YAP promotes the proliferation of neuroblastoma cells through decreasing the nuclear location of p27 ^Kip1 mediated by Akt

Abstract

Objective

We aimed to investigate the roles and underlying mechanisms of YAP in the proliferation of neuroblastoma cells.

Methods

The expression level of YAP was evaluated by Western blotting and immunocytochemistry. Cell viability, cell proliferation and growth were detected by CCK‐8, PH3 and Ki67 immunostaining, and the real‐time cell analyser system. The nuclear and cytoplasmic proteins of p27 ^Kip1 were dissociated by the nuclear‐cytosol extraction kit and were detected by Western blotting and immunocytochemistry. mRNA levels of Akt, CDK5 and CRM1 were determined by qRT‐PCR.

Results

YAP was enriched in SH‐SY5Y cells (a human neuroblastoma cell line). Knock‐down of YAP in SH‐SY5Y cells or SK‐N‐SH cell line (another human neuroblastoma cell line) significantly decreased cell viability, inhibited cell proliferation and growth. Mechanistically, knock‐down of YAP increased the nuclear location of p27 ^Kip1, whereas serum‐induced YAP activation decreased the nuclear location of p27 ^Kip1 and was required for cell proliferation. Meanwhile, overexpression of YAP in these serum‐starved SH‐SY5Y cells decreased the nuclear location of p27 ^Kip1, promoted cell proliferation and overexpression of p27 ^Kip1 in YAP‐activated cells inhibited cell proliferation. Furthermore, knock‐down of YAP reduced Akt mRNA and protein levels. Overexpression of Akt in YAP‐downregulated cells decreased the nuclear location of p27 ^Kip1 and accelerated the proliferation of SH‐SY5Y cells.

Conclusions

Our studies suggest that YAP promotes the proliferation of neuroblastoma cells through negatively controlling the nuclear location of p27 ^Kip1 mediated by Akt.