Fig. 6

PHB2 induces the ubiquitination degradation of SHIP2 by enhancing the interaction between NEDD4 and SHIP2. A Co-IP assays showing that NEDD4 was specifically co-precipitated with SHIP2 and PHB2 in HGC-27 cells. E3 ligases SIAH2, WWP2, MUL1 and Cul4A were used as negative controls. Data shown represent three independent experiments. B SiRNA knockdown of NEDD4 decreased the ubiquitination of SHIP2. MG132: 10 μM. C Deletion-mapping experiments showing that NEDD4 was precipitated with the 5-Ptase domain of SHIP2 but not other SHIP2 domains in HEK293T cells. D Overexpression of NEDD4 increased the ubiquitination full-length SHIP2 but not tGFP-SHIP2-del 5Ptase in HEK293T and HGC-27 cells. MG132:10 μM. E Silencing of PHB2 reduced the interaction between SHIP2 and NEDD4 in HEK293T cells. F Silencing of NEDD4 abolished the ubiquitination of SHIP2 caused by PHB2 overexpression. G, Overexpression of PHB2 mainly increased K48-linked polyubiquitination of SHIP2. MG132:10 μM. Data are representatives of three independent experiments