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: EC:2.7.11.17 (CaMKII)
4,029 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The mitogenic neuropeptides bombesin and vasopressin markedly increased tyrosine and serine phosphorylation of multiple substrates in quiescent Swiss 3T3 fibroblasts, including two major bands of Mr 90,000 and 115,000. Tyrosine phosphorylation of these proteins was increased as judged by immunoprecipitation of 32Pi-labeled cells and immunoblotting of unlabeled cells with monoclonal antiphosphotyrosine antibodies, elution with phenyl phosphate, and phospho amino acid analysis. Phosphotyrosyl proteins generated by bombesin and vasopressin did not correspond either by apparent molecular weight or by immunological and biochemical criteria to several known tyrosine kinase substrates, including phospholipase C gamma, the microtubule-associated protein 2 kinase, GTPase-activating protein, or phosphatidylinositol kinase. The effect was rapid (within seconds), concentration dependent, and inhibited by specific receptor antagonists for both bombesin and vasopressin. The endothelin-related peptide, vasoactive intestinal contractor, also elicited a rapid and concentration-dependent tyrosine/serine phosphorylation of a similar set of substrates. These results demonstrate that neuropeptides, acting through receptors linked to GTP-binding proteins, stimulate tyrosine phosphorylation of a common set of substrates in quiescent Swiss 3T3 cells and suggest the existence of an additional signal transduction pathway in neuropeptide-induced mitogenesis.
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PMID:Bombesin, vasopressin, and endothelin rapidly stimulate tyrosine phosphorylation in intact Swiss 3T3 cells. 164 10

Recent studies have identified protein tyrosine phosphorylation as a major intracellular signaling pathway. However, little is known about regulation of this signaling pathway in neuronal systems. To help identify changes in levels of protein tyrosine phosphorylation in brain, we have utilized specific anti-phosphotyrosine antibodies to detect phosphotyrosine-containing proteins by immunoblotting techniques. We have found that electroconvulsive treatment induces a selective increase in tyrosine phosphorylation of a soluble 40-kDa protein. The rise is rapid and transient, reaching maximal levels at 1-2 min and returning to basal levels by 8 min. The phosphotyrosine-containing 40-kDa protein is most prominent in hippocampus, smaller in neocortex, and not detected in brainstem or cerebellum. A phosphotyrosine-containing 42-kDa protein present in several cell types has recently been identified as a serine/threonine phosphotransferase, referred to as microtubule-associated protein 2 kinase. Comparison of the levels of tyrosine phosphorylation of the 40-kDa protein and microtubule-associated protein 2 kinase activity during column chromatography of hippocampal extracts demonstrates that the phosphotyrosine-containing 40-kDa protein and microtubule-associated protein 2 co-purify. Moreover, the tyrosine phosphorylation of the 40-kDa protein and microtubule-associated protein 2 kinase activity are increased to a similar extent following electroconvulsive treatment. These findings suggest that the phosphotyrosine-containing 40-kDa protein identified in brain is closely related to microtubule-associated protein 2 kinase.
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PMID:Electroconvulsive treatment induces a rapid and transient increase in tyrosine phosphorylation of a 40-kilodalton protein associated with microtubule-associated protein 2 kinase activity. 170 29

Soluble, monomeric simian virus 40 (SV40) small-t antigen (small-t) was purified from bacteria and assayed for its ability to form complexes with protein phosphatase 2A (PP2A) and to modify its catalytic activity. Different forms of purified PP2A, composed of combinations of regulatory subunits (A and B) with a common catalytic subunit (C), were used. The forms used included free A and C subunits and AC and ABC complexes. Small-t associated with both the free A subunit and the AC form of PP2A, resulting in a shift in mobility during nondenaturing polyacrylamide gel electrophoresis. Small-t did not interact with the free C subunit or the ABC form. These data demonstrate that the primary interaction is between small-t and the A subunit and that the B subunit of PP2A blocks interaction of small-t with the AC form. The effect of small-t on phosphatase activity was determined by using several exogenous substrates, including myosin light chains phosphorylated by myosin light-chain kinase, myelin basic protein phosphorylated by microtubule-associated protein 2 kinase/ERK1, and histone H1 phosphorylated by protein kinase C. With the exception of histone H1, small-t inhibited the dephosphorylation of these substrates by the AC complex. With histone H1, a small stimulation of dephosphorylation by AC was observed. Small-t had no effect on the activities of free C or the ABC complex. A maximum of 50 to 75% inhibition was obtained, with half-maximal inhibition occurring at 10 to 20 nM small-t. The specific activity of the small-t/AC complex was similar to that of the ABC form of PP2A with myosin light chains or histone H1 as the substrate. These results suggested that small-t and the B subunit have similar qualitative and quantitative effects on PP2A enzyme activity. These data show that SV40 small-antigen binds to purified PP2A in vitro, through interaction with the A subunit, and that this interaction inhibits enzyme activity.
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PMID:Control of protein phosphatase 2A by simian virus 40 small-t antigen. 170 74

