RESUMO
In this study, we investigated the regulation of FOXM1 expression by estrogen receptor alpha (ERalpha) and its role in hormonal therapy and endocrine resistance. FOXM1 protein and mRNA expression was regulated by ER-ligands, including estrogen, tamoxifen (OHT) and fulvestrant (ICI182780; ICI) in breast carcinoma cell lines. Depletion of ERalpha by RNA interference (RNAi) in MCF-7 cells downregulated FOXM1 expression. Reporter gene assays showed that ERalpha activates FOXM1 transcription through an estrogen-response element (ERE) located within the proximal promoter region. The direct binding of ERalpha to the FOXM1 promoter was confirmed in vitro by mobility shift and DNA pull-down assays and in vivo by chromatin immunoprecipitation (ChIP) analysis. Our data also revealed that upon OHT treatment ERalpha recruits histone deacetylases to the ERE site of the FOXM1 promoter, which is associated with a decrease in histone acetylation and transcription activity. Importantly, silencing of FOXM1 by RNAi abolished estrogen-induced MCF-7 cell proliferation and overcame acquired tamoxifen resistance. Conversely, ectopic expression of FOXM1 abrogated the cell cycle arrest mediated by the anti-estrogen OHT. OHT repressed FOXM1 expression in endocrine sensitive but not resistant breast carcinoma cell lines. Furthermore, qRT-PCR analysis of breast cancer patient samples revealed that there was a strong and significant positive correlation between ERalpha and FOXM1 mRNA expression. Collectively, these results show FOXM1 to be a key mediator of the mitogenic functions of ERalpha and estrogen in breast cancer cells, and also suggest that the deregulation of FOXM1 may contribute to anti-estrogen insensitivity.
Assuntos
Neoplasias da Mama/genética , Antagonistas de Estrogênios/farmacologia , Receptor alfa de Estrogênio/fisiologia , Estrogênios/farmacologia , Fatores de Transcrição Forkhead/genética , Regulação Neoplásica da Expressão Gênica/fisiologia , Western Blotting , Neoplasias da Mama/tratamento farmacológico , Ciclo Celular , Linhagem Celular Tumoral , Proliferação de Células , Imunoprecipitação da Cromatina , Resistencia a Medicamentos Antineoplásicos , Ensaio de Desvio de Mobilidade Eletroforética , Estradiol/análogos & derivados , Estradiol/farmacologia , Feminino , Proteína Forkhead Box M1 , Fatores de Transcrição Forkhead/metabolismo , Fulvestranto , Humanos , Mutagênese Sítio-Dirigida , Regiões Promotoras Genéticas/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Elementos de Resposta , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células Tumorais CultivadasRESUMO
The B-MYB proto-oncogene is a transcription factor belonging to the MYB family that is frequently overexpressed or amplified in different types of human malignancies. While it is suspected that B-MYB plays a role in human cancer, there is still no direct evidence of its causative role. Looking for mutations of the B-MYB gene in human cell lines and primary cancer samples, we frequently isolated two nonsynonymous B-MYB polymorphic variants (rs2070235 and rs11556379). Compared to the wild-type protein, the B-MYB isoforms display altered conformation, impaired regulation of target genes and decreased antiapoptotic activity, suggesting that they are hypomorphic variants of the major allele. Importantly, the B-MYB polymorphisms are common; rs2070235 and rs11556379 are found, depending on the ethnic background, in 10-50% of human subjects. We postulated that, if B-MYB activity is important for transformation, the presence of common, hypomorphic variants might modify cancer risk. Indeed, the B-MYB polymorphisms are underrepresented in 419 cancer patients compared to 230 controls (odds ratio 0.53; (95%) confidence interval 0.385-0.755; P=0.001). This data imply that a large fraction of the human population is carrier of B-MYB alleles that might be associated with a reduced risk of developing neoplastic disease.
