Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.17 (CaMKII)
 4,029 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Chinese hamster ovary cells exhibit several characteristic morphological and physiological responses upon treatment with agents which increase the intracellular level of adenosine 3':5'-phosphate (cyclic AMP). To better understand the mechanism of these cyclic AMP-mediated responses, we separated two cyclic AMP-dependent protein kinases (ATP:protein phosphotransferase, EC 2.7.1.37) (protein kinase I and protein kinase II) from the cytosol of Chinese hamster ovary cells by DEAE-cellulose chromatography and studied their properties. Protein kinase I is eluted at a lower salt concentration than protein kinase II and is stimulable to 10 times its basal catalytic activity, while protein kinase II is stimulable only 2-fold. Both kinases are completely dissociated by cyclic AMP and inhibited by specific cyclic AMP-dependent protein kinase inhibitor. They have similar Km values for magnesium (approximately 1 mM), cyclic AMP (approximately 60 nM), and ATP (approximately 0.1 mM), and the dissociation constant (Kdis) for cyclic AMP (approximately 13 nM) is the same for both enzymes. However, they appear to have different substrate preferences and cyclic AMP-binding properties in that cyclic AMP bound to protein kinase II exchanges readily with free cyclic AMP, while that bound to protein kinase I is not exchangeable. The native enzymes have different sedimentation coefficients (6.4 S for protein kinase I and 4.8 S for protein kinase II), whereas those of the activated enzymes are the same (2.9--3.0 S). It appears that the two cyclic AMP-dependent protein kinases which differ from each other in their regulatory subunits may play different roles in the mediation of cyclic AMP action in Chinese hamster ovary cells.
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PMID:Characterization of two adenosine 3':5'-phosphate-dependent protein kinase species from Chinese hamster ovary cells. 21 11

A regulatory region involved in both autoinhibition and calmodulin (CaM) binding has previously been identified in the multifunctional Ca2+/CaM-dependent protein kinase (CaM kinase II). We have tested the role of various segments of the regulatory region in autoinhibition by the analysis of a series of truncation, substitution, and deletion mutants of the CaM kinase II alpha subunit (CaM kinase II alpha). Unexpectedly, the sequence Lys-Lys-Phe-Asn at positions 291-294, adjacent to the CaM binding domain, was found to be sufficient to maintain an inhibited state in a truncated form of the kinase. However, these residues are not essential in the context of the full-length protein, indicating the importance of additional residues from the overlapping CaM binding domain. We propose here a molecular model for CaM kinase II alpha based on the three-dimensional structure of the cAPK-PKI-(5-24) (protein kinase inhibitor fragment) complex. It is predicted from this model that autoinhibition is of the pseudosubstrate variety and that autophosphorylation of Thr-286 could occur by an intersubunit reaction in the holoenzyme complex.
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PMID:Regulation of intrasteric inhibition of the multifunctional calcium/calmodulin-dependent protein kinase. 133 58

We determined the effect of nucleotides and protein kinase A (PKA) on the Ca(2+)-dependent gating of the cloned intermediate conductance, Ca(2+)-dependent K(+) channel, hIK1. In Xenopus oocytes, during two-electrode voltage-clamp, forskolin plus isobutylmethylxanthine induced a Ca(2+)-dependent increase in hIK1 activity. In excised inside-out patches, addition of ATP induced a Ca(2+)-dependent increase in hIK1 activity (NP(o)). In contrast, neither nonhydrolyzable (AMP-PNP, AMP-PCP) nor hydrolyzable ATP analogs (GTP, CTP, UTP, and ITP) activated hIK1. The ATP-dependent activation of hIK1 required Mg(2+) and was reversed by either exogenous alkaline phosphatase or the PKA inhibitor PKI(5-24). The Ca(2+) dependence of hIK1 activation was best fit with a stimulatory constant (K(s)) of 350 nM and a Hill coefficient (n) of 2.3. ATP increased NP(o) at [Ca(2+)] >100 nM while having no effect on K(s) or n. Mutation of the single PKA consensus phosphorylation site at serine 334 to alanine (S334A) had no effect on the PKA-dependent activation during either two-electrode voltage-clamp or in excised inside-out patches. When expressed in HEK293 cells, ATP activated hIK1 in a Mg(2+)-dependent fashion, being reversed by alkaline phosphatase. Neither PKI(5-24) nor CaMKII(281-309) or PKC(19-31) affected the ATP-dependent activation. Northern blot analysis revealed hIK1 expression in the T84 colonic cell line. Endogenous hIK1 was activated by ATP in a Mg(2+)- and PKI(5-24)-dependent fashion and was reversed by alkaline phosphatase, whereas CaMKII(281-309) and PKC(19-31) had no effect on the ATP-dependent activation. The Ca(2+)-dependent activation (K(s) and n) was unaffected by ATP. In conclusion, hIK1 is activated by a membrane delimited PKA when endogenously expressed. Although the oocyte expression system recapitulates this regulation, expression in HEK293 cells does not. The effect of PKA on hIK1 gating is Ca(2+)-dependent and occurs via an increase in NP(o) without an effect on either Ca(2+) affinity or apparent cooperativity.
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PMID:Kinase-dependent regulation of the intermediate conductance, calcium-dependent potassium channel, hIK1. 1061 55

