The role of 3-phosphoinositide-dependent protein kinase 1 in activating AGC kinases defined in embryonic stem cells.
BACKGROUND
Protein kinase B (PKB), and the p70 and p90 ribosomal S6 kinases (p70 S6 kinase and p90 Rsk, respectively), are activated by phosphorylation of two residues, one in the 'T-loop' of the kinase domain and, the other, in the hydrophobic motif carboxy terminal to the kinase domain. The 3-phosphoinositide-dependent protein kinase 1 (PDK1) activates many AGC kinases in vitro by phosphorylating the T-loop residue, but whether PDK1 also phosphorylates the hydrophobic motif and whether all other AGC kinases are substrates for PDK1 is unknown.


RESULTS
Mouse embryonic stem (ES) cells in which both copies of the PDK1 gene were disrupted were viable. In PDK1(-/-) ES cells, PKB, p70 S6 kinase and p90 Rsk were not activated by stimuli that induced strong activation in PDK1(+/+) cells. Other AGC kinases - namely, protein kinase A (PKA), the mitogen- and stress-activated protein kinase 1 (MSK1) and the AMP-activated protein kinase (AMPK) - had normal activity or were activated normally in PDK1(-/-) cells. The insulin-like growth factor 1 (IGF1) induced PKB phosphorylation at its hydrophobic motif, but not at its T-loop residue, in PDK1(-/-) cells. IGF1 did not induce phosphorylation of p70 S6 kinase at its hydrophobic motif in PDK1(-/-) cells.


CONCLUSIONS
PDK1 mediates activation of PKB, p70 S6 kinase and p90 Rsk in vivo, but is not rate-limiting for activation of PKA, MSK1 and AMPK. Another kinase phosphorylates PKB at its hydrophobic motif in PDK1(-/-) cells. PDK1 phosphorylates the hydrophobic motif of p70 S6 kinase either directly or by activation of another kinase.
Williams
M R
MR
MRC Protein Phosphorylation Unit, MSI/WTB Complex, University of Dundee, Dundee, DD1 5EH, Scotland. m.r.williams@dundee.ac.uk
Arthur
J S
JS
Balendran
A
A
van der Kaay
J
J
Poli
V
V
Cohen
P
P
Alessi
D R
DR
D064413:3-Phosphoinositide-Dependent Protein Kinases; D055372:AMP-Activated Protein Kinases; D000818:Animals; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D002478:Cells, Cultured; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects / Q000528:radiation effects; D017353:Gene Deletion; D038362:Glycogen Synthase Kinase 3; D015151:Immunoblotting; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology; D051379:Mice; D009097:Multienzyme Complexes / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology / Q000502:physiology*; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D019893:Ribosomal Protein S6 Kinases / Q000378:metabolism; D038744:Ribosomal Protein S6 Kinases, 90-kDa; D013234:Stem Cells / Q000201:enzymology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D009097:Multienzyme Complexes; D011518:Proto-Oncogene Proteins; D007334:Insulin-Like Growth Factor I; D064413:3-Phosphoinositide-Dependent Protein Kinases; C578564:Pdpk1 protein, mouse; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D019893:Ribosomal Protein S6 Kinases; D038744:Ribosomal Protein S6 Kinases, 90-kDa; C116919:mitogen and stress-activated protein kinase 1; D017868:Cyclic AMP-Dependent Protein Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D038362:Glycogen Synthase Kinase 3; D055372:AMP-Activated Protein Kinases; D013755:Tetradecanoylphorbol Acetate