首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献,搜索用时 31 毫秒
1.
We describe a Triton-insoluble cytoskeletal fraction extracted from cerebral cortex of young rats retaining an endogenous Ca2+-mediated mechanism acting in vitro on Ca2+/calmodulin-dependent protein kinase II (CaM-KII) activity and on phosphorylation and proteolysis of the 150 kDa neurofilament subunit (NF-M), α and β tubulin. Exogenous Ca2+ induced a 70% decrease in the in vitro phosphorylation of the NF-M and tubulins and a 30–50% decrease in the total amount of these proteins. However, when calpastatin was added basal phosphorylation and NF-M and tubulin content were recovered. Furthermore, exogenous Ca2+/calmodulin induced increased in vitro phosphorylation of the cytoskeletal proteins and CaM-KII activity only in the presence of calpastatin, suggesting the presence of Ca2+-induced calpain-mediated proteolysis. This fraction could be an interesting model to further studies concerning the in vitro effects of Ca2+-mediated protein kinases and proteases associated with the cytoskeletal fraction.  相似文献   

2.
Ca2+ plays a major role in neurotransmission and synaptic modulation. Evidence is presented to support the calmodulin hypothesis of neurotransmission developed in this laboratory stating that calmodulin, a major Ca2+ binding protein in brain, mediates the effects of Ca2+ on neurotransmission. Calmodulin was isolated from highly enriched preparations of synaptic vesicles and nerve terminal cytoplasm. Ca2+ and calmodulin were shown to regulate several synaptic processes in isolated and intact preparations, including endogenous synaptic Ca2+-calmodulin protein kinase activity, neurotransmitter release, and synaptic vesicle and synaptic membrane interactions. Ca2+ and calmodulin were shown to activate a synaptic tubulin kinase system which was shown to be a distinct enzyme system from the cyclic AMP protein kinase. Ca2+ and calmodulin stimulated phosphorylation of tubulin altered the properties of tubulin, forming insoluble tubulin fibrils. Evidence for the role of Ca2+-calmodulin kinase activity, especially the calmodulin-tubulin kinase, in neurotransmission are presented. The effects of several neuroactive drugs on the synaptic calmodulin system are presented. The results support the hypothesis that calmodulin mediates many of calcium's actions at the synapse, and that the effects of Ca2+ on synaptic protein phosphorylation, especially synaptic tubulin, may provide a biochemical mechanism for converting the Ca2+ signal into a motor force in the process of neurotransmission.  相似文献   

3.
Purified myelin fraction isolated from rat brain white matter contained Mg2+-dependent protein kinase capable of phosphorylation of myelin basic proteins. The Mg2+-supported kinase was markedly stimulated (two- to fivefold) by micromolar concentrations of free Ca2+ with and without Triton X-100 in the assay, the degree of stimulation being greater with the detergent present. Cyclic AMP, on the other hand, failed to show any effect on phosphorylation of myelin in the absence of Triton X-100 and in the presence of Triton caused only 25–30% stimulation. The phosphorylation reaction was temperature dependent and exhibited a pH optimum at pH 6.5. Apparent affinity toward MgATP2? was found to be about 70 μm and Ca2+ had no effect on this parameter. Dependence on MgCl2 of myelin phosphorylation indicated the presence of high- and low-affinity sites toward Mg2+; Ca2+ appeared to influence the low-affinity site. Maximal level of phosphorylation was attained by 10–15 min at 30 °C and it declined at longer incubation times due to phosphatase activity present in the preparation. Stimulatory effect of Ca2+ on phosphorylation was not due to inhibition of phosphatase activity. Dephosphorylation experiments showed that neither cyclic AMP nor Ca2+ influenced the myelin phosphatase activity. Autoradiographic analysis revealed that phosphorylation of myelin basic proteins accounted for nearly 90% of total myelin phosphorylation. This was supported by the observation that the HCl extract of myelin contained 85% of total activity and comigrated with purified myelin basic proteins. Basal and Ca2+-stimulated phosphorylation of basic proteins were due to phosphorylation of serines mainly, although threonine was phosphorylated to a minor extent. Within myelin, Ca2+ and cyclic AMP kinases are differentially bound. It appears that the myelin kinase (studied in vitro) is primarily influenced by Ca2+ rather than cyclic AMP. Inhibitors (Type I and Type II) of cyclic nucleotide-stimulated protein kinases had no effect on the Ca2+-stimulated phosphorylation although basal and cyclic AMP-stimulated phosphorylation was inhibited, indicating that the Ca2+ kinase is a separate and distinct enzyme from the cyclic AMP-stimulated and basal kinase(s). Also, leupeptin, a protease inhibitor, did not influence basal, cyclic AMP-stimulated, or Ca2+-stimulated myelin phosphorylation, indicating that under the conditions used protease(s) did not alter the myelin kinase activity. The potential significance of phosphorylation of myelin basic proteins and the stimulatory action of Ca2+ on this reaction are discussed.  相似文献   

