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

---

Query: EC:2.7.11.1 (protein kinase)
81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In order to examine a possible role of protein kinases in the signal transduction of platelet activation, thrombin-stimulated human platelets were analyzed for protein kinase activity with a denaturation/renaturation method. Treatment of platelets with thrombin resulted in a rapid activation of a 33-kDa protein kinase (PK33) using casein as an in vitro substrate. The concentration of thrombin to activate PK33 was proportional to that required to induce platelet aggregation. PK33 was also activated by a thrombin receptor agonist peptide, but not by hirudin-treated or diisopropylphosphate-inactivated thrombin. Phosphoamino acid analysis showed that PK33 is a serine/threonine kinase. Comparative analysis using specific substrate and inhibitors revealed that PK33 is distinct from casein kinase I, casein kinase II, P34cdc2, and mitogen activated protein kinase. These findings suggest that platelet activation mediated by thrombin receptor requires PK33 activation.
...
PMID:The thrombin receptor transmits signals through a 33-kDa protein kinase in human platelets. 809 74

Treatment of quiescent rat aortic smooth muscle cells with either alpha-thrombin or a thrombin receptor-derived agonist peptide (SFLLRNP) resulted in pronounced increases in [3H]thymidine incorporation that were concentration dependent and reached a maximum of approximately 15-fold above serum-starved controls. However, in contrast to FBS, PDGF-BB, or basic fibroblast growth factor (bFGF), that initiated DNA synthesis promptly after 16-19 h, thymidine incorporation in response to thrombin was delayed by an additional 3-6 h. Delayed mitogenesis correlated with the appearance of a potent mitogenic activity in conditioned media samples obtained from thrombin-stimulated rat aortic smooth muscle cells, as assayed using Swiss 3T3 fibroblasts. This activity was not inhibited by neutralizing antibodies directed against PDGF or bFGF. Furthermore, in the Swiss 3T3 cells, simple addition of either alpha-thrombin or SFLLRNP failed to elicit a significant mitogenic response. In signal transduction studies, both thrombin and SFLLRNP treatment led to rapid tyrosine phosphorylation of proteins with apparent molecular masses of 42, 44, 75, 120, and 190 kD, respectively, as assessed by antiphosphotyrosine immunoblotting. The overall pattern of protein tyrosine phosphorylation was distinct from that observed after PDGF-BB addition. Activation of Raf-1 and the mitogen-activated protein (MAP) kinases p44mapk and p42mapk was also observed. However, the time course and duration of Raf-1/MAP kinase activation after thrombin stimulation were similar to those elicited by PDGF-BB. Taken together, our results indicate that thrombin-stimulated vascular smooth muscle proliferation is delayed and requires the de novo expression of one or more autocrine mitogens. In addition, the rapid induction of discrete intracellular signaling mechanisms by thrombin, including the Raf-1/MAP kinase pathway, appears to be insufficient alone to promote vascular smooth muscle cell mitogenesis.
...
PMID:Thrombin receptor activation elicits rapid protein tyrosine phosphorylation and stimulation of the raf-1/MAP kinase pathway preceding delayed mitogenesis in cultured rat aortic smooth muscle cells: evidence for an obligate autocrine mechanism promoting cell proliferation induced by G-protein-coupled receptor agonist. 863 28

