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. 2015 Sep 28;35(23):3049–3061. doi: 10.1038/onc.2015.361

Figure 3.

Figure 3

Akt and SGK3 cooperatively regulates colon cancer cell proliferation downstream of INPP4B. (a) WiDr and HCT116 cells stably transduced with the control shRNA (shControl) or INPP4B shRNA (shINPP4B1) were transduced with the vector alone or myr-Akt cDNA. Forty-eight hours later, whole-cell lysates were subjected to western blotting analysis of phosphorylated Akt (pSer473-Akt and pThr308-Akt), Akt and GAPDH (as a loading control). Data are representative of three individual experiments. (b) WiDr and HCT116 cells stably transduced with the shControl or shINPP4B1 were transduced with the vector alone or myr-Akt cDNA. Forty-eight hours later, cells were subjected to bromodeoxyuridine (BrdU) incorporation assays. Data are represented as mean±s.e.m. of three individual experiments. Student's t-test. (c) Whole-cell lysates from WiDr and HCT116 cells stably transduced with shControl or two individual INPP4B shRNAs (shINPP4B1 and shINPP4B2) were subjected to western blotting analysis of phosphorylated SGK3 (pThr320-SGK3), SGK3, phosphorylated SGK1 (pSer422-SGK1), SGK1 and GAPDH (as a loading control). Data are representative of three individual experiments. (d) WiDr and HCT116 cells stably transduced shINPP4B1 were transduced with the vector alone, myr-SGK3 cDNA, myr-Akt cDNA or myr-SGK3 cDNA plus myr-Akt cDNA. Forty-eight hours later, whole-cell lysates were subjected to western blotting analysis of phosphorylated SGK3 (pThr320-SGK3), SGK3, phosphorylated Akt (pSer473-Akt and pThr308-Akt), Akt and GAPDH (as a loading control). Data are representative of three individual experiments. (e) WiDr cells stably transduced with shINPP4B1 were transduced with the vector alone, myr-SGK3 cDNA, myr-Akt cDNA or myr-SGK3 cDNA plus myr-Akt cDNA. Forty-eight hours later, cells were subjected to BrdU incorporation assays. Data are represented as mean±s.e.m. of three individual experiments. *P<0.05, Student's t-test.