Construction of a recombinant lentivirus-mediated shRNA expression vector targeting the human PSMD10 gene and validation of RNAi efficiency in RPMI-8226 multiple myeloma cells

摘要

Multiple myeloma (MM) is one of the most common malignant blood cancers. Previous studies have reported that proteasome 26S subunit non-ATPase 10 (PSMD10) is an oncoprotein with complex roles in hepatocellular carcinoma and other malignant tumors. Notably, research on the relationship between PSMD10 and tumorigenesis of MM has rarely been reported. The present study was designed to explore the possibility of PSMD10 as a therapeutic target in the treatment of MM, and the use of RNA interference (RNAi) to determine the function PSMD10. A recombinant lentivirus-mediated short hairpin RNA (shRNA) targeting human PSMD10 mRNA was constructed and used to decrease endogenous PSMD10 expression in the MM RPMI-8226 cell line in vitro. Expression of the PSMD10-targeting shRNA in RPMI-8226 cells transduced with the recombinant vector could be tracked by observing the expression of green fluorescent protein after infection. A transient transgenic RPMI-8226 cell line was generated by transducing cells with the packaged viral particles. Western blot analysis indicated that the protein levels of PSMD10 in the PSMD10-shRNA MM cells were significantly lower than those in the cells transduced with the negative control shRNA. Notably, RT-qPCR analysis did not reveal a marked change in the PSMD10 mRNA level; thus, the knockdown effect of the PSMD10-shRNA may occur during translation. Furthermore, apoptosis of MM cells was increased by silencing PSMD10 expression. Overall, the results demonstrated that the lentivirus-mediated shRNA vector-based RNAi expression system is an efficient method to silence PSMD10 gene expression in the MM RPMI-8226 cell line. It may provide a basis to study the role of PSMD10 in tumor cells, and may be a reliable gene therapy strategy in the clinic.