Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: UNIPROT:P31749 (AKT)
22,954 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Protein kinase B and p70 S6 kinase are members of the cyclic AMP-dependent/cyclic GMP-dependent/protein kinase C subfamily of protein kinases and are activated by a phosphatidylinositol 3-kinase-dependent pathway when cells are stimulated with insulin or growth factors. Both of these kinases are activated in cells by phosphorylation of a conserved residue in the kinase domain (Thr-308 of protein kinase B (PKB) and Thr-252 of p70 S6 kinase) and another conserved residue located C-terminal to the kinase domain (Ser-473 of PKB and Thr-412 of p70 S6 kinase). Thr-308 of PKBalpha and Thr-252 of p70 S6 kinase are phosphorylated by 3-phosphoinositide-dependent protein kinase-1 (PDK1) in vitro. Recent work has shown that PDK1 interacts with a region of protein kinase C-related kinase-2, termed the PDK1 interacting fragment (PIF). Interaction with PIF converts PDK1 from a form that phosphorylates PKB at Thr-308 alone to a species capable of phosphorylating Ser-473 as well as Thr-308. This suggests that PDK1 may be the enzyme that phosphorylates both residues in vivo. Here we demonstrate that PDK1 is capable of phosphorylating p70 S6 kinase at Thr-412 in vitro. We study the effect of PIF on the ability of PDK1 to phosphorylate p70 S6 kinase. Surprisingly, we find that PDK1 bound to PIF is no longer able to interact with or phosphorylate p70 S6 kinase in vitro at either Thr-252 or Thr-412. The expression of PIF in cells prevents insulin-like growth factor 1 from inducing the activation of the p70 S6 kinase and its phosphorylation at Thr-412. Overexpression of PDK1 in cells induces the phosphorylation of p70 S6 kinase at Thr-412 in unstimulated cells, and a catalytically inactive mutant of PDK1 prevents the phosphorylation of p70 S6K at Thr-412 in insulin-like growth factor 1-stimulated cells. These observations indicate that PDK1 regulates the activation of p70 S6 kinase and provides evidence that PDK1 mediates the phosphorylation of p70 S6 kinase at Thr-412.
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PMID:Evidence that 3-phosphoinositide-dependent protein kinase-1 mediates phosphorylation of p70 S6 kinase in vivo at Thr-412 as well as Thr-252. 1060 11

Growth factors interact with their cell surface receptors and activate the enzyme PI 3-kinase (PI 3-K) resulting in the formation of 3-phosphorylated phosphatidylinositols, which in turn activate the serine/threonine kinase AKT/PKB. AKT functions, in part, to promote cell survival by phosphorylating the BCL-2 family member BAD and the cell death pathway enzyme, caspase-9. Although induction of apoptosis by ultraviolet (UV) irradiation is well documented, little is known about UV activation of cell survival pathways in human skin cells. We have investigated whether UV activates the PI 3-K/AKT pathway in human skin in vivo. UV irradiation (2MED from UVB source) stimulated PI 3-kinase activity within 15 min. PI 3-K activity was maximal (2.5-fold, n=6) 30 min post UV and remained elevated for 4 h. UV stimulated AKT activity within 30 min. Maximal activity (4-fold, n=11) was observed 1 h post UV. UV also stimulated phosphorylation of the downstream AKT effectors, S6 kinase and BAD. S6 kinase was maximally stimulated 4 h post UV (15-fold, n=6). Increased BAD phosphorylation was observed 1 h post UV and remained elevated for 4 h. Western blot analysis revealed that UV-induced phosphorylation of BAD at Ser112, a site known to be phosphorylated by AKT. Inhibitors of EGFR and PI 3-kinase blocked UV-induced phosphorylation of BAD, suggesting that EGFR mediates UV-activated cell survival pathway. Collectively, both positive and negative roles for UV activation of the PI 3-K/AKT pathway in human skin can be envisioned. The PI 3-K/AKT pathway likely plays a critical role in balancing UV-induced apoptotic signals, thereby preventing widespread skin cell death. Conversely UV activation of the PI 3-K/AKT pathway may enhance survival of mutated cells, thereby promoting skin cancer, as has been found in several other types of cancer.
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PMID:Ultraviolet irradiation activates PI 3-kinase/AKT survival pathway via EGF receptors in human skin in vivo. 1117 72

