Mitogenic activation, phosphorylation, and nuclear translocation of protein kinase B beta

Protein kinase B (PKB) is a member of the second messenger-dependent family of serine/threonine kinases that has been implicated in signaling pathways downstream of growth factor receptor tyrosine kinases and phosphatidylinositol 3-kinase. Here we report the characterization of the human beta-isoform of PKB (PKB beta). PKB beta is ubiquitously expressed in a number of human tissues, with mRNA and protein levels elevated in heart, liver, skeletal muscle, and kidney. After transfection into HEK-293 or COS-1 cells, PKB beta is activated 2- to 12-fold by mitogens and survival factors. Activation was due to phosphorylation on Thr-309 and Ser-474, which correspond to Thr-308 and Ser-473 implicated in the regulation of PKB alpha, Both phosphorylation and activation were prevented by the phosphatidylinositol 3-kinase inhibitor wortmannin, Moreover, membrane-targeted PKB beta was constitutively activated when overexpressed in HEK-293 cells, Although the specific activity of PKB beta was lower than that of PKB alpha toward Crosstide as a substrate (23 nmol/min/mg compared with 170 nmol/ min/mg for PKB alpha), both enzymes showed similar substrate specificities, Using confocal microscopy, we show that activation of PKB beta results in its nuclear translocation within 20 to 30 min after stimulation, These observations provide evidence that PKB beta undergoes nuclear translocation upon mitogenic activation and support a role for PKB in signaling from receptor tyrosine kinases to the nucleus through phosphatidylinositol 3-kinase.