4.
Calmodulin and the regulation of smooth muscle contraction   总被引:8,自引:0,他引:8  
Calmodulin, the ubiquitous and multifunctional Ca2+-binding protein, mediates many of the regulatory effects of Ca2+, including the contractile state of smooth muscle. The principal function of calmodulin in smooth muscle is to activate crossbridge cycling and the development of force in response to a [Ca2+]i transientvia the activation of myosin light-chain kinase and phosphorylation of myosin. A distinct calmodulin-dependent kinase, Ca2+/calmodulin-dependent protein kinase II, has been implicated in modulation of smooth-muscle contraction. This kinase phosphorylates myosin light-chain kinase, resulting in an increase in the calmodulin concentration required for half-maximal activation of myosin light-chain kinase, and may account for desensitization of the contractile response to Ca2+. In addition, the thin filament-associated proteins, caldesmon and calponin, which inhibit the actin-activated MgATPase activity of smooth-muscle myosin (the cross-bridge cycling rate), appear to be regulated by calmodulin, either by the direct binding of Ca2+/calmodulin or indirectly by phosphorylation catalysed by Ca2+/calmodulin-dependent protein kinase II. Another level at which calmodulin can regulate smooth-muscle contraction involves proteins which control the movement of Ca2+ across the sarcolemmal and sarcoplasmic reticulum membranes and which are regulated by Ca2+/calmodulin, e.g. the sarcolemmal Ca2+ pump and the ryanodine receptor/Ca2+ release channel, and other proteins which indirectly regulate [Ca2+]i via cyclic nucleotide synthesis and breakdown, e.g. NO synthase and cyclic nucleotide phosphodiesterase. The interplay of such regulatory mechanisms provides the flexibility and adaptability required for the normal functioning of smooth-muscle tissues.  相似文献   

5.
The regulation of the phosphorylation of the acetylcholine receptor in electroplax membranes from Torpedo californica and of purified acetylcholine receptor was investigated. The phosphorylation of the membrane-bound acetylcholine receptor was not stimulated by Ca2+/calmodulin, nor was it inhibited by EGTA, but it was stimulated by the catalytic subunit of cAMP-dependent protein kinase, and was blocked by the protein inhibitor of cAMP-dependent protein kinase. Purified acetylcholine receptor was not phosphorylated by Ca2+/calmodulin-dependent protein kinase activity in electroplax membranes, nor by partially purified Ca2+/calmodulin-dependent protein kinases from soluble or particulate fractions from the electroplax. Of the four acetylcholine receptor subunits, termed α, β, γ and δ, only the γ- and δ-subunits were phosphorylated by the cAMP-dependent protein kinase (+cAMP), or by its purified catalytic subunits.  相似文献   

