Activation of an S6/H4 Kinase (PAK 65) from Human Placenta by Intramolecular and Intermolecular Autophosphorylation

Benner, Gretchen E.; Dennis, Patrick B.; Masaracchia, Ruthann A.

doi:10.1074/jbc.270.36.21121

Cited by 69 publications

(80 citation statements)

References 25 publications

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“…Consistent with this proposal, autophosphorylation of the 54-kDa C-terminal fragment of native kinase (19) and of the expressed catalytic domain (data not shown) are much faster than autophosphorylation of native kinase. This explanation would also be consistent with the need for phosphorylation of a second residue in the catalytic region of S6͞H4 kinase prepared from the holoenzyme (21) extension N terminal to the catalytic domain, and for activation by autophosphorylation in vitro of kinases such as Rhoactivated protein kinase N (26), in which the residue corresponding to MIHCK Ser-627 seems to be constitutively phosphorylated.…”

Section: Discussionmentioning

confidence: 49%

“…The activities of many other serine͞threonine kinases have been shown to be dependent on phosphorylation of Ser or Thr residues at the same or neighboring position(s). These include two other members of the PAK family (Table 2): S6͞H4 kinase from human placenta (21) and, as inferred from its activation by substitution of Glu for the putative phosphorylation site (T422E), rat brain ␣-PAK (22). However, the two corresponding Thr residues of a third member of the PAK͞ STE20 family, yeast Ste20p (Table 2), can both be replaced by Ala residues in a double mutant (T772A, T773A) with no phenotypic effect in vivo or effect on autophosphorylation in vitro whereas the T777A mutant is inactive in both assays (23).…”

Section: Discussionmentioning

confidence: 99%

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Identification by mass spectrometry of the phosphorylated residue responsible for activation of the catalytic domain of myosin I heavy chain kinase, a member of the PAK/STE20 family

Szczepanowska

Zhang

Herring

et al. 1997

Proc. Natl. Acad. Sci. U.S.A.

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Biochemistry. In the article ''Identification by mass spectrometry of the phosphorylated residue responsible for activation of the catalytic domain of myosin I heavy chain kinase, a member of the PAK͞STE20 family'' by Joanna Szczepanowska, Xiaolong Zhang, Christopher J. Herring, Jun Qin, Edward D. Korn, and Hanna Brzeska, which appeared in number 16, August 5, 1997, of Proc. Natl. Acad. Sci. USA (94,(8503)(8504)(8505)(8506)(8507)(8508), the authors wish to note that in Fig. 3, the ions of m͞z 1345.3 and 1247.1 were incorrectly identified as b 13 and b 13 ⌬ , respectively, produced by cleavage of the peptide AS(Pi)VVGTTYW-MAPEVVK between E and V (Fig. 3 Inset). In fact, these ions are b 12 and b 12 ⌬ , produced by cleavage of the peptide between P and E. This correction has no effect on the conclusion that the phosphorylated residue is serine. Also, in Table 2, reference numbers 22-24 should be 21-23 (the reference in the legend is cited correctly) and the MIHCK sequence is from residue 624 to residue 638.Cell Biology. In the article ''Subtraction hybridization identifies a transformation progression-associated gene PEG-3 with sequence homology to a growth arrest and DNA damageinducible gene'' by Zao

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Section: Discussionmentioning

confidence: 49%

Section: Discussionmentioning

confidence: 99%

Identification by mass spectrometry of the phosphorylated residue responsible for activation of the catalytic domain of myosin I heavy chain kinase, a member of the PAK/STE20 family

Szczepanowska

Zhang

Herring

et al. 1997

Proc. Natl. Acad. Sci. U.S.A.

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“…In vitro kinase assays showed hpak1-(1-234) is capable to inhibit GST-rpak1-(233-544) kinase activity (Fig. 1D, lanes [3][4][5][6]. With increasing amounts of the NH 2 terminus, autophosphorylation and phosphorylation of myelin basic protein were decreased.…”

Section: Interaction Between the Pak1 Kinase Domain And A Nh 2 -mentioning

confidence: 99%

“…Several lines of evidence suggested that the amino-terminal nonkinase region of Pak, in which the Cdc42/Rac-binding site is located, is crucial for the regulation of kinase activity. It has been shown by several groups that removal of the NH 2 -terminal portion of Pak by protease digestion leads to activation of the kinase fragment (4,5). A physiologically relevant example is known for the 62-kDa isoform Pak2, which has been shown to be cleaved and activated by the cysteine protease caspase-3 in response to apoptosis-inducing stimuli (6 -8).…”

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confidence: 99%

Identification of a Central Phosphorylation Site in p21-activated Kinase Regulating Autoinhibition and Kinase Activity

