RESUMEN
BACKGROUND: Thyroid carcinoma (THCA) is a common malignancy of the endocrine system. Further understanding of the molecular mechanism underlying THCA is crucial to develop effective diagnostic therapy and improve its treatments. In this study, we intended to provide novel direction for THCA targeted therapy from the aspect of lncRNA-miRNA-mRNA interaction. We aimed to investigate the function and molecular mechanism of lncRNA ATP1A1-AS1 in THCA. METHODS: Gene expression was assessed by reverse transcription quantitative polymerase chain reaction (RT-qPCR). Cell growth was detected by CCK-8 and EdU assays. Flow cytometry was applied for analyzing cell apoptosis. The binding of ATP1A1-AS1 or IRF2BP2 to miR-620 was validated by RNA pulldown and luciferase reporter assays. The protein levels were examined by western blotting. RESULTS: ATP1A1-AS1 was decreased in THCA cells and tissues. ATP1A1-AS1 overexpression attenuated cell growth and promoted apoptosis. MiR-620, which was upregulated in THCA, was identified as a direct target of ATP1A1-AS1. Furthermore, IRF2BP2 was discovered to be a target of miR-620, which displayed low expression in THCA cells and tissues. Importantly, IRF2BP2 knockdown reversed the influence of ATP1A1-AS1 overexpression on THCA cell proliferation and apoptosis. CONCLUSIONS: LncRNA ATP1A1-AS1 inhibited cell growth and promotes apoptosis in THCA via the miR-620/IRF2BP2 axis.
Asunto(s)
MicroARNs , ARN Largo no Codificante , Neoplasias de la Tiroides , Humanos , ARN Largo no Codificante/genética , ARN Largo no Codificante/metabolismo , Línea Celular Tumoral , MicroARNs/genética , MicroARNs/metabolismo , Neoplasias de la Tiroides/genética , Proliferación Celular/genética , Apoptosis/genética , Regulación Neoplásica de la Expresión Génica , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , ATPasa Intercambiadora de Sodio-Potasio/genética , ATPasa Intercambiadora de Sodio-Potasio/metabolismoRESUMEN
Breast cancer is the leading cause of death in female cancers, and what's worse, tamoxifen resistance occurs in almost 30% breast cancer patients and has seriously attenuated the therapeutic effect. It is widely studied that epigenetic regulation has played important role in the development of tamoxifen resistance. FOXA1 is a pioneer transcription factor that can translate epigenetic signature into transcription regulation and also drive genome-wide enhancer reprogramming in breast cancer. However, the chromatin super enhancer landscape orchestrated by FOXA1 and the key downstream targets of the FOXA1 oncogenic network in tamoxifen resistance remain elusive. Through analyzing the FOXA1 ChIP-seq data in tamoxifen sensitive MCF7 and tamoxifen resistant MCF7/TamR cells, we show that the FOXA1 chromatin occupancy is enhanced in both the promoter and enhancer regions, and the recruitment events may be E2 dependent in both MCF7 and MCF7/TamR cells. By integratively analyzing the FOXA1 ChIP-seq data and RNA-seq data of MCF7 and MCF7/TamR cells, we find that the enhanced or reduced FOXA1 chromatin binding densities may synchronize the transcriptional activity in tamoxifen resistance. Besides, we identify 1003 super enhancer associated protein coding genes and five super enhancer associated lncRNAs (ATP1A1-AS1, CASC11, CASC15, KCTD21-AS1, LINC00885) in tamoxifen resistance. By KM survival analysis, we find that high expression level of ATP1A1-AS1 and its sense transcript ATP1A1 indicates favorable clinical outcome among the luminal endocrine treated breast cancer patients. Further coexpression analysis indicates that ATP1A1-AS1 is significantly correlated with ATP1A1, and RT-qPCR results show that they both are downregulated in MCF7/TamR cells. Our study shows that the FOXA1 transcriptional regulatory network may promote the development of tamoxifen resistance, and identifies one super enhancer associated lncRNA ATP1A1-AS1 that may work as promising biomarker or drug target in tamoxifen resistance.
RESUMEN
Non-coding RNAs are important regulators of protein-coding genes. The current study characterized an antisense long non-coding RNA, ATP1A1-AS1, which is located on the opposite strand of the Na/K-ATPase α1 gene. Our results show that four splice variants are expressed in human adult kidney cells (HK2 cells) and embryonic kidney cells (HEK293 cells). These variants can be detected in both cytosol and nuclear fractions. We also found that the inhibition of DNA methylation has a differential effect on the expression of ATP1A1-AS1 and its sense gene. To investigate the physiological role of this antisense gene, we overexpressed the ATP1A1-AS1 transcripts, and examined their effect on Na/K-ATPase expression and related signaling function in human kidney cells. The results showed that overexpression of the ATP1A1-AS1-203 transcript in HK2 cells reduced the Na/K-ATPase α1 (ATP1A1) gene expression by approximately 20% (p < 0.05), while reducing the Na/K-ATPase α1 protein synthesis by approximately 22% (p < 0.05). Importantly, overexpression of the antisense RNA transcript attenuated ouabain-induced Src activation in HK2 cells. It also inhibited the cell proliferation and potentiated ouabain-induced cell death. These results demonstrate that the ATP1A1-AS1 gene is a moderate negative regulator of Na/K-ATPase α1, and can modulate Na/K-ATPase-related signaling pathways in human kidney cells.