6.
In both cardiac and slow-twitch skeletal muscle sarcoplasmic reticulum (SR) there are several systems involved in the regulation of Ca2+-ATPase function. These include substrate level regulation, covalent modification via phosphorylation-dephosphorylation of phospholamban by both cAMP-dependent protein kinase (PKA) and Ca2+/calmodulin-dependent protein kinase (CaM kinase) as well as direct CaM kinase phosphorylation of the Ca2+-ATPase. Studies comparing, the effects of PKA and CaM kinase on cardiac Ca2+-ATPase function have yielded differing results; similar studies have not been performed in slow-twitch skeletal muscle. It has been suggested recently, however, that phospholamban is not tightly coupled to the Ca2+-ATPase in SR vesicles from slow-twitch skeletal muscle. Our results indicate that assay conditions strongly influence the extent of CaM kinase-dependent Ca2+-ATPase stimulation seen in both cardiac and slow-twitch skeletal muscle. Addition of calmodulin (0.2 M) directly to the Ca2+ transport assay medium results in minimal ( 112–130% of control) stimulation of Ca2+ uptake activity when the Ca2+ uptake reaction is initiated by the addition of either ATP or Ca2+/EGTA. On the other hand, prephosphorylation of the SR by the endogenous CaM kinase and subsequent transfer of the membranes to the Ca2+ transport assay medium results in stimulation of Ca2+ uptake activity (202% of control). These effects are observable in both cardiac and slow-twitch skeletal muscle SR. PKA stimulates Ca2+ uptake markedly (215% of control) when the Ca2+ uptake reaction is initiated by the addition of prephosphorylated SR membranes or by Ca2+/EGTA but minimally (130% of control) when the Ca2+ uptake reaction is initiated by the addition of ATP. These findings imply that (a) phospholamban is coupled to the Ca2+-ATPase in slow-twitch skeletal muscle SR (as in cardiac SR), and (b) the amount of Ca2+ uptake stimulation seen upon the addition of calmodulin or PKA depends strongly on the assay conditions employed. Our observations help to explain the wide range of effects of calmodulin or PKA addition reported in previous studies. It should be noted that, since CaM kinase is now known to phosphorylate the Ca2+-ATPase in addition to phospholamban, further studies are required to determine the relative contributions of phospholambanversus Ca2+-ATPase phosphorylation in the stimulation of Ca2+-ATPase function by CaM kinase. Also, earlier studies attributing all of the effects of CaM kinase stimulation of Ca2+ uptake and Ca2+-ATPase activity to phospholamban phosphorylation need to be re-examined.  相似文献   

7.
This work investigated the role of Ca2+ mobilization and heterotrimeric G protein activation in mediating angiotensin II-dependent tyrosine phosphorylation signaling patterns. We demonstrate that the predominant, angiotensin II-dependent, tyrosine phosphorylation signaling patterns seen in vascular smooth muscle cells are blocked by the intracellular Ca2+ chelator BAPTA-AM, but not by the Ca2+ channel blocker verapamil. Activation of heterotrimeric G proteins with NaF resulted in a divergent signaling effect; NaF treatment was sufficient to increase tyrosine phosphorylation levels of some proteins independent of angiotensin II treatment. In the same cells, NaF alone had no effect on other cellular proteins, but greatly potentiated the ability of angiotensin II to increase the tyrosine phosphorylation levels of these proteins. Two proteins identified in these studies were paxillin and Jak2. We found that NaF treatment alone, independent of angiotensin II stimulation, was sufficient to increase the tyrosine phosphorylation levels of paxillin. Furthermore, the ability of either NaF and/or angiotensin II to increase tyrosine phosphorylation levels of paxillin is critically dependent on intracellular Ca2+. In contrast, angiotensin II-mediated Jak2 tyrosine phosphorylation was independent of intracellular Ca2+ mobilization and extracellular Ca2+ entry. Thus, our data suggest that angiotensin II-dependent tyrosine phosphorylation signaling cascades are mediated through a diverse set of signaling pathways that are partially dependent on Ca2+ mobilization and heterotrimeric G protein activation.  相似文献   