Phospholipid-dependent kinase 1 (PDK 1) is a 3'-phospholipid-responsive serine/threonine kinase that plays a critical role in cell survival by phosphorylating and activating the anti-apoptotic AKT/PKB kinase. While PDK 1 is clearly an important component of the cell survival machinery, the potential for phospholipid-independent activation of the AKT/PKB survival pathway has not been extensively examined at the molecular level. We have identified a second form of PDK 1 in the nematode Caenorhabditis elegans that we have termed PIAK (phospholipid-independent AKT/PKB kinase). PIAK is highly homologous to C. elegans and mammalian PDK 1 with the exception that the novel kinase lacks a phospholipid binding pleckstrin homology domain. The domain structure of PIAK suggests that it might be a phospholipid-independent kinase, and PIAK phosphorylates mammalian AKT/PKB at the activating Thr(308) residue in the presence of the phosphatidylinositol (PI) 3-kinase inhibitors as well as in the absence of growth factors. In addition, PIAK is capable of inducing the phospholipid-independent, AKT/PKB-induced phosphorylation of the AFX-type forkhead transcription factor, resulting in its cytoplasmic localization. Because the nuclear localization of this transcription factor induces an apoptotic state, this PIAK-mediated cytoplasmic sequestration allows for cell survival. Finally, PIAK activity appears to be induced by various inhibitors of cell cycle G(1) progression. These data suggest an alternate, phosphatidylinositol 3-kinase-independent mechanism for the activation of the AKT/PKB survival pathway that may be utilized during periods of cellular quiescence.
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PMID:Caenorhabditis elegans PIAK, a phospholipid-independent kinase that activates the AKT/PKB survival kinase. 1127 60

The phenotype of Bcr-Abl-transformed cells is characterized by a growth factor-independent survival and a reduced susceptibility to apoptosis. Furthermore, Bcr-Abl kinase alters adhesion features by phosphorylating cytoskeletal and/or signaling proteins important for integrin function. Integrin-mediated adhesion to extracellular matrix molecules is critical for the regulation of growth and apoptosis. However, effects of integrin signaling on regulation of apoptosis in cells expressing Bcr-Abl are largely unknown. The influence of adhesion on survival and apoptosis in Bcr-Abl+ and Bcr-Abl- BaF3 cells was investigated. p185bcr-abl-transfected BaF3 cells preadhered to immobilized fibronectin had a significant survival advantage and reduced susceptibility to apoptosis following gamma-irradiation when compared with the same cells grown on laminin, on polylysin, or in suspension. Both inhibition of Bcr-Abl kinase by STI571 and inhibition of specific adhesion reversed the fibronectin-mediated antiapoptotic effect in BaF3p185. The DNA damage response of Bcr-Abl- BaF3 cells was not affected by adhesion to fibronectin. In contrast to parental BaF3 cells, BaF3p185 adherent to fibronectin did not release cytochrome c to the cytosol following irradiation. The fibronectin-mediated antiapoptotic mechanism in Bcr-Abl-active cells was not mediated by overexpression of Bcl-XL or Bcl-2 but required an active phosphatidylinositol 3-kinase (PI-3K). Kinase-active Bcr-Abl in combination with fibronectin-induced integrin signaling led to a hyperphosphorylation of AKT. Thus, cooperative activation of PI-3K/AKT by Bcr-Abl and integrins causes synergistic protection of Bcr-Abl+ cells from DNA damage-induced apoptosis.
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PMID:Adhesion to fibronectin selectively protects Bcr-Abl+ cells from DNA damage-induced apoptosis. 1152 Aug 4

Phosphatidylinositol 3'-kinase (PI3K) and the serine/threonine kinase AKT have critical roles in phosphorylating and transactivating the p65 subunit of nuclear factor kappaB (NF-kappaB) in response to the pro-inflammatory cytokines interleukin-1 (IL-1) and tumor necrosis factor (TNF). Mouse embryo fibroblasts (MEFs) lacking either the alpha or beta subunit of IkappaB kinase (IKK) were deficient in NF-kappaB-dependent transcription following treatment with IL-1 or TNF. However, in contrast to IKKbeta-null MEFs, IKKalpha-null MEFs were not substantially defective in the cytokine-stimulated degradation of Ikappabetaalpha or in the nuclear translocation of NF-kappaB. The IKK complexes from IKKalpha- or IKKbeta-null MEFs were both deficient in PI3K-mediated phosphorylation of the transactivation domain of the p65 subunit of NF-kappaB in response to IL-1 and TNF, and constitutively activated forms of PI3K or AKT did not potentiate cytokine-stimulated activation of NF-kappaB in either IKKalpha- or IKKbeta-null MEFs. Collectively, these data indicate that, in contrast to IKKbeta, which is required for both NF-kappaB liberation and p65 phosphorylation, IKKalpha is required solely for the cytokine-induced phosphorylation and activation of the p65 subunit of NF-kappaB that are mediated by the PI3K/AKT pathway.
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PMID:Distinct roles of the Ikappa B kinase alpha and beta subunits in liberating nuclear factor kappa B (NF-kappa B) from Ikappa B and in phosphorylating the p65 subunit of NF-kappa B. 1173 37

