PDKl

Tyrosine phosphorylated IRS interacts with and activates PI 3-kinase [3]. Binding takes place via the SRC homology 2 (SH2) domain of the PI 3-kinase regulatory subunit. The resulting complex consisting of INSR, IRS, and PI 3-kinase facilitates interaction of the activated PI 3-kinase catalytic subunit with the phospholipid substrates in the plasma membrane. Generation of PI 3-phosphates in the plasma membrane reemits phospholipid dependent kinases (PDKl and PDK2) which subsequently phosphorylate and activate the serine/threonine kinase Akt (synonym protein... 

Kuhiak We found in mouse oocytes undergoing GVBD that Rsk is phosphorylated by two successive steps, one of which is dependent on Cdc2 and the following one on MAP kinase. There is also evidence of phosphorylation of Rsk by another kinase, which is Pdkl. 

Mailer For the last 15 years since Rsk was discovered no one could activate it in vitro from a recombinant source. This turns out to be because in addition to MAPK, Pdkl is required. There is no phosphorylation of Rsk by Cdc2. What we essentially made with our constitutively active construct is an enzyme solely dependent on Pdkl. 

Anderson, K.E., CoadweU, J., Stephens, L.R., and Hawkins, P.T., 1998, Translocation of PDKl to the plasma membrane is important in allowing PDK-1 to activate protein kinase B. Curr. Biol, 8 684-691. 

Keywords Hsp90 IGF-IR Inositol polyphosphate mTor PDKl PI3K Phosphatidylinositol Phospholipids PKB... 

The 3-phosphoinositide-dependent protein kinase-1 (PDKl) is a 556-amino acid enzyme composed of three well-differentiated motifs an N-terminal domain, a constitutively activated serine/threonine kinase domain, and a Pleck-strin homology (PH) domain at its C-terminus [87-91]. The attractiveness of PDKl as a potential anticancer target is hnked to its ability to control the activity of a diverse set of AGC kinase members, in particular the three PKB isoforms [92], Full activation of PKB requires phosphorylation at two sites. 

Contrary to the other major components of the PI3K/PKB survival pathway, the development of PDKl inhibitors would be, to a certain extent, simpler as only a single PDKl isoform exists in human cells. Moreover, the observation that PDKl hypomorphic mice expressing only approximately 10% of the normal level of PDKl display no obvious harmful phenotype—they are 40-50% smaller than control animals — [100], indicates that a PDKl inhibitor may provide effective anticancer therapy at an acceptable therapeutic index. 

The most potent PDKl kinase inhibitor reported to date is UCN-01 (compound 16 Fig. 4 IC50 = 6 to 33 nM) [101], a staurosporine analogue isolated from the culture broth of Streptomyces sp. Originally developed as an inhibitor of calcium-dependent PKC, UCN-01 has the capacity to inhibit a broad spectrum of kinases [102], including other members of the AGC subfamily of kinases (e.g., IC5o = 491nM for PKB) [103]. UCN-Ol-induced PDKl inhibition has also been observed in in vivo murine and human tumor xenografts [101]. 

In addition to pulmonary toxicity, nausea/vomiting, lactic acidosis and transaminitis, UCN-01 induced insulin resistance during Phase I clinical trials. As shown recently with rat adipose cells, this effect may be due to UCN-01 inhibition of PKB Thr-308 phosphorylation—no effect on Ser-473 was observed in this study—and subsequent blockade of GLUT4 translocation in response to insulin [104]. If this mode of action is confirmed in the ongoing clinical trials and contrary to what was observed in the PDKl hypomorphic mice (vide supra) [100], insulin resistance may represent an important hurdle in the development of PDKl inhibitors and, in general, of any agent that blocks the PI3K/PKB pathway in adipose and muscle cells. 

In addition to UCN-01 and other staurosporine or maleimide derivatives (e.g., compounds 17 and 18 Fig. 4) [102,103,105-107], PDKl kinase activity is inhibited by aminopyrimidines. A representative example of this compound class is BX-320 (compound 19, Fig. 4), a PDKl inhibitor (IC50 = 30 nM) that displays good selectivity over protein kinase A (PKA, 35-fold) [108]. BX-320 blocks the growth in soft agar of a wide range of tumor cell lines (IC50 = 0.093 to 1.32 xM), and shows efficacy in a metastasis mouse model (200 mg/kg bid). 

