RESUMEN

Lung cancer is one familiar cancer that threatens the lives of humans. circCTNNB1 has been disclosed to have regulatory functions in some diseases. However, the functions and related regulatory mechanisms of circCTNNB1 in lung cancer remain largely indistinct. The mRNA and protein expression levels were examined through real-time polymerase chain reaction (RT-qPCR) and western blot. The cell proliferation was tested through CCK-8 assay. The cell migration and invasion were confirmed through Transwell assays. The cell senescence was evaluated through SA-ß-gal assay. The binding ability between miR-186-5p and circCTNNB1 (or YY1) was verified through luciferase reporter and RIP assays. In this study, the higher expression of circCTNNB1 was discovered in lung cancer tissues and cell lines and resulted in poor prognosis. In addition, circCTNNB1 facilitated lung cancer cell proliferation, migration, invasion, and suppressed cell senescence. Knockdown of circCTNNB1 retarded the Wnt pathway. Mechanism-related experiments revealed that circCTNNB1 combined with miR-186-5p to target YY1. Through rescue assays, YY1 overexpression could rescue decreased cell proliferation, migration, invasion, increased cell senescence, and retarded Wnt pathway mediated by circCTNNB1 suppression. Furthermore, YY1 acts as a transcription factor that can transcriptionally activate circCTNNB1 to form YY1/circCTNNB1/miR-186-5p/YY1 positive loop. Through in vivo assays, circCTNNB1 accelerated tumour growth in vivo. All findings revealed that a positive loop YY1/circCTNNB1/miR-186-5p/YY1 aggravated lung cancer progression by modulating the Wnt pathway.

Asunto(s)

Proliferación Celular , Neoplasias Pulmonares , MicroARNs , ARN Circular , Vía de Señalización Wnt , Factor de Transcripción YY1 , Animales , Femenino , Humanos , Masculino , Ratones , Células A549 , Línea Celular Tumoral , Movimiento Celular , Progresión de la Enfermedad , Regulación Neoplásica de la Expresión Génica , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Neoplasias Pulmonares/metabolismo , Ratones Desnudos , MicroARNs/genética , MicroARNs/metabolismo , ARN Circular/genética , ARN Circular/metabolismo , Factor de Transcripción YY1/metabolismo , Factor de Transcripción YY1/genética

RESUMEN

BACKGROUND: Long non-coding RNA (lncRNA) HOTAIR has been reported to be associated with cisplatin (DDP) resistance in different human cancers including non-small cell lung cancer (NSCLC). However, the mechanism of HOTAIR in cisplatin resistance of NSCLC remains largely undefined. MATERIALS AND METHODS: Expression of HOTAIR, miR-149-5p and doublecortin-like kinase 1 (DCLK1) was detected using real-time quantitative PCR (RT-qPCR) and Western blotting. Cisplatin resistance was determined with cell counting kit (CCK)-8 assay and transwell assays in vitro, and xenograft tumor models in vivo. The target binding between miR-149-5p and either HOTAIR or DCLK1 was predicted on Diana Tools website, and confirmed by dual-luciferase reporter assay and RNA immunoprecipitation. RESULTS: Expression of HOTAIR was upregulated in DDP-resistant NSCLC tumor tissues and cell lines (A549/DDP and H1299/DDP). Knockdown of HOTAIR decreased the acquired cisplatin resistance of A549/DDP and H1299/DDP cells, as evidenced by attenuated 50% inhibitory concentration (IC50) of DDP, cell proliferation, migration and invasion in vitro, as well as tumor growth inhibition in vivo. Mechanically, HOTAIR negatively regulated miR-149-5p expression via targeting, and DCLK1 was a downstream target for miR-149-5p. DCLK1 was indirectly regulated by HOTAIR in DDP-resistant NSCLC cells as well. Functionally, miR-149-5p deletion could counteract the inhibitory effect of HOTAIR knockdown on cisplatin resistance; contrarily, restoring miR-149-5p exhibited the similar inhibition on cisplatin resistance in DDP-resistant cells in vitro, which was then abated by DCLK1 upregulation. CONCLUSION: Knockdown of HOTAIR enhances DDP-resistant NSCLC cells to overcome cisplatin resistance partially via regulating miR-149-5p/DCLK1 axis.