Protein kinase B (PKB), also known as Akt, is a serine/threonine protein kinase controlled by insulin, various growth factors, and phosphatidylinositol 3-kinase. Full activation of the PKB enzyme requires phosphorylation of a threonine in the activation loop and a serine in the C-terminal tail. PDK1 has clearly been shown to phosphorylate the threonine, but the mechanism leading to phosphorylation of the serine, the PDK2 site, is unclear. A yeast two-hybrid screen using full-length human PKBgamma identified protein kinase C (PKC) zeta, an atypical PKC, as an interactor with PKBgamma, an association requiring the pleckstrin homology domain of PKBgamma. Endogenous PKBgamma was shown to associate with endogenous PKCzeta both in cos-1 cells and in 3T3-L1 adipocytes, demonstrating a physiological interaction. Immunoprecipitates of PKCzeta, whether endogenous PKCzeta from insulin-stimulated 3T3-L1 adipocytes or overexpressed PKCzeta from cos-1 cells, phosphorylated S472 (the C-terminal serine phosphorylation site) of PKBgamma, in vitro. In vivo, overexpression of PKCzeta stimulated the phosphorylation of approximately 50% of the PKBgamma molecules, suggesting a physiologically meaningful effect. However, pure PKCzeta protein was incapable of phosphorylating S472 of PKBgamma. Antisense knockout studies and use of a PDK1 inhibitor showed that neither PKB autophosphorylation nor phosphorylation by PDK1 accounted for the S472 phosphorylation in PKCzeta immunoprecipitates. Staurosporine inhibited the PKCzeta activity but not the PDK2 activity in PKCzeta immunoprecipitates. Together these results indicate that an independent PDK2 activity exists that physically associates with PKCzeta and that PKCzeta, by binding PKBgamma, functions to deliver the PDK2 to a required location. PKCzeta thus functions as an adaptor, associating with a staurosporine-insensitive PDK2 enzyme that catalyzes the phosphorylation of S472 of PKBgamma. Because both PKCzeta and PKB have been proposed to be required for mediating a number of crucial insulin responses, formation of an active signaling complex containing PKCzeta, PKB, and PDK2 is an attractive mechanism for ensuring that all the critical sites on targets such as glycogen synthase kinase-3 are phosphorylated.
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PMID:Characterization of PDK2 activity against protein kinase B gamma. 1216 51

Grb2-associated binder 1 (Gab1) is known to play an important role in hepatocyte growth factor (HGF) signaling, which rapidly becomes tyrosine-phosphorylated upon HGF stimulation. In this study, we found that the tyrosine phosphorylation of Gab1 in the cells derived from Src/Yes/Fyn null mouse embryos was approximately 40% lower than that in their wild type counterparts upon HGF stimulation. Increased expression of wild-type Src enhanced HGF-induced phosphorylation of Gab1, and, in contrast, expression of the Src kinase-deficient mutant or treatment of the specific Src inhibitor PP1 suppressed it. Expression of a constitutively active Src mutant (Y527F) or oncogenic v-Src led to a prominent increase in Gab1 phosphorylation independent of HGF stimulation. Moreover, Src interacted with Gab1 via both its Src homology 2 and 3 domains and was capable of phosphorylating purified Gab1 in vitro. Finally, the increased phosphorylation of Gab1 by Src selectively potentiated HGF-induced activation of ERK and AKT. Taken together, our results establish a new role for Src in HGF-induced Gab1 phosphorylation.
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PMID:Src phosphorylates Grb2-associated binder 1 upon hepatocyte growth factor stimulation. 1294 62

