Effect of overexpression and nuclear translocation of constitutively active PKB-α on cellular survival and proliferation in HepG2 cells

✍ Scribed by Dhananjay Gupta; Noor A. Syed; William J. Roesler; Ramji L. Khandelwal

• Publisher: John Wiley and Sons
• Year: 2004
• Tongue: English
• Weight: 375 KB
• Volume: 93
• Category: Article
• ISSN: 0730-2312
• DOI: 10.1002/jcb.20200

✦ Synopsis

Abstract

Protein kinase B (Akt/PKB) is a key component in the PI 3‐kinase mediated cell survival pathway and has oncogenic transformation potential. Although the over‐expression of PKB‐α can prevent cell death following growth factor withdrawal, the long‐term effects of stable over‐expression of PKB‐α on cell survival in the absence of growth factors remain to be resolved. In the present study, we generated HepG2 cells with stable expression of active PKB‐α and compared its characteristics with HepG2 cells. Basal as well as insulin‐stimulated levels of Ser^473^ and Thr^308^ phosphorylation in PKB‐α transfected HepG2 cells were much higher than HepG2 cells. Constitutive expression of active PKB‐α enabled HepG2 cells to survive up to 96 h without serum in growth media while HepG2 cells fail to survive after 48 h of serum withdrawal. A strong positive correlation (R^2^ = 0.71) between cell proliferation and phosphorylated form of PKB‐α at Thr^308^ was observed along with higher levels of phosphorylated 3′‐phosphoinositide‐dependent kinase‐1 (PDK‐1). HepG2 cells with constitutive expression of active PKB‐α also showed higher levels of phosphorylated p65 subunit of nuclear factor‐κB (NFκB) in comparison with HepG2 cells. Predominant nuclear localization of phosphorylated PKB‐α was observed in stably transfected HepG2 cells. These results indicate that constitutive expression of active PKB‐α renders HepG2 cells independent of serum based growth factors for survival and proliferation. © 2004 Wiley‐Liss, Inc.