scholarly journals Phosphoinositide 3-kinase: the key switch mechanism in insulin signalling

1998 ◽  
Vol 333 (3) ◽  
pp. 471-490 ◽  
Author(s):  
Peter R. SHEPHERD ◽  
Dominic J. WITHERS ◽  
Kenneth SIDDLE
Keyword(s):  
Protein Kinase ◽  
Insulin Signalling ◽  
Protein Substrates ◽  
Cellular Processes ◽  
Glucose And Lipid Metabolism ◽  
Wide Range ◽  
Dependent Protein ◽  
Switch Mechanism ◽  
Phosphoinositide 3 Kinase ◽  
Allosteric Regulators

Insulin plays a key role in regulating a wide range of cellular processes. However, until recently little was known about the signalling pathways that are involved in linking the insulin receptor with downstream responses. It is now apparent that the activation of class 1a phosphoinositide 3-kinase (PI 3-kinase) is necessary and in some cases sufficient to elicit many of insulin's effects on glucose and lipid metabolism. The lipid products of PI 3-kinase act as both membrane anchors and allosteric regulators, serving to localize and activate downstream enzymes and their protein substrates. One of the major ways these lipid products of PI 3-kinase act in insulin signalling is by binding to pleckstrin homology (PH) domains of phosphoinositide-dependent protein kinase (PDK) and protein kinase B (PKB) and in the process regulating the phosphorylation of PKB by PDK. Using mechanisms such as this, PI 3-kinase is able to act as a molecular switch to regulate the activity of serine/threonine-specific kinase cascades important in mediating insulin's effects on endpoint responses.

Signalling functions for sphingolipid long-chain bases in Saccharomyces cerevisiae

2005 ◽  
Vol 33 (5) ◽  
pp. 1170-1173 ◽  
Author(s):  
K. Liu ◽  
X. Zhang ◽  
C. Sumanasekera ◽  
R.L. Lester ◽  
R.C. Dickson
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Protein Kinase ◽  
Second Messengers ◽  
Dependent Protein Kinase ◽  
Yeast Saccharomyces Cerevisiae ◽  
Signalling Molecules ◽  
Cellular Processes ◽  
Wide Range ◽  
Dependent Protein ◽  
Long Chain

Over the past several years, studies of sphingolipid functions in the baker's yeast Saccharomyces cerevisiae have revealed that the sphingoid LCBs (long-chain bases), dihydrosphingosine and PHS (phytosphingosine), are important signalling molecules or second messengers under heat stress and during non-stressed conditions. LCBs are now recognized as regulators of AGC-type protein kinase (where AGC stands for protein kinases A, G and C) Pkh1 and Pkh2, which are homologues of mammalian phosphoinositide-dependent protein kinase 1. LCBs were previously shown to activate Pkh1 and Pkh2, which then activate the downstream protein kinase Pkc1. We have recently demonstrated that PHS stimulates Pkh1 to activate additional downstream kinases including Ypk1, Ypk2 and Sch9. We have also found that PHS acts downstream of Pkh1 and partially activates Ypk1, Ypk2 and Sch9. These kinases control a wide range of cellular processes including growth, cell wall integrity, stress resistance, endocytosis and aging. As we learn more about the cellular processes controlled by Ypk1, Ypk2 and Sch9, we will have a far greater appreciation of LCBs as second messengers.

scholarly journals Domain Swapping Used To Investigate the Mechanism of Protein Kinase B Regulation by 3-Phosphoinositide-Dependent Protein Kinase 1 and Ser473 Kinase

1999 ◽  
Vol 19 (7) ◽  
pp. 5061-5072 ◽  
Author(s):  
Mirjana Andjelković ◽  
Sauveur-Michel Maira ◽  
Peter Cron ◽  
Peter J. Parker ◽  
Brian A. Hemmings
Keyword(s):  
Protein Kinase ◽  
Kinase Inhibitors ◽  
Protein Kinase B ◽  
Second Messengers ◽  
Activation Mechanism ◽  
Dependent Protein Kinase ◽  
Dependent Protein ◽  
Phosphoinositide 3 Kinase ◽  
Regulatory Sites

ABSTRACT Protein kinase B (PKB or Akt), a downstream effector of phosphoinositide 3-kinase (PI 3-kinase), has been implicated in insulin signaling and cell survival. PKB is regulated by phosphorylation on Thr308 by 3-phosphoinositide-dependent protein kinase 1 (PDK1) and on Ser473 by an unidentified kinase. We have used chimeric molecules of PKB to define different steps in the activation mechanism. A chimera which allows inducible membrane translocation by lipid second messengers that activate in vivo protein kinase C and not PKB was created. Following membrane attachment, the PKB fusion protein was rapidly activated and phosphorylated at the two key regulatory sites, Ser473 and Thr308, in the absence of further cell stimulation. This finding indicated that both PDK1 and the Ser473 kinase may be localized at the membrane of unstimulated cells, which was confirmed for PDK1 by immunofluorescence studies. Significantly, PI 3-kinase inhibitors prevent the phosphorylation of both regulatory sites of the membrane-targeted PKB chimera. Furthermore, we show that PKB activated at the membrane was rapidly dephosphorylated following inhibition of PI 3-kinase, with Ser473 being a better substrate for protein phosphatase. Overall, the results demonstrate that PKB is stringently regulated by signaling pathways that control both phosphorylation/activation and dephosphorylation/inactivation of this pivotal protein kinase.

