Phosphatidylinositol 3-kinase regulates Raf1 through Pak phosphorylation of serine 338

Chaudhary, Ajay Kumar; King, W. G.; Mattaliano, Mark D.; Frost, Jeffrey A.; Diaz, Bruce; Morrison, Deborah K.; Cobb, Melanie H.; Marshall, Mark S.; Brugge, Joan S.

doi:10.1016/s0960-9822(00)00475-9

Cited by 206 publications

(146 citation statements)

References 18 publications

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“…PI3K has been shown to stimulate integrin-mediated Raf activation in synergy with Ras (Chaudhary et al, 2000). Cooperative effects between these two signaling cascades have also been demonstrated in the regulation of the platelet-derived growth factor-induced proliferation (Choudhury et al, 1997) or in cell cycle progression and transformation (Sheng et al, 2001a, b).…”

Section: Discussionmentioning

confidence: 99%

Akt Down-Regulates ERK1/2 Nuclear Localization and Angiotensin II-induced Cell Proliferation through PEA-15

Gervais

Dugourd

Muller

et al. 2006

MBoC

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Angiotensin II (AngII) type 1 receptors (AT1) regulate cell growth through the extracellular signal-regulated kinase (ERK)1/2 and phosphatidylinositol 3-kinase (PI3K) pathways. ERK1/2 and Akt/protein kinase B, downstream of PI3K, are independently activated but both required for mediating AngII-induced proliferation when expressed at endogenous levels. We investigate the effect of an increase in the expression of wild-type Akt1 by using Chinese hamster ovary (CHO)-AT1 cells. Unexpectedly, Akt overexpression inhibits the AT1-mediated proliferation. This effect could be generated by a cross-talk between the PI3K and ERK1/2 pathways. A functional partner is the phosphoprotein enriched in astrocytes of 15 kDa (PEA-15), an Akt substrate known to bind ERK1/2 and to regulate their nuclear translocation. We report that Akt binds to PEA-15 and that Akt activation leads to PEA-15 stabilization, independently of PEA-15 interaction with ERK1/2. Akt cross-talk with PEA-15 does not affect ERK1/2 activation but decreases their nuclear activity as a result of the blockade of ERK1/2 nuclear accumulation. In response to AngII, PEA-15 overexpression displays the same functional consequences on ERK1/2 signaling as Akt overactivation. Thus, Akt overactivation prevents the nuclear translocation of ERK1/2 and the AngII-induced proliferation through interaction with and stabilization of endogenous PEA-15.

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Section: Discussionmentioning

confidence: 99%

Akt Down-Regulates ERK1/2 Nuclear Localization and Angiotensin II-induced Cell Proliferation through PEA-15

Gervais

Dugourd

Muller

et al. 2006

MBoC

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“…The recent suggestion that P13-K and Pak3 regulate Raf-1 activity through S338 phosphorylation (7,25,49) a good deal of attention. We have tested this model using a monoclonal antibody that is highly specific for S338 phosphorylation on Raf-1 (40) and found, in contradiction to the conclusions of Sun et al (49), that S338 phosphorylation could still be observed when P13-K activity was inhibited.…”

Section: Discussionmentioning

confidence: 99%

“…There is much interest in the cross talk that exists between these different, parallel Ras pathways, and in some but not all cell types, inhibition of Pl3-K that leads to suppression of ERK has been shown (8,16,24,56). In different studies, Pl3-K has been shown to regulate ERK activation at the level of Ras or at the level of Raf-1 (7,8,27,56). More recently, it was suggested that Pl3-K regulates Raf-1 by activating protein kinases that can phosphorylate Raf-1 directly.…”

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confidence: 99%

S338 Phosphorylation of Raf-1 Is Independent of Phosphatidylinositol 3-Kinase and Pak3

