Silencing NUPR1 down-regulates autophagy and promotes migration and invasion in human multiple myeloma U266 cells

Abstract

Objective To explore the effect of silencing nuclear protein 1 (NUPR1) on autophagy, migration and invasion in human multiple myeloma (MM) U266 cells and its possible mechanism. Methods NUPR1-shRNA was transfected into MM U266 cells, and the transfection rate was detected by flow cytometry. Then the cells were divided into negative control cells (transfected with unrelated sequence RNA lentiviral vector) and NUPR1 knockdown cells (transfected with NUPR1-shRNA lentiviral vector). The interference effect of NUPR1 was detected by qRT-PCR, and the expression levels of autophagy related proteins (Beclin1, ATG5, LC3Ⅱ/LC3Ⅰ, P62), migration and invasion associated proteins (MMP9, CXCR4) and pathway proteins (p-AKT/T-AKT, p-mTOR/T-mTOR) were measured by Western blotting. Transmission electron microscopy (TEM) was used to observe autophagosomes. Transwell test was employed to detect cell migration and invasion. Results The expression of NUPR1 at mRNA and protein level was significantly decreased in NUPR1 knockdown cells (P < 0.05). The protein levels of autophagy related proteins, ATG5, Beclin1 and LC3Ⅱ/LC3Ⅰwere significantly lower, while those of f MMP9, CXCR4, P62, p-AKT/T-AKT and p-mTOR/T-mTOR were increased obviously in the NUPR1 knockdown cells than the negative control cells (P < 0.05). There were significantly reduced autophagosomes in NUPR1 knockdown cells. While, migratory and invasive abilities were enhanced in NUPR1 knock down group, and rapamycin reduced the promoting effects. Conclusion Silencing NUPR1 may promotes migration and invasion by down-regulating autophagy in U266 cells, which may be related to activation of the AKT/mTOR pathway.

Keywords

• nuclear protein 1
• multiple myeloma
• autophagy
• migration
• invasion