Modulation of postsynaptic AMPA receptors in the brain by phosphorylation may play a role in the expression of synaptic plasticity at central excitatory synapses. It is known from biochemical studies that GluR1 AMPA receptor subunits can be phosphorylated within their C terminal by cAMP-dependent protein kinase A (PKA), which is colocalized with the phosphatase calcineurin (i.e., phosphatase 2B). We have examined the effect of PKA and calcineurin on the time course, peak open probability (P(O, PEAK)), and single-channel properties of glutamateevoked responses for neuronal AMPA receptors and homomeric GluR1(flip) receptors recorded in outside-out patches. Inclusion of purified catalytic subunit Calpha-PKA in the pipette solution increased neuronal AMPA receptor P(O,PEAK) (0.92) compared with recordings made with calcineurin included in the pipette (P(O,PEAK) 0.39). Similarly, Calpha-PKA increased P(O,PEAK) for recombinant GluR1 receptors (0. 78) compared with patches excised from cells cotransfected with a cDNA encoding the PKA peptide inhibitor PKI (P(O,PEAK) 0.50) or patches with calcineurin included in the pipette (P(O,PEAK) 0.42). Neither PKA nor calcineurin altered the amplitude of single-channel subconductance levels, weighted mean unitary current, mean channel open period, burst length, or macroscopic response waveform for recombinant GluR1 receptors. Substitution of an amino acid at the PKA phosphorylation site (S845A) on GluR1 eliminated the PKA-induced increase in P(O,PEAK), whereas the mutation of a Ca(2+), calmodulin-dependent kinase II and PKC phosphorylation site (S831A) was without effect. These results suggest that AMPA receptor peak response open probability can be increased by PKA through phosphorylation of GluR1 Ser845.
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PMID:Control of GluR1 AMPA receptor function by cAMP-dependent protein kinase. 1062 85

Whole-cell patch clamp experiments were used to investigate the transduction mechanism of adenosine A(2A) receptors in modulating N-methyl-D-aspartate (NMDA)-induced currents in rat striatal brain slices. The A(2A) receptor agonist 2-p-(2-carboxyethyl)phenethylamino-5'-N-ethylcarboxamidoadenosine (CGS 21680) inhibited the NMDA, but not the (S)-alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) current in a subset of striatal neurons. Lucifer yellow-filled pipettes in combination with immunostaining of A(2A) receptors were used to identify CGS 21680-sensitive cells as typical medium spiny striatal neurons. Dibutyryl cyclic AMP and the protein kinase A activator Sp-cyclic AMPs, but not the protein kinase A inhibitors Rp-cyclic AMPS or PKI(14 - 24)amide abolished the inhibitory effect of CGS 21680. The phospholipase C inhibitor U-73122, but not the inactive structural analogue U-73343 also interfered with CGS 21680. The activation of protein kinase C by phorbol 12-myristate 13-acetate or the blockade of this enzyme by staurosporine did not alter the effect of CGS 21680. Heparin, an antagonist of inositol 1, 4,5-trisphosphate (InsP(3)) and a more efficient buffering of intracellular Ca(2+) by BAPTA instead of EGTA in the pipette solution, abolished the CGS 21680-induced inhibition. The calmodulin antagonist W-7 and cytochalasin B which enhances actin depolymerization also prevented the effect of CGS 21680; the calmodulin kinase II inhibitors CaM kinase II(281 - 309) and KN-93 but not the inactive structural analogue KN-92 were also effective. The calcineurin inhibitor deltamethrin did not interfere with CGS 21680. It is suggested that the transduction mechanism of A(2A) receptors to inhibit NMDA receptor channels is the phospholipase C/InsP(3)/calmodulin and calmodulin kinase II pathway. The adenylate cyclase/protein kinase A and phospholipase C/protein kinase C pathways do not appear to be involved.
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PMID:Inhibition by adenosine A(2A) receptors of NMDA but not AMPA currents in rat neostriatal neurons. 1080 62