Thrombin is one of the first regulatory molecules present at sites of CNS trauma or injury. Exposure of neuronal and glial cells to thrombin produces potent morphological as well as cytoprotective and cytotoxic effects, but little is known about how this important modulator affects neurotransmitter signaling. In astrocyte cultures that have been morphologically differentiated by exposure to transforming growth factor-alpha, addition of thrombin induced a retraction of astrocytic processes and suppressed the stimulation of phosphoinositide hydrolysis by the selective metabotropic glutamate receptor (mGluR) agonist 1-aminocyclopentane-1S,3R-dicarboxylic acid. In addition to the suppression of phosphoinositide hydrolysis, thrombin treatment produced a corresponding reduction in level of mGluR5 mRNA as demonstrated with ribonuclease protection assay and reduced content of mGluR5 receptor protein as seen with western blotting. In contrast, thrombin exposure up-regulated astrocyte beta-actin mRNA levels. A synthetic hexapeptide with a sequence corresponding to the amino-terminus of the thrombin receptor's tethered ligand also mimicked the ability of thrombin to suppress mGluR5 levels and to increase beta-actin mRNA content, suggesting that these effects of thrombin are mediated by proteolytically activated cell surface thrombin receptors. Thrombin's suppressive effect on mGluR5 was resistant to pretreatment with pertussis toxin or various protein kinase and protein phosphatase inhibitors. However, the serine/threonine protein kinase inhibitor H-7 did prevent thrombin-induced reversal of astrocyte stellation and induction of beta-actin mRNA levels, indicating that these effects of thrombin involve a signaling pathway distinct from the one that mediates the suppressive effects of thrombin on mGluR5.
...
PMID:Exposure of astrocytes to thrombin reduces levels of the metabotropic glutamate receptor mGluR5. 885 25

Protein kinase C (PKC) is a family of serine/threonine protein kinase isoforms that is important to intracellular enzymes for both tyrosine kinase receptors and G protein coupled receptors. However, which isoforms are linked to which class of receptors in endothelial cell signaling is not known. Moreover, the PKC isoforms in endothelial cells have not been thoroughly characterized. We tested the hypothesis that specific PKC isoforms are involved in different signaling pathways. PKC isoform expression was assessed by using reverse transcription polymerase chain reaction and Western blotting. The spatial distribution of PKC after stimulation of the cells with basic fibroblast growth factor (bFGF) and thrombin was examined by using confocal microscopy. Expression of PKC alpha, delta, epsilon, theta, and zeta was detectable on both the mRNA and protein levels. In resting cells, PKC alpha and epsilon were mostly distributed in the cytosol, while PKC alpha and epsilon were also present in the nucleus. Nuclear immunoreactivity of PKC alpha and epsilon increased significantly between passages 1 and 3. The phorbol ester TPA induced a rearrangement of PKC delta and a translocation of PKC alpha and epsilon to the nucleus. Treatment of endothelial cells with TPA for 24 hours caused PKC alpha, delta, and epsilon to disappear, while PKC zeta was not influenced by TPA. bFGF induced a rapid assembly of PKC alpha along cytosolic structures, followed by a translocation of the isoform toward the perinuclear region and into the nucleus. bFGF had a smaller effect on PKC epsilon. In contrast, thrombin had a similar effect on nuclear translocation of PKC alpha, did not influence PKC epsilon, and induced a rapid nuclear translocation of PKC zeta. Thus, tyrosine kinase receptor activation via bFGF induced a rapid association of PKC alpha and epsilon with nuclear structures, while activation of the G protein-coupled thrombin receptor increased mostly nuclear PKC zeta. The translocation of PKC isoforms into the nucleus by growth-promoting factors may be important for the induction of endothelial cell growth.
...
PMID:Endothelial cell tyrosine kinase receptor and G protein-coupled receptor activation involves distinct protein kinase C isoforms. 896 26

We examined the effects of platelet activators and inhibitors of platelet function on the voltage-gated delayed rectifier K+ current of human megakaryocytes. We found that both the activators such as thrombin, the thrombin receptor peptide (TRP42-47) and ADP and the inhibitors such as prostacyclin suppressed the delayed rectifier current through two different mechanisms. The cAMP dependent protein kinase (A-kinase) inhibitor IP20 blocked the suppression of the delayed rectifier current by prostacyclin and failed to block the suppression by thrombin, TRP42-47 and ADP. The effects of IP20 suggest that the action of prostacyclin is mediated by A-kinase and the action of the three activators is not mediated by A-kinase. Pertussis toxin (PTX) an inhibitor of the inhibitory GTP-binding proteins (Gi) blocked the suppression of the delayed rectifier current by thrombin, TRP42-47 and ADP and failed to block the suppression by prostacyclin. The effects of PTX suggests that the action of the three activators is mediated by Gi or some other PTX-sensitive GTP-binding protein. We speculate that thrombin and other platelet activators that activate Gi may be suppressing the delayed rectifier current via a direct interaction of Gi or a subunit of it with the delayed rectifier potassium channel itself.
...
PMID:Suppression of the voltage-gated K+ current of human megakaryocytes by thrombin and prostacyclin. 906 Oct 4

