RESUMO
BACKGROUND: MiR-27a is significantly overexpressed in triple-negative breast cancer (TNBC). However, the exact biological function of MiR-27a in TNBC is not fully understood. In this study, we verified miR-27a expression in TNBC cells and explored how its overexpression modulates radiosensitivity of the cells. MATERIAL/METHODS: qRT-PCR analysis was performed to study miR-27a expression in TNBC lines MDA-MB-435 and MDA-MB-231 and in normal human breast epithelial cell line MCF10A. Dual luciferase assay was performed to verify a putative downstream target of miR-27a, CDC27. CCK-8 assay was used to assess the influence of miR-27a-CDC27 axis on cell proliferation under irradiation (IR) treatment. RESULTS: We confirmed significantly higher miR-27a expression in 2 TNBC cell lines--MDA-MB-435 and MDA-MB-231--than in human breast epithelial cell line MCF10A. miR-27a could modulate proliferation and radiosensitivity of TNBC cells. CDC-27 is a direct target of miR-27a and its downregulation conferred increased radioresistance of the cells. CONCLUSIONS: The miR-27a-CDC27 axis might play an important role in modulating response to radiotherapy in TNBC cells. Testing miR-27a expression might be a useful way to identify a subgroup of patients who will benefit from an IR-based therapeutic approach.
Assuntos
Subunidade Apc3 do Ciclossomo-Complexo Promotor de Anáfase/antagonistas & inibidores , MicroRNAs/fisiologia , Proteínas de Neoplasias/antagonistas & inibidores , Neoplasias de Mama Triplo Negativas/radioterapia , Subunidade Apc3 do Ciclossomo-Complexo Promotor de Anáfase/biossíntese , Subunidade Apc3 do Ciclossomo-Complexo Promotor de Anáfase/genética , Subunidade Apc3 do Ciclossomo-Complexo Promotor de Anáfase/fisiologia , Sítios de Ligação , Mama/citologia , Linhagem Celular Tumoral/efeitos da radiação , Células Cultivadas , Sequência Conservada , Regulação para Baixo , Células Epiteliais/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Células HEK293 , Humanos , Terapia de Alvo Molecular , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/fisiologia , Oligonucleotídeos/farmacologia , Interferência de RNA , RNA Interferente Pequeno/farmacologia , Tolerância a Radiação/genética , Proteínas Recombinantes de Fusão/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/patologia , Ensaio Tumoral de Célula-TroncoRESUMO
UNLABELLED: CDC27 is a core component of the anaphase-promoting complex/cyclosome (APC/C), a multisubunit E3 ubiquitin ligase, whose oscillatory activity is responsible for the metaphase-to-anaphase transition and mitotic exit. Here, in normal murine mammary gland epithelial cells (NMuMG), CDC27 expression is controlled posttranscriptionally through the RNA binding protein poly(rC) binding protein 1 (PCBP1)/heterogeneous nuclear ribonucleoprotein E1 (HNRNP E1). shRNA-mediated knockdown of HNRNP E1 abrogates translational silencing of the Cdc27 transcript, resulting in constitutive expression of CDC27. Dysregulated expression of CDC27 leads to premature activation of the G2-M-APC/C-CDC20 complex, resulting in the aberrant degradation of FZR1/CDH1, a cofactor of the G1 and late G2-M-APC/C and a substrate normally reserved for the SCF-ßTRCP ligase. Loss of CDH1 expression and of APC/C-CDH1 activity, upon constitutive expression of CDC27, results in mitotic aberrations and aneuploidy in NMuMG cells. Furthermore, tissue microarray of breast cancer patient tumor samples reveals high CDC27 levels compared with nonneoplastic breast tissue and a significant correlation between disease recurrence and CDC27 expression. These results suggest that dysregulation of HNRNP E1-mediated translational regulation of Cdc27 leads to chromosomal instability and aneuploidy and that CDC27 expression represents a significant predictor of breast cancer recurrence. IMPLICATIONS: The RNA-binding protein HNRNP E1 mediates translational regulation of the cell-cycle regulator CDC27 and that dysregulation of CDC27 leads to aneuploidy. In addition, high CDC27 expression in breast cancer patient tumor specimens significantly predicts disease recurrence, suggesting a novel role for CDC27 as a predictor of relapse. Mol Cancer Res; 14(7); 634-46. ©2016 AACR.