Protein kinase B (PKB)/Akt is known to promote cell migration, and this may contribute to the enhanced invasiveness of malignant cells. To elucidate potential mechanisms by which PKB/Akt promotes the migration phenotype, we have investigated its role in the endosomal transport and recycling of integrins. Whereas the internalization of alpha v beta 3 and alpha 5 beta 1 integrins and their transport to the recycling compartment were independent of PKB/Akt, the return of these integrins (but not internalized transferrin) to the plasma membrane was regulated by phosphatidylinositol 3-kinases and PKB/Akt. The blockade of integrin recycling and cell spreading on integrin ligands effected by inhibition of PKB/Akt was reversed by inhibition of glycogen synthase kinase 3 (GSK-3). Moreover, expression of nonphosphorylatable active GSK-3 beta mutant GSK-3 beta-A9 suppressed recycling of alpha 5 beta 1 and alpha v beta 3 and reduced cell spreading on ligands for these integrins, indicating that PKB/Akt promotes integrin recycling by phosphorylating and inactivating GSK-3. We propose that the ability of PKB/Akt to act via GSK-3 to promote the recycling of matrix receptors represents a key mechanism whereby integrin function and cell migration can be regulated by growth factors.
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PMID:Protein kinase B/Akt acts via glycogen synthase kinase 3 to regulate recycling of alpha v beta 3 and alpha 5 beta 1 integrins. 1474 68

We sought to elucidate the role of AKT in follicle-stimulating hormone (FSH)-mediated granulosa cell (GC) differentiation. Our results define a signaling pathway in GCs whereby the inactivating phosphorylation of tuberin downstream of phosphatidylinositol (PI) 3-kinase/AKT activity leads to Rheb (Ras homolog enriched in brain) and subsequent mTOR (mammalian target of rapamycin) activation. mTOR then stimulates translation by phosphorylating p70 S6 kinase and, consequently, the 40 S ribosomal protein S6. Activation of this pathway is required for FSH-mediated induction of several follicular differentiation markers, including luteinizing-hormone receptor (LHR), inhibin-alpha, microtubule-associated protein 2D, and the PKA type IIbeta regulatory subunit. FSH also promotes activation of the transcription factor hypoxia-inducible factor-1 (HIF-1). FSH-stimulated HIF-1 activity is inhibited by the PI 3-kinase inhibitor LY294002, the Rheb inhibitor FTI-277 (farnesyltransferase inhibitor-277), and the mTOR inhibitor rapamycin. Finally, we find that the FSH-mediated up-regulation of reporter activities for LHR, inhibin-alpha, and vascular endothelial growth factor is dependent upon HIF-1 activity, because a dominant negative form of HIF-1alpha interferes with the up-regulation of these genes. These results show that FSH enhances HIF-1 activity downstream of the PI 3-kinase/AKT/Rheb/mTOR pathway in GCs and that HIF-1 activity is necessary for FSH to induce multiple follicular differentiation markers.
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PMID:Follicle-stimulating hormone activation of hypoxia-inducible factor-1 by the phosphatidylinositol 3-kinase/AKT/Ras homolog enriched in brain (Rheb)/mammalian target of rapamycin (mTOR) pathway is necessary for induction of select protein markers of follicular differentiation. 1498 27

The hepatitis C virus NS5A protein plays a critical role in virus replication, conferring interferon resistance to the virus through perturbation of multiple intracellular signaling pathways. Since NS5A is a phosphoprotein, it is of considerable interest to understand the role of phosphorylation in NS5A function. In this report, we investigated the phosphorylation of NS5A by taking advantage of 119 glutathione S-transferase-tagged protein kinases purified from Saccharomyces cerevisiae to perform a global screening of yeast kinases capable of phosphorylating NS5A in vitro. A database BLAST search was subsequently performed by using the sequences of the yeast kinases that phosphorylated NS5A in order to identify human kinases with the highest sequence homologies. Subsequent in vitro kinase assays and phosphopeptide mapping studies confirmed that several of the homologous human protein kinases were capable of phosphorylating NS5A. In vivo phosphopeptide mapping revealed phosphopeptides common to those generated in vitro by AKT, p70S6K, MEK1, and MKK6, suggesting that these kinases may phosphorylate NS5A in mammalian cells. Significantly, rapamycin, an inhibitor commonly used to investigate the mTOR/p70S6K pathway, reduced the in vivo phosphorylation of specific NS5A phosphopeptides, strongly suggesting that p70S6 kinase and potentially related members of this group phosphorylate NS5A inside the cell. Curiously, certain of these kinases also play a major role in mRNA translation and antiapoptotic pathways, some of which are already known to be regulated by NS5A. The findings presented here demonstrate the use of high-throughput screening of the yeast kinome to facilitate the major task of identifying human NS5A protein kinases for further characterization of phosphorylation events in vivo. Our results suggest that this novel approach may be generally applicable to the screening of other protein biochemical activities by mechanistic class.
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PMID:High-throughput screening of the yeast kinome: identification of human serine/threonine protein kinases that phosphorylate the hepatitis C virus NS5A protein. 1501 73


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