scholarly journals Proteomic Profile of Carbonylated Proteins Screen Regulation of Apoptosis via CaMK Signaling in Response to Regular Aerobic Exercise

2018 ◽  
Vol 2018 ◽  
pp. 1-13 ◽  
Author(s):  
Wenfeng Liu ◽  
Li Li ◽  
Heyu Kuang ◽  
Yan Xia ◽  
Zhiyuan Wang ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Aerobic Exercise ◽  
Protein Kinase B ◽  
Mammalian Target Of Rapamycin ◽  
Rat Striatum ◽  
Protective Effects ◽  
Sprague Dawley ◽  
Calcium Calmodulin Dependent ◽  
Dependent Protein ◽  
Phosphoinositide 3 Kinase

To research carbonylated proteins and screen molecular targets in the rat striatum on regular aerobic exercise, male Sprague-Dawley rats (13 months old, n = 24) were randomly divided into middle-aged sedentary control (M-SED) and aerobic exercise (M-EX) groups (n = 12 each). Maximum oxygen consumption (VO2max) gradually increased from 50%–55% to 65%–70% for a total of 10 weeks. A total of 36 carbonylated proteins with modified oxidative sites were identified by Electrospray Ionization-Quadrupole-Time of Flight-Mass Spectrometer (ESI-Q-TOF-MS), including 17 carbonylated proteins unique to the M-SED group, calcium/calmodulin-dependent protein kinase type II subunit beta (CaMKIIβ), and heterogeneous nuclear ribonucleoprotein A2/B1 (Hnrnpa2b1), among others, and 19 specific to the M-EX group, ubiquitin carboxyl-terminal hydrolase isozyme L1 (UCH-L1), and malic enzyme, among others. Regular aerobic exercise improved behavioral and stereological indicators, promoted normal apoptosis (P < 0.01), alleviated carbonylation of the CaMKIIβ and Hnrnpa2b1, but induced carbonylation of the UCH-L1, and significantly upregulated the expression levels of CaMKIIβ, CaMKIIα, and Vdac1 (p < 0.01) and Hnrnpa2b1 and UCH-L1 (p < 0.01), as well as the phosphoinositide 3-kinase/protein kinase B/mammalian target of rapamycin pathways (PI3K/Akt/mTOR) pathway-related genes Akt and mTOR. Regular aerobic exercise for 10 weeks (incremental for the first 6 weeks followed by constant loading for 4 weeks) enhanced carbonylation of CaMKIIβ, Hnrnpa2b1, and modulated apoptosis via activation of CaMK and phosphoinositide 3-kinase/protein kinase B/mTOR signaling. It also promoted normal apoptosis in the rat striatum, which may have protective effects in neurons.

scholarly journals Vanadium oxoanions and cAMP-dependent protein kinase: an anti-substrate inhibitor

1997 ◽  
Vol 321 (2) ◽  
pp. 333-339 ◽  
Author(s):  
Scott PLUSKEY ◽  
Mohammad MAHROOF-TAHIR ◽  
Debbie C. CRANS ◽  
David S. LAWRENCE
Keyword(s):  
Protein Kinase ◽  
Protein Kinases ◽  
Competitive Inhibitor ◽  
Serine Threonine Kinase ◽  
Dependent Protein Kinase ◽  
Phosphorylated Proteins ◽  
Camp Dependent Protein Kinase ◽  
Wide Range ◽  
Dependent Protein ◽  
Substrate Inhibitor

