Mitogenic activation, phosphorylation, and nuclear translocation of protein kinase Bbeta.
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 (PKBbeta). PKBbeta 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, PKBbeta 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 PKBalpha. Both phosphorylation and activation were prevented by the phosphatidylinositol 3-kinase inhibitor wortmannin. Moreover, membrane-targeted PKBbeta was constitutively activated when overexpressed in HEK-293 cells. Although the specific activity of PKBbeta was lower than that of PKBalpha toward Crosstide as a substrate (23 nmol/min/mg compared with 178 nmol/min/mg for PKBalpha), both enzymes showed similar substrate specificities. Using confocal microscopy, we show that activation of PKBbeta results in its nuclear translocation within 20 to 30 min after stimulation. These observations provide evidence that PKBbeta 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.
Meier
R
R
Friedrich Miescher Institute, P. O. Box 2543, CH-4002 Basel, Switzerland.
Alessi
D R
DR
Cron
P
P
Andjelković
M
M
Hemmings
B A
BA
D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport; D019556:COS Cells; D002451:Cell Compartmentation; D002467:Cell Nucleus / Q000201:enzymology; D004789:Enzyme Activation; D006133:Growth Substances / Q000494:pharmacology; D006801:Humans; D007328:Insulin / Q000494:pharmacology; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology; D008934:Mitogens / Q000494:pharmacology; D008969:Molecular Sequence Data; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D010768:Phosphoserine / Q000378:metabolism; D010769:Phosphothreonine / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D051381:Rats; D013379:Substrate Specificity; D014018:Tissue Distribution
D006133:Growth Substances; D007328:Insulin; D007334:Insulin-Like Growth Factor I; D008934:Mitogens; D019869:Phosphatidylinositol 3-Kinases; D010768:Phosphoserine; D010769:Phosphothreonine; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt