW Cyr Anthony R AR Kulak Mikhail V MV White-Baer Lola S LS Bair Thomas B TB Weigel Ronald J RJ

Annals of surgical oncology 20120810 7

<h4>Background</h4>The RET proto-oncogene is expressed as part of the estrogen receptor (ER) cluster in breast cancer. We sought to determine if TFAP2C regulates Ret expression directly or indirectly through ER.<h4>Methods</h4>Chromatin immunoprecipitation sequencing (ChIP-Seq) and gel-shift assay were used to identify TFAP2C binding sites in the RET promoter in four breast cancer cell lines. Ret mRNA and protein levels were evaluated in ER-positive and ER-negative breast cancer cell lines after ...[more]

PMID: 22878616

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Project description:BackgroundKIT is a proto-oncogene involved in diverse neoplastic processes. Aberrant kinase activity of the KIT receptor has been targeted by tyrosine kinase inhibitor (TKI) therapy in different neoplasias. In all the earlier studies, KIT expression was reported to be absent in meningiomas. However, we observed KIT mRNA expression in some meningioma cases. This prompted us to undertake its detailed analyses in meningioma tissues resected during 2008-2009.MethodsTumor tissues and matched peripheral blood samples collected from meningioma patients were used for detailed molecular analyses. KIT expression was ascertained immunohistochemically and validated by immunoblotting. KIT and KITLG transcript levels were discerned by reverse transcription quantitative real-time PCR (RT-qPCR). Similarly, KIT amplification and allele loss were assessed by quantitative real-time (qPCR) and validated by fluorescence in situ hybridization (FISH) on the neoplastic tissues. Possible alterations of the gene at the nucleotide level were analyzed by sequencing.ResultsContrary to earlier reports, KIT expression, was detected immunohistochemically in 20.6% meningioma cases (n = 34). Receptor (KIT) and ligand (KITLG) transcripts monitored by RT-qPCR were found to co-express (p = 0.048) in most of the KIT immunopositive tumors. 1/7 KIT positive meningiomas showed allele loss corroborated by reduced FISH signal in the corresponding neoplastic tissue. Sequence analysis of KIT showed M541L substitution in exon 10, in one of the immunopositive cases. However, its biological consequence remains to be uncovered.ConclusionsThis study clearly demonstrates KIT over-expression in the human meningiomas. The data suggest that up-regulated KIT transcription (p < 0.001), instead of gene amplification (p > 0.05), is a likely mechanism responsible for altered KIT expression. Thus, KIT is a potential candidate for detailed investigation in the context of meningioma pathogenesis.

Dataset Information

Expression of the RET proto-oncogene is regulated by TFAP2C in breast cancer independent of the estrogen receptor.

Publications

Expression of the RET proto-oncogene is regulated by TFAP2C in breast cancer independent of the estrogen receptor.

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