In order to examine a possible role of protein kinases in the signal transduction of platelet activation, thrombin-stimulated human platelets were analyzed for protein kinase activity with a denaturation/renaturation method. Treatment of platelets with thrombin resulted in a rapid activation of a 33-kDa protein kinase (PK33) using casein as an in vitro substrate. The concentration of thrombin to activate PK33 was proportional to that required to induce platelet aggregation. PK33 was also activated by a thrombin receptor agonist peptide, but not by hirudin-treated or diisopropylphosphate-inactivated thrombin. Phosphoamino acid analysis showed that PK33 is a serine/threonine kinase. Comparative analysis using specific substrate and inhibitors revealed that PK33 is distinct from casein kinase I, casein kinase II, P34cdc2, and mitogen activated protein kinase. These findings suggest that platelet activation mediated by thrombin receptor requires PK33 activation.
...
PMID:Thrombin-induced activation of the novel 33-kDa serine/threonine kinase in human platelets. 911 37

Plasmalemmal vesicles (PVs) or caveolae are plasma membrane invaginations and associated vesicles of regular size and shape found in most mammalian cell types. They are particularly numerous in the continuous endothelium of certain microvascular beds (e.g., heart, lung, and muscles) in which they have been identified as transcytotic vesicular carriers. Their chemistry and function have been extensively studied in the last years by various means, including several attempts to isolate them by cell fractionation from different cell types. The methods so far used rely on nonspecific physical parameters of the caveolae and their membrane (e.g., size-specific gravity and solubility in detergents) which do not rule out contamination from other membrane sources, especially the plasmalemma proper. We report here a different method for the isolation of PVs from plasmalemmal fragments obtained by a silica-coating procedure from the rat lung vasculature. The method includes sonication and flotation of a mixed vesicle fraction, as the first step, followed by specific immunoisolation of PVs on anticaveolin-coated magnetic microspheres, as the second step. The mixed vesicle fraction, is thereby resolved into a bound subfraction (B), which consists primarily of PVs or caveolae, and a nonbound subfraction (NB) enriched in vesicles derived from the plasmalemma proper. The results so far obtained indicate that some specific endothelial membrane proteins (e.g., thrombomodulin, functional thrombin receptor) are distributed about evenly between the B and NB subfractions, whereas others are restricted to the NB subfraction (e.g., angiotensin converting enzyme, podocalyxin). Glycoproteins distribute unevenly between the two subfractions and antigens involved in signal transduction [e.g., annexin II, protein kinase C alpha, the G alpha subunits of heterotrimeric G proteins (alpha s, alpha q, alpha i2, alpha i3), small GTP-binding proteins, endothelial nitric oxide synthase, and nonreceptor protein kinase c-src] are concentrated in the NB (plasmalemma proper-enriched) subfraction rather than in the caveolae of the B subfraction. Additional work should show whether discrepancies between our findings and those already recorded in the literature represent inadequate fractionation techniques or are accounted for by chemical differentiation of caveolae from one cell type to another.
...
PMID:Immunoisolation and partial characterization of endothelial plasmalemmal vesicles (caveolae). 924 41