Chiloeches

Mason

Marais

2001

Molecular and Cellular Biology

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The Raf-1 serine/threonine protein kinase requires phosphorylation of the serine at position 338 (S338) for activation. Ras is required to recruit Raf-1 to the plasma membrane, which is where S338 phosphorylation occurs. The recent suggestion that Pak3 could stimulate Raf-1 activity by directly phosphorylating S338 through a Ras/phosphatidylinositol 3-kinase (Pl3-K)/-Cdc42-dependent pathway has attracted much attention. Using a phospho-specific antibody to S338, we have reexamined this model. Using LY294002 and wortmannin, inhibitors of Pl3-K, we find that growth factor-mediated S338 phosphorylation still occurs, even when Pl3-K activity is completely blocked. Although high concentrations of LY294002 and wortmannin did suppress S338 phosphorylation, they also suppressed Ras activation. Additionally, we show that Pak3 is not activated under conditions where S338 is phosphorylated, but when Pak3 is strongly activated, by coexpression with V12Cdc42 or by mutations that make it independent of Cdc42, it did stimulate S338 phosphorylation. However, this occurred in the cytosol and did not stimulate Raf-1 kinase activity. The inability of Pak3 to activate Raf-1 was not due to an inability to stimulate phosphorylation of the tyrosine at position 341 but may be due to its inability to recruit Raf-1 to the plasma membrane. Taken together, our data show that growth factorstimulated Raf-1 activity is independent of Pl3-K activity and argue against Pak3 being a physiological mediator of S338 phosphorylation in growth factor-stimulated cells.The Raf-1 serine/threonine-specific protein kinase is the first component of a three-tiered protein kinase cascade that regulates many biological events such as cell growth, differentiation, and apoptosis (for reviews, see references 12, 39, and 44). Raf-1 phosphorylates and activates the dual-specificity mitogen-activated protein kinase (MAPK) kinases MEK1 and MEK2, which in turn activate the MAPKs ERK1 and ERK2. The ERKs phosphorylate and regulate the activity of transcription factors, cytoskeletal proteins, metabolic enzymes, and other protein kinases to modulate cellular responses to extracellular signals. Raf-1 regulation is highly complex. It is cytosolic in unstimulated cells, but following activation of the small G-protein Ras, it translocates to the plasma membrane, where activation takes place (for reviews, see references 23, 38, and 41). Interaction with Ras alone is not sufficient to activate Raf-1 and other membrane-localized events such as oligomerization, interaction with other proteins, and interactions with lipids all appear to play a role.Phosphorylation also plays a key role in Raf-1 activation, and both positive and negative regulatory sites have been mapped (23,38,41). Two sites whose phosphorylation has been shown to be necessary for activation are the serine located at position 338 (S338) and the tyrosine located at position 341 (Y341) (3,15,18,36,40,42). These amino acids are located 10 to 15 amino acids N terminal to the glycine-rich loop of the ATP-binding domain...

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“…5 Phosphorylation at S338 is critical for Raf-1 activation and ERK stimulation by a variety of factors including activated integrins and growth factors. 6,7 Maximal Raf-1 activation is achieved by concurrent phosphorylation at Y341 and S338. 8 While concordant data indicates Src as the kinase phosphorylating Raf-1 at Y341, it is still controversial which is the kinase phosphorylating Raf-1 at S338.…”

Section: Introductionmentioning

confidence: 99%

“…9 Phosphatidyl-inositol-3-kinase (PI3-K) inhibitors (Wortmannin and Ly-294002) were shown to inhibit both Raf-1 kinase activity and its phosphorylation at S338, stimulated by EGF and by integrins in COS-7 cells. 6,10 However, a careful re-examination of those results raised some doubts on the effective role proposed for Pak3. Chiloeches et al argued against the physiological role proposed for Pak3, mainly because supporting data were obtained by overexpression of recombinant factors or by using PI3-K inhibitors at concentrations that block also Ras activity.…”

Section: Introductionmentioning

confidence: 99%

Calcium/calmodulin-dependent protein kinase II (CaMKII) phosphorylates Raf-1 at serine 338 and mediates Ras-stimulated Raf-1 activation

et al. 2012

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The calcium/calmodulin-dependent kinase II (CaMKII) participates with Ras to Raf-1 activation, and it is necessary for activation of the extracellular signal-regulated kinase (ERK) by different factors in epithelial and mesenchimal cells. Raf-1 activation is a complex multistep process, and its maximal activation is achieved by phosphorylation at Y341 by Src and at S338 by other kinase/s. Although early data proposed the involvement of p21-activated kinase 3 (Pak3), the kinase phosphorylating S338 remains to be definitively identified. In this study, we verified the hypothesis that CaMKII phosphorylates Raf-1 at Ser338. To do so, we determined the role of CaMKII in Raf-1 and ERK activation by oncogenic Ras and other factors. Serum, fibronectin, Src(Y527) and Ras(V12) activated CaMKII and ERK, at different extents. The inhibition of CaMKII attenuated Raf-1 and ERK activation by all these factors. CaMKII was also necessary for the phosphorylation of Raf-1 at S338 by serum, fibronectin and Ras. Conversely, inhibition of Pak3 activation by blocking phosphatidylinositol-3-kinase was ineffective. The direct phosphorylation of S338 Raf-1 by CaMKII was demonstrated in vitro by interaction of purified kinases. These results demonstrate that Ras activates CaMKII, which, in turn, phosphorylates Raf-1 at S338 and participates in ERK activation upon different stimuli

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Phosphatidylinositol 3-kinase regulates Raf1 through Pak phosphorylation of serine 338

Cited by 206 publications

References 18 publications

Akt Down-Regulates ERK1/2 Nuclear Localization and Angiotensin II-induced Cell Proliferation through PEA-15

Akt Down-Regulates ERK1/2 Nuclear Localization and Angiotensin II-induced Cell Proliferation through PEA-15

S338 Phosphorylation of Raf-1 Is Independent of Phosphatidylinositol 3-Kinase and Pak3

Calcium/calmodulin-dependent protein kinase II (CaMKII) phosphorylates Raf-1 at serine 338 and mediates Ras-stimulated Raf-1 activation

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