The N-methyl-D-aspartate (NMDA) receptor, a subtype of glutamate receptors, plays a key role in synaptic plasticity in the nervous system. After NMDA receptor activation, calcium entry into the postsynaptic neuron is a critical initial event. However, the subsequent mechanisms by which the NMDA receptor signal is processed are incompletely understood. Stimulation of cultured rat hippocampal cells with glutamate resulted in the rapid and transient tyrosine phosphorylation of a 39-kilodalton protein (p39). Tyrosine phosphorylation of p39 was triggered by the NMDA receptor and required an influx of Ca2+ from the extracellular medium. Because p39 was found to be highly related or identical to the microtubule-associated protein 2 kinase, the NMDA receptor signal may be processed by a sequential activation of protein kinases.
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PMID:Stimulation of protein tyrosine phosphorylation by NMDA receptor activation. 171 95

Protein phosphorylation is considered an early cellular mechanism of signal transduction by surface immunoglobulins (sIg) and other receptors of B cells. Using intact human peripheral blood B cells of young subjects labeled with orthophosphate, increased phosphorylation levels of serine/threonine and tyrosine substrates were demonstrated on indicator phosphoproteins corresponding to the CD20 isoforms and microtubule-associated protein 2 kinase after cross-linking sIg and costimulation with phorbol diesters. By contrast, stimulated B cells from certain elderly subjects displayed substantial alterations in the phosphorylation patterns of serine/threonine or tyrosine indicator phosphoproteins. Also, age-related impairments in sIg stimulated mobilization of cytosolic protein kinase C (PKC) enzymatic activity and in cytosolic calcium [Ca2+]i responses of B cells were observed with the altered phosphorylation reactions. Comparison of the substrate phosphorylation profiles to the proliferative responses of stimulated B cells from individual elderly subjects suggested a model of signal transduction in which differing stimuli have different dependencies on phosphorylation reactions. Diminished proliferative responses after sIg ligation coincided with decreased phosphorylations of either tyrosine or serine/threonine indicator substrates. However, the decreased proliferative responses of B cells from elderly subjects with substantial reductions of tyrosine phosphorylation after sIg ligation were enhanced by the direct stimulation of serine/threonine kinase activity with phorbol diesters or CD40 ligation. Experiments with kinase inhibitors evaluated the relative dependency of different B cell stimuli on tyrosine and serine/threonine phosphorylation reactions. The proliferative responses of normal B cells to sIg ligation were quite sensitive to the tyrosine kinase inhibitor genistein whereas those observed following costimulations with phorbol diesters or CD40 ligation were more resistant. However, treatment of B cells with H7, an inhibitor of PKC activity, led to a more uniform reduction of B-cell responses after different stimuli. Results from RNase protection assays of c-myc expression also suggested that different B-cell stimuli might utilize distinct intracellular signaling pathways. Both the type of stimuli and mode of sIg ligation were important in determining the stimulated levels of c-myc mRNA expression. Thus, the current findings suggest that age-related defects are present in human B cell signaling pathways as reflected by tyrosine and serine/threonine phosphorylation reactions. Also, these age-related defects can coexist with altered mobilization of PKC enzymatic activity and with alterations in [Ca2+]i and proliferative responses.
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PMID:Signal transduction in human B cells during aging: alterations in stimulus-induced phosphorylations of tyrosine and serine/threonine substrates and in cytosolic calcium responsiveness. 180 9

In Swiss 3T3 fibroblasts a peptide mitogen bombesin, which acts through the phospholipase C-protein kinase C signaling pathway, stimulates DNA synthesis in a manner strictly dependent on the medium calcium concentration: [3H]thymidine incorporation into DNA in the presence of a saturating concentration of bombesin (10(-8) M) is 4-fold greater at 3.0 mM extracellular calcium as compared with a value obtained at 0.03 mM calcium. In the present study we attempted to identify the site and the mechanism of action of Ca2+ influx along the bombesin-induced mitogenic signaling pathway, by comparing bombesin effects at 0.03 and 3.0 mM of medium calcium. Bombesin induces the same extent of increases in [3H]inositol phosphates after 1 min, and comparable sustained increases in the cellular content of 1,2-diacylglycerol for up to 4 h, at either 0.03 or 3.0 mM calcium. Bombesin induces the same extent of phosphorylation of MARCKS protein, the major cellular substrate for protein kinase C, irrespective of the medium calcium concentration for at least 4 h. Moreover, diverse cellular responses elicited by bombesin, including c-fos expression, activation of microtubule-associated protein 2 kinase and S6 kinase, glucose uptake, and protein synthesis but not the release of arachidonic acid and its metabolites, are induced similarly at either 0.03 or 3.0 mM calcium. Down-regulation of cellular protein kinase C nearly completely abolishes bombesin effects on c-fos expression, S6 kinase activation, glucose uptake, and DNA synthesis. These results suggest that the target of Ca2+ influx in bombesin-induced mitogenic signaling pathway is not located along the phospholipase C-protein kinase C signal transduction system including cellular events in early G1 phase that exist downstream to protein kinase C action.
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PMID:Role of Ca2+ influx in bombesin-induced mitogenesis in Swiss 3T3 fibroblasts. 184 53