The membrane-associated Akt kinase is now a substrate for protein kinase PDKl that phosphorylates a specific Thr and Ser residue of Akt kinase. The double phosphorylation converts Akt kinase to the active form. It is assumed that the Akt kinase now dissociates from the membrane and phosphorylates cytosolic substrates such as glycogen synthase kinase, 6-phosphofructo-2-kinase and ribosomal protein S6 kinase, p70 . According to this mechanism, Akt kinase regulates central metabolic pathways of the cell. Furthermore, it has a promoting influence on cell division and an inhibitory influence on programmed cell death, apoptosis. A role in apoptosis is suggested by the observation that a component of the apoptotic program. Bad protein (see Chapter 15) has been identified as a substrate of Akt kinase. 

P. A. (1993). A comparison of the multiple alleles of xylS carried by TOL plasmids pWW53and pDKl and its implications for their evolutionary relationship. Journal of General Microbiology, 139. 557—68. 

Wu, S. and Kaufman, R.J., fran,y-autophosphorylation by the isolated kinase domain is not sufficient for dimerization of activation of the dsRNA-activated protein kinase PKR, Biochemistry 43, 11027-11034, 2004 Shi, G.W., Chen, J., Concepcion, F. et ah. Light causes phosphorylation of nonactivated visual pigments in intact mouse rod photoreceptor cells, J. Biol. Chem. 280, 41184-41191, 2005 Gao, X. and Harris, T.K., Steady-state kinetic mechanism of PDKl, J. Biol. Chem., 281, 21670-21681, 2006. 

PI3K-AKT Pathway Phospho-specific AKT antibody Antibody for Western blot detection of phosphorylated and activated AKT when phosphorylated by PDKl at T308... 

Gell survival AKT, PDKl, IKK, NFkB, Bad, Casp9, FKHRLl Cancer, Huntington s, Alzheimer s... 

Protein kinase B (PKB) becomes partially activated by binding to PI 3-phosphates. Its full activation requires phosphorylation by another kinase (PDKl), which also is recruited to the membrane by binding to PI 3-phosphates (see Figure 14-27). 

Mora, A., Komander, D., van Aalten, D. M., and Alessi, D. R. PDKl, the master regulator of AGC kinase signal transduction. Semin Cell Dev Biol 15 (2004) 161-170. 

PKB bound to PIP3 is phosphorylated by PDKl (not shown). Thus activated, PKB phosphorylates GSK3 on a Ser residue, inactivating it. 

Fig. 12. Some signaling targets and pathways affected by sphingolipid backbones that are metabolically interrelated. PKA, protein kinase A PDKl, 3-phosphoinositide-dependent kinase 1 SDKl, sphingosine-dependent kinase 1 PKC, protein kinase C PPl, protein phosphatase 1 PP2A, protein phosphatase 2A aSMase, acid sphingomyelinase PLA, phospholipase Aj SIP, sphingosine-l-phosphase MATIA, methionine adenosyl-transferase (liver specific) SFl, steroidogenic factor 1. The biophysical properties of ceramides and ceramide 1-phosphates may play important roles in membrane structure, including tendencies to form rafts, membrane curvature, and leakiness.

F. 11.14. The insulin receptor-protein kinase B signaling pathway. Abbreviations Ins, insulin IRS, insulin receptor substrate PH domains, pleckstrin homology domains PDKl, phosphoinositide-dependent protein kinase 1 PKB, protein kinase B. The final phosphorylation step that activates PKB is shown in blue. 

B and PDKl (phosphoinositide-dependent kinase-1) are recruited to the membrane by their PH domains, where PDKl phosphorylates and activates protein kinase B. Many other signal transducer proteins have PH domains and are docked at the membrane, where they can find and bind each other. Thus, the insulin signal diverges again and again. Insulin is covered in more detail in Chapters 26, 36 and 43. 

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