Vanadium oxoions have been shown to elicit a wide range of effects in biological systems, including an increase in the quantity of phosphorylated proteins. This response has been attributed to the inhibition of protein phosphatases, the indirect activation of protein kinases via stimulation of enzymes at early steps in signal transduction pathways and/or the direct activation of protein kinases. We have evaluated the latter possibility by exploring the effects of vanadate, decavanadate and vanadyl cation species on the activity of the cAMP-dependent protein kinase (PKA), a serine/threonine kinase. Vanadate, in the form of monomer, dimer, tetramer and pentamer species, neither inhibits nor activates PKA. In marked contrast, decavanadate is a competitive inhibitor (Ki = 1.8ŷ0.1 mM) of kemptide (Leu-Arg-Arg-Ala-Ser-Leu-Gly), a peptide-based substrate. This inhibition pattern is especially surprising, since the negatively charged decavanadate would not be predicted to bind to the region of the active site of the enzyme that accommodates the positively charged kemptide substrate. Our studies suggest that decavanadate can associate with kemptide in solution, which would prevent kemptide from interacting with the enzyme. Vanadium(IV) also inhibits the PKA-catalysed phosphorylation of kemptide, but with an IC50 of 366ŷ10 ƁM. However, in this case V4+ appears to bind to the Mg2+-binding site, since it can substitute for Mg2+. In the absence of Mg2+, the optimal concentration of vanadium(IV) for the PKA-catalysed phosphorylation of kemptide is 100 ƁM, with concentrations above 100 ƁM being markedly inhibitory. However, even at the optimal 100 ƁM V4+ concentration, the Vmax and Km values (for kemptide) are significantly less favourable than those obtained in the presence of 100 ƁM Mg2+. In summary, we have found that oxovanadium ions can directly alter the activity of the serine/threonine-specific PKA.

scholarly journals Where in the cell is SIRT3? – functional localization of an NAD+-dependent protein deacetylase

2008 ◽  
Vol 411 (2) ◽  
pp. e11-e13 ◽  
Author(s):  
William C. Hallows ◽  
Brittany N. Albaugh ◽  
John M. Denu
Keyword(s):  
Nuclear Localization ◽  
The Elderly ◽  
Active Protein ◽  
Cellular Processes ◽  
Age Related ◽  
Biochemical Journal ◽  
Wide Range ◽  
Functional Localization ◽  
Dependent Protein ◽  
Protein Deacetylase

Sirtuins are NAD+-dependent enzymes that have been implicated in a wide range of cellular processes, including pathways that affect diabetes, cancer, lifespan and Parkinson's disease. To understand their cellular function in these age-related diseases, identification of sirtuin targets and their subcellular localization is paramount. SIRT3 (sirtuin 3), a human homologue of Sir2 (silent information regulator 2), has been genetically linked to lifespan in the elderly. However, the function and localization of this enzyme has been keenly debated. A number of reports have indicated that SIRT3, upon proteolytic cleavage in the mitochondria, is an active protein deacetylase against a number of mitochondrial targets. In stark contrast, some reports have suggested that full-length SIRT3 exhibits nuclear localization and histone deacetylase activity. Recently, a report comparing SIRT3−/− and SIRT+/+ mice have provided compelling evidence that endogenous SIRT3 is mitochondrial and appears to be responsible for the majority of protein deacetylation in this organelle. In this issue of the Biochemical Journal, Cooper et al. present additional results that address the mitochondrial and nuclear localization of SIRT3. Utilizing fluorescence microscopy and cellular fractionation studies, Cooper et al. have shown that SIRT3 localizes to the mitochondria and is absent in the nucleus. Thus this study provides additional evidence to establish SIRT3 as a proteolytically modified, mitochondrial deacetylase.

scholarly journals Fyn Regulates Binding Partners of Cyclic-AMP Dependent Protein Kinase A

Proteomes ◽  
2018 ◽  
Vol 6 (4) ◽  
pp. 37
Author(s):  
Anna Schmoker ◽  
Samuel Barritt ◽  
Marion Weir ◽  
Jacqueline Mann ◽  
Tyler Hogan ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Quantitative Mass Spectrometry ◽  
Serine Threonine Kinase ◽  
Dependent Protein Kinase ◽  
Threonine Kinase ◽  
Cellular Processes ◽  
Binding Partners ◽  
Dependent Protein ◽  
Kinase A

The cAMP-dependent protein kinase A (PKA) is a serine/threonine kinase involved in many fundamental cellular processes, including migration and proliferation. Recently, we found that the Src family kinase Fyn phosphorylates the catalytic subunit of PKA (PKA-C) at Y69, thereby increasing PKA kinase activity. We also showed that Fyn induced the phosphorylation of cellular proteins within the PKA preferred target motif. This led to the hypothesis that Fyn could affect proteins in complex with PKA. To test this, we employed a quantitative mass spectrometry approach to identify Fyn-dependent binding partners in complex with PKA-C. We found Fyn enhanced the binding of PKA-C to several cytoskeletal regulators that localize to the centrosome and Golgi apparatus. Three of these Fyn-induced PKA interactors, AKAP9, PDE4DIP, and CDK5RAP2, were validated biochemically and were shown to exist in complex with Fyn and PKA in a glioblastoma cell line. Intriguingly, the complexes formed between PKA-C and these known AKAPs were dependent upon Fyn catalytic activity and expression levels. In addition, we identified Fyn-regulated phosphorylation sites on proteins in complex with PKA-C. We also identified and biochemically validated a novel PKA-C interactor, LARP4, which complexed with PKA in the absence of Fyn. These results demonstrate the ability of Fyn to influence the docking of PKA to specific cellular scaffolds and suggest that Fyn may affect the downstream substrates targeted by PKA.

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