The dominant insulin-stimulated ribosomal protein S6 kinase activity was purified to near homogeneity from insulin-treated 32P-labeled rat H4 hepatoma cells and found to copurify with a 70-kDa 32P-labeled polypeptide. The dominant S6 kinase purified from livers of cycloheximide-treated rats is also a 70-kDa polypeptide. Antiserum raised against rat liver S6 kinase specifically immunoprecipitates the purified 32P-labeled H4 hepatoma insulin-stimulated S6 kinase. This antiserum also specifically precipitates insulin-stimulated S6 kinase activity directly from cytosolic extracts of H4 cells. Immune complexes prepared from the cytosol of 32P-labeled H4 cells contain several 32P-labeled polypeptides; only a 70-kDA 32P-labeled peptide, however, is specifically displaced by preadsorption of the antiserum with nonradioactive rat liver S6 kinase. Insulin treatment increases the 32P content of the immunoprecipitated 70-kDa S6 kinase polypeptide 3- to 4-fold over basal levels; 32P-labeled serine, some 32P-labeled threonine, but no 32P-labeled tyrosine are detected after partial acid hydrolysis. Tryptic peptide maps indicate that the insulin-stimulated S6 kinase purified from 32P-labeled H4 cells is phosphorylated at multiple sites distinct from those which participate in autophosphorylation in vitro. Autophosphorylation of rat liver S6 kinase in vitro does not modify S6 kinase activity. The S6 kinases purified from liver of cycloheximide-treated rat and H4 hepatoma insulin-stimulated enzyme are each completely deactivated by incubation with protein phosphatase type 2A in both autophosphorylating and 40S S6 phosphorylating activities. The phosphatase 2A-deactivated 70-kDa S6 kinase is neither reactivated nor phosphorylated by partially purified insulin-stimulated microtubule-associated protein 2 kinase, in experiments where Xenopus S6 kinase II undergoes phosphorylation and partial reactivation. Thus insulin activates the 70-kDa S6 kinase by promoting phosphorylation of specific serine/threonine residues on the enzyme polypeptide, probably through activating an as-yet-unidentified serine/threonine protein kinase distinct from microtubule-associated protein 2 kinase.
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PMID:Insulin activates a 70-kDa S6 kinase through serine/threonine-specific phosphorylation of the enzyme polypeptide. 212 50

Growth factor activation of serine/threonine protein kinases was studied by treating quiescent Swiss 3T3 cells with epidermal growth factor (EGF) and examining cytosolic extracts for protein kinase activity under conditions inhibitory to calcium- and cyclic nucleotide-dependent kinases. Cytosolic extracts of cells stimulated for 5 min were fractionated by Mono Q fast protein liquid chromatography. Eight peaks of kinase activity were resolved, of which five were stimulated by EGF treatment of cells. These peaks were revealed using the synthetic peptide Arg-Arg-Leu-Ser-Ser-Leu-Arg-Ala (S6 peptide), 40 S ribosomal S6 protein, glycogen synthase, microtubule-associated protein 2, and myelin basic protein as substrates. The peaks varied in the kinetics of their activation by EGF and in their response to insulin. Selected peaks were resolved further by sizing gel chromatography. The results together indicate that at least seven distinct fractions of cytosolic kinase activities are stimulated in Swiss 3T3 cells by EGF. One of these, which phosphorylates both S6 protein and S6 peptide, is similar to the S6 kinase characterized previously in this cell line by others. Four additional activities that also phosphorylate the S6 protein and S6 peptide appear unrelated to this enzyme. Finally, two kinase activities that phosphorylate both myelin basic protein and microtubule associated protein 2 are EGF stimulated. One is similar to an insulin-stimulated microtubule-associated protein 2 kinase described in other cell lines whereas the other seems to represent a novel activity. Several of these EGF-stimulated activities were inactivated by protein phosphatases, suggesting that they might be regulated by phosphorylation.
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PMID:Identification of multiple epidermal growth factor-stimulated protein serine/threonine kinases from Swiss 3T3 cells. 214 53

