RBBP4 downregulation increases the sensitivity of A549 and HeLa cells to cisplatin by inhibiting cyclin D1 expression

Abstract

Introduction: Retinoblastoma-Binding Protein 4 (RBBP4), belonging to the WD-40 family, is an important member of the Polycomb Repressor Complex 2 (PRC2), the Nucleosome Remodeling and Deacetylation complex (NuRD), and is involved in chromatin remodeling, histone deacetylation, and H3K27 methylation. Methods: The effects of cisplatin treatment on cell viability were evaluated using the MTT assay. Western blotting was employed to analyze protein expression, and RNA interference-mediated knockdown of RBBP4 and cyclin D1 was conducted using Lipofectamine 2000. The formation of colonies was evaluated following a 14-day cisplatin treatment period. Cisplatin uptake was quantified by atomic absorption spectrophotometry. RNA sequencing was conducted on total RNA extracted from cells, and lentiviral vectors were employed for gene overexpression, followed by puromycin selection. Immunohistochemistry was performed on tissue microarrays of lung and cervical adenocarcinoma in order to evaluate RBBP4 expression. Results: In this study, cisplatin was found to induce RBBP4 expression in human lung cancer A549 cells, and RBBP4 expression in cisplatin-resistant A549/DDP cells was significantly higher than in A549 cells. Down regulating RBBP4 expression by small interfering RNA significantly increased the sensitivity of A549 and A549/DDP cells to cisplatin. Conversely, lentiviral-mediated RBBP4 overexpression reduced sensitivity to cisplatin. Mechanistic studies showed that downregulated RBBP4 increased cell sensitivity to cisplatin mainly by inhibiting cyclinD1 expression, and lentiviral-mediated cyclinD1 caused the opposite effects. These same results were verified in human HeLa cells and in cisplatin-resistant HeLa/DDP cells. Discussion: This study showed that RBBP4 regulates the sensitivity of tumor cells to cisplatin and is a potential target for reversing cisplatin resistance in tumor cells.