Zenke¹,

King²,

Bohl

et al. 1999

Journal of Biological Chemistry

225

221

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p21-activated kinases (Pak)/Ste20 kinases are regulated in vitro and in vivo by the small GTP-binding proteins Rac and Cdc42 and lipids, such as sphingosine, which stimulate autophosphorylation and phosphorylation of exogenous substrates. The mechanism of Pak activation by these agents remains unclear. We investigated Pak kinase activation in more detail to gain insight into the interplay between the GTPase/sphingosine binding, an intramolecular inhibitory interaction, and autophosphorylation. We present biochemical evidence that an autoinhibitory domain (ID) contained within amino acid residues 67-150 of Pak1 interacts with the carboxyl-terminal kinase domain and that this interaction is regulated in a GTPase-dependent fashion. Cdc42-and sphingosine-stimulated Pak1 activity can be inhibited in trans by recombinant ID peptide, indicating similarities in their mode of activation. However, Pak1, which was autophosphorylated in response to either GTPase or sphingosine, is highly active and is insensitive to inhibition by the ID peptide. We identified phospho-acceptor site threonine 423 in the kinase activation loop as a critical determinant for the sensitivity to autoinhibition and enzymatic activity. Phosphorylation studies suggested that the stimulatory effect of both GTPase and sphingosine results in exposure of the activation loop, making it accessible for intermolecular phosphorylation.Localized regulation of protein kinase activity is an essential means to ensure spatial and temporal control of signaling events in a cellular environment. Hormonal or other stimuli are usually necessary to switch a kinase into a catalytically competent state, allowing phosphorylation of substrates to take place. An emerging regulatory theme is that inhibitory mechanisms exist to keep protein kinases in an inactive state (1, 2), and that relief of such inhibition allows activation to occur. Kinases often act autocatalytically to phosphorylate key amino acid residues that relieve autoinhibition and enhance catalytic efficiency. Alternatively, exogenous kinases may also serve this role. However, activation must be reversed in the absence of the stimulus, and dephosphorylation by protein phosphatases is thought to mediate switching the active kinase back to an inactive or basally activated state. p21-activated kinases (Paks) 1 belong to a growing family of serine/threonine kinases involved in the control of various cellular processes, including the cell cycle, dynamics of the cytoskeleton, apoptosis, and transcription (3). Pak kinase activity is regulated by members of the Rho family of GTPases, specifically Cdc42 and Rac. These GTPases bind to Pak kinase solely in their active forms, i.e. the GTP-bound state, resulting in stimulation of the kinase activity both in vitro and in vivo. The molecular details of how the GTPases exert their effect on the kinase to induce its activation remain unclear, however. Several lines of evidence suggested that the amino-terminal nonkinase region of Pak, in which the Cdc42/Rac-binding site is loca...

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“…In addition to its effects on the actin cytoskeleton, PAKs may play a role in disassembling intermediate filaments composed of desmin (21). Studies on PAK as a protease-activated kinase (22) indicated that the catalytic domain needs to undergo ATP-dependent autotransphosphorylation prior to conversion to its active form (23). The behavior of PAK as a dimer in solution (9) suggests that transphosphorylation of the kinase can be rapid.…”

mentioning

confidence: 99%

The Mechanism of PAK Activation

Chong

Tan

Lim

et al. 2001

Journal of Biological Chemistry

219

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The p21-activated kinases (PAKs), in common with many kinases, undergo multiple autophosphorylation events upon interaction with appropriate activators. The Cdc42-induced phosphorylation of PAK serves in part to dissociate the kinase from its partners PIX and Nck. Here we investigate in detail how autophosphorylation events affect the catalytic activity of PAK by altering the autophosphorylation sites in both ␣-and ␤PAK. Both in vivo and in vitro analyses demonstrate that, although most phosphorylation events in the PAK Nterminal regulatory domain play no direct role in activation, a phosphorylation of ␣PAK serine 144 or ␤PAK serine 139, which lie in the kinase inhibitory domain, significantly contribute to activation. By contrast, sphingosine-mediated activation is independent of this residue, indicating a different mode of activation. Thus two autophosphorylation sites direct activation while three others control association with focal complexes via PIX and Nck.

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Activation of an S6/H4 Kinase (PAK 65) from Human Placenta by Intramolecular and Intermolecular Autophosphorylation

Cited by 69 publications

References 25 publications

Identification by mass spectrometry of the phosphorylated residue responsible for activation of the catalytic domain of myosin I heavy chain kinase, a member of the PAK/STE20 family

Identification by mass spectrometry of the phosphorylated residue responsible for activation of the catalytic domain of myosin I heavy chain kinase, a member of the PAK/STE20 family

Identification of a Central Phosphorylation Site in p21-activated Kinase Regulating Autoinhibition and Kinase Activity

The Mechanism of PAK Activation

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