The CD4R has been shown to exert variable effects on T cell activation responses. Depending on the manner of ligation, the CD4R has been demonstrated to have positive as well as negative effects on the generation of [Ca2+]i flux by the CD3R. Coaggregation of CD3 with CD4 enhanced Ca2+ flux while their independent ligation and aggregation diminished this response. To further elucidate these paradoxical CD4 effects, we studied induction of a microtubule-associated protein 2 kinase (MAP-2K) activity during ligation of the CD3R. Lymphoid MAP-2K activation by CD3 is an evanescent event that is dependent on phosphorylation of 43-kDa MAP-2K via a pathway that involves protein kinase C. Coaggregation of CD4 and CD3 with cross-linking antibodies and avidin enhanced the CD3-mediated MAP-2K response almost twofold. In contrast, independent ligation and cross-linking of CD4 reduced the CD3-induced MAP-2K response by approximately 50%. An important requirement for this inhibitory effect was that CD4 be ligated before stimulation with anti-CD3. The negative effect of anti-CD4 mAb was specific as other mAb failed to simulate this event. The PMA-induced MAP-2K response was not inhibited by anti-CD4. Intact 32P-labeled Jurkat and normal human T cells demonstrated the appearance of a single 43-kDa tyrosine phosphoprotein during stimulation with PMA and anti-CD3. When these crude cellular extracts were extensively fractionated across DEAE- and hydrophobic columns, MAP-2K was resolved into two peaks of activity, each containing a single tyrosine phosphoprotein around 43 kDa. In addition to tyrosine-specific labeling, mitogenic stimulation of normal human T cells also induced threonine-specific labeling of MAP-2K. These results imply that activation of lymphoid MAP-2K is a dual process requiring at least two independent kinases for optimal activity. Inasmuch as CD3 activates protein kinase C and CD4 is associated with a tyrosine kinase, pp56lck, we suggest that their coaggregation may create the conditions whereby MAP-2K may be activated by dual phosphorylation. Independent aggregation of these receptors may lead to physical separation and breakdown of this interactive mechanism.
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PMID:CD-3-mediated activation of MAP-2 kinase can be modified by ligation of the CD4 receptor. Evidence for tyrosine phosphorylation during activation of this kinase. 216 97

2,3,7,8-Tetrachloro-p-dioxin (TCDD) induced a modest stimulation of nuclear protein phosphorylation in explant tissue cultures in 10 min, followed by a substantial decrease in the level of total protein phosphorylation activity in the nucleus. Curiously, this TCDD-induced decline in nuclear protein phosphorylation was accompanied by an increase in cytosolic and extranuclear protein phosphorylation activity. One of the main causes for such a decrease in the protein phosphorylation activity in the nucleus appears to be related to some increase in protein phosphatase activities as judged by the counteractions of okadaic acid and Na3VO4 to the above effect. In addition, TCDD induced changes in nuclear protein kinase activities as well. Manganese-stimulated protein kinase was found to be the predominant type of nuclear protein phosphorylating activity affected by TCDD, with 60% of the total activity due to heparin-sensitive casein kinase II (CK II), a major nuclear protein kinase. The level of CK II activity in the nuclear protein preparation from adipose tissue of TCDD-treated guinea pigs (1 microgram/kg) in the presence of 100 nM heparin was only 35% of the control value after 24 hr. In addition, TCDD was found to increase the protein kinase C and microtubule-associated protein 2 kinase activities as early as 15 min after treatment in isolated adipose tissues in culture. Under in situ incubation conditions with explant tissues in culture, TCDD rapidly enhanced the DNA binding activity of the transcriptional factor AP-1, whereas the same treatment reduced c-Myc DNA binding activity. Genistein, a specific protein tyrosine kinase inhibitor, abolished the stimulatory effect of TCDD on AP-1 binding activity, but not on DNA binding activity of c-Myc. Phorbol ester (TPA) increased the binding activity of AP-1 and c-Myc, as expected. However, TCDD in combination with TPA caused a slight reduction in binding activity of both transcriptional factors. On the other hand, in the presence of forskolin, the stimulatory effect of TCDD on AP-1 binding activity and the inhibitory effect on c-Myc were still apparent. Okadaic acid almost abolished the binding activity of c-Myc, whereas in combination with TCDD a stimulatory effect was found. These observations are consistent with the idea that TCDD regulates the DNA binding activity of AP-1 and c-Myc mainly through modulating their states of phosphorylation by altering protein kinase and phosphatase activities.
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PMID:Regulation by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) of the DNA binding activity of transcriptional factors via nuclear protein phosphorylation in guinea pig adipose tissue. 748 34


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