While activation of tyrosine kinase growth factor receptors is accompanied by hyperphosphorylation of Raf-1, stimulation of receptors coupled to G-proteins has in some cases been shown to result in activation of a non-Raf MEKK rather than of Raf itself. Our finding (Weiss, R. H., and Nuccitelli, R. (1992) J. Biol. Chem. 267, 5608-5613) that the thrombin receptor requires tyrosine phosphorylation for its mitogenic effect in vascular smooth muscle cells led us to search for the molecules which are being tyrosine phosphorylated by this receptor. To determine whether mitogenic signalling of G-protein-coupled growth factor receptors results in tyrosine phosphorylation of Raf, we examined activation of Raf by two such receptors. Both thrombin and angiotensin II are mitogenic in NIH3T3 cells, but only thrombin causes hyperphosphorylation of Raf-1. Activation of Raf by thrombin is associated with phosphorylation of Raf-1 on tyrosine residues, whereas activation of Raf by angiotensin II does not involve significant tyrosine phosphorylation. However, Shc is tyrosine phosphorylated by both thrombin and angiotensin II. Thus, there exists a divergence in the mitogenic signalling pathways of the G-protein-coupled receptors relative to the Raf signalling cascade. While both thrombin and angiotensin II phosphorylate Shc and activate Raf catalytic activity, only thrombin phosphorylated Raf-1 on tyrosine. This signalling through disparate Raf-coupled pathways suggests one means by which the G-protein-coupled receptors may confer specificity in their signalling properties.
...
PMID:Divergence in the G-protein-coupled receptor mitogenic signalling pathway at the level of Raf kinase. 937 25

Phosphatidylinositol 3-kinase (PI3K) is a heterodimer lipid kinase consisting of an 85-kD subunit bound to a 110-kD catalytic subunit that also possesses intrinsic, Mn(2+)-dependent protein serine kinase activity capable of phosphorylating the 85-kD subunit. Here, we examine the Mn(2+)-dependent protein kinase activity of PI3K alpha immunoprecipitated from normal resting or thrombin-stimulated platelets, and characterize p85/p110 phosphorylation, in vitro. Phosphoamino acid analysis of phosphorylated PI3K alpha showed p85 and p110 were phosphorylated on serine, but in contrast to previous results, were also phosphorylated on threonine and tyrosine. Wortmannin and LY294002 inhibited p85 phosphorylation; however, p110 phosphorylation was also inhibited suggesting p110 autophosphorylation on serine/threonine. The protein tyrosine kinase inhibitor, erbstatin analog, partially inhibited p85 and p110 phosphorylation but did not appear to affect PI3K lipid kinase activity. The in vitro phosphorylation of p85 alpha or p110 alpha derived from thrombin-stimulated platelets was no different than that of resting platelets, but we confirm that in thrombin receptor-stimulated platelets enhanced levels of p85 alpha and PI3K lipid kinase activity were recovered in antiphosphotyrosine antibody immunoprecipitates. These results suggest PI3K alpha can autophosphorylate on serine and threonine, and both p85 alpha and p110 alpha are substrates for a constitutively-associated protein tyrosine kinase in platelets.
...
PMID:The p85 and p110 subunits of phosphatidylinositol 3-kinase-alpha are substrates, in vitro, for a constitutively associated protein tyrosine kinase in platelets. 944 54

Dramatic transient changes resulting in a stellate morphology are induced in many cell types on treatment with agents that enhance intracellular cAMP levels. Thrombin fully protects cells from this inductive effect of cAMP through the thrombin receptor. The protective effect of thrombin was shown to be Rho-dependent. Clostridium botulinum C3 exoenzyme, which inactivates RhoA functions, abolished the ability of thrombin to protect cells from responding to increased cAMP levels. A constitutively activated RhoAV14 mutant protein also prevented cells from responding to cAMP. RhoA can be specifically phosphorylated at Ser-188 by the cAMP-activated protein kinase A (PKA). We demonstrate that RhoAV14A188, which cannot be phosphorylated by PKA in vitro, is more effective than RhoAV14 in preventing cells from responding to cAMP and in inducing actin stress fiber formation. This suggests that PKA phosphorylation of RhoA impairs its biological activity in vivo. ROKalpha, a RhoA-associated serine/threonine kinase can also prevent cells from responding to cAMP with shape changes. Phosphorylation of RhoA by PKA in vitro decreases the binding of RhoA to ROKalpha. These results indicate that RhoA and cAMP have antagonistic roles in regulating cellular morphology and suggest that cAMP-mediated down-regulation of RhoA binding to its effector ROKalpha may be involved in this antagonism.
...
PMID:cAMP-induced morphological changes are counteracted by the activated RhoA small GTPase and the Rho kinase ROKalpha. 971 82


<< Previous 1 2 3 4 Next >>

---