The nuclear envelope mediates key functions by interacting with chromatin. We recently reported an interaction between the chromatin- and nuclear matrix-associated protein HA95 and the inner nuclear membrane integral protein LAP2beta, implicated in initiation of DNA replication (Martins et al. (2003) J. Cell Biol. 160, 177-188). Here, we show that in vitro, interaction between HA95 and LAP2beta is modulated by cAMP signaling via PKA. Exposure of an anti-HA95 immune precipitate from interphase HeLa cells to a mitotic extract promotes ATP-dependent release of LAP2beta from the HA95 complex. This coincides with Ser and Thr phosphorylation of HA95 and LAP2beta. Inhibition of PKA with PKI abolishes phosphorylation of HA95 and dissociation of LAP2beta from HA95, although LAPbeta remains phosphorylated. Antagonizing cAMP signaling in mitotic extract also abolishes the release of LAP2beta from HA95; however, disrupting PKA anchoring to A-kinase anchoring proteins has no effect. Inhibition of CDK activity in the extract greatly reduces LAP2beta phosphorylation but does not prevent LAP2beta release from HA95. Inhibition of PKC, MAP kinase, or CaM kinase II does not affect mitotic extract-induced dissociation of LAP2beta from HA95. PKA phosphorylates HA95 but not LAP2beta in vitro and elicits a release of LAP2beta from HA95. CDK1 or PKC phosphorylates LAP2beta within the HA95 complex, but neither kinase induces LAP2beta release. Our results indicate that in vitro, the interaction between HA95 and LAP2beta is influenced by a PKA-mediated phosphorylation of HA95 rather than by CDK1- or PKC-mediated phosphorylation of LAP2beta. This suggests an additional level of regulation of a chromatin-nuclear envelope interaction in dividing cells.
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PMID:In vitro modulation of the interaction between HA95 and LAP2beta by cAMP signaling. 1295 Jan 72

Histamine H(3) autoreceptors induce a negative feedback on histamine synthesis and release. While it is known that cAMP/cAMP dependent protein kinase (PKA) and Ca(2+)/CaMKII transduction pathways mediate H(3) effects on histamine synthesis, the pathways regulating neuronal histamine release are poorly known. Given the potential use of H(3) ligands in cognitive diseases, we have developed a technique for the determination of H(3) effects on histamine synthesis and release in brain cortical miniprisms. Potassium-induced depolarization effects were impaired by blockade of calcium entry through N and P/Q channels, as well as of CaMKII, but release was not affected by activators or inhibitors of the cAMP/PKA pathway (1-methyl-3-isobutylxanthine (IBMX), N6,2'-O-dibutyryladenosine 3',5'-cyclic monophosphate sodium salt (db-cAMP) or myristoyl PKA inhibitor peptide 14-22 (PKI(14-22)). In contrast, forskolin stimulated histamine release, although independently of PKA. Stimulation of histamine H(3) receptors with the agonist imetit markedly reduced the depolarization increase of histamine release, apparently through P/Q calcium channel inhibition. The H(3) antagonist/inverse agonist thioperamide modestly stimulated histamine release. Thioperamide effect on release was not modified by the PKA inhibitor PKI(14-22), but it was blocked by the CaMKII inhibitor KN-62. These results indicate that H(3) autoreceptors regulate neuronal histamine release (1) independently of the cAMP/PKA cascade, and (2) through modulation of calcium entry and CaMKII activation during depolarization.
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PMID:Different role of cAMP dependent protein kinase and CaMKII in H3 receptor regulation of histamine synthesis and release. 1973