8.
Presynaptic group III metabotropic glutamate receptors (mGluRs) and Ca2+ channels are the main neuronal activity-dependent regulators of synaptic vesicle release, and they use common molecules in their signaling cascades. Among these, calmodulin (CaM) and the related EF-hand Ca2+-binding proteins are of particular importance as sensors of presynaptic Ca2+, and a multiple of them are indeed utilized in the signaling of Ca2+ channels. However, despite its conserved structure, CaM is the only known EF-hand Ca2+-binding protein for signaling by presynaptic group III mGluRs. Because the mGluRs and Ca2+ channels reciprocally regulate each other and functionally converge on the regulation of synaptic vesicle release, the mGluRs would be expected to utilize more EF-hand Ca2+-binding proteins in their signaling. Here I show that calcium-binding protein 1 (CaBP1) bound to presynaptic group III mGluRs competitively with CaM in a Ca2+-dependent manner and that this binding was blocked by protein kinase C (PKC)-mediated phosphorylation of these receptors. As previously shown for CaM, these results indicate the importance of CaBP1 in signal cross talk at presynaptic group III mGluRs, which includes many molecules such as cAMP, Ca2+, PKC, G protein, and Munc18-1. However, because the functional diversity of EF-hand calcium-binding proteins is extraordinary, as exemplified by the regulation of Ca2+ channels, CaBP1 would provide a distinct way by which presynaptic group III mGluRs fine-tune synaptic transmission.  相似文献   

9.
Plasma membranes were prepared from guinea pig ventricle by a procedure which involved differential centrifugation at low gravitational forces, extraction with KCl, and centrifugation in a discontinuous sucrose gradient. Adenylate cyclase was purified 10–15-fold over the starting homogenate with a yield of 75%. The membranes contained an active Ca2+ binding and uptake system as well as Ca2+-activated adenosine triphosphatase; protein kinase and phosphoprotein phosphatase activities were also present. The membranes could be phosphorylated by either intrinsic or exogenous protein kinase, and phosphorylation was stimulated by cyclic AMP and was reversible. Phosphorylated membranes accumulated twice as much Ca2+ as control preparations.  相似文献   

10.
Robert W. Wrenn 《Life sciences》1983,32(20):2385-2392
Phospholipid-sensitive Ca2+-dependent protein kinase and its endogenous substrate proteins were examined in acinar cells from rat pancreas. The enzyme was clearly demonstrable by DEAE-cellulose chromatography of acinar cell extract. At least four endogenous substrate proteins (Mr = 38K, 30K, 22K and 15K) for this Ca2+-activated kinase were found in the acinar cell extract. These substrate proteins were maximally phosphorylated in the combined presence of Ca2+ and phosphatidylserine. Calmodulin was partially effective as a cofactor for phosphorylation of the 38K substrate protein, but ineffective for the other three. A slight Ca2+/phospholipid-dependent phosphorylation of 38K and 30K proteins, but not of 22K and 15K proteins was seen in extract of isolated pancreatic islets. The Ka for Ca2+ for phosphorylation of the endogenous acinar cell proteins was decreased more than ten-fold in the combined presence of phosphatidylserine and unsaturated diacylglycerol. The presence of this Ca2+/phospholipid-dependent protein kinase/ protein phosphorylation system provides a potential mechanism of action for Ca2+ as a regulator of exocrine pancreatic function.  相似文献   

11.
The regulation of the phosphorylation of the acetylcholine receptor in electroplax membranes from Torpedo californica and of purified acetylcholine receptor was investigated. The phosphorylation of the membrane-bound acetylcholine receptor was not stimulated by Ca2+/calmodulin, nor was it inhibited by EGTA, but it was stimulated by the catalytic subunit of cAMP-dependent protein kinase, and was blocked by the protein inhibitor of cAMP-dependent protein kinase. Purified acetylcholine receptor was not phosphorylated by Ca2+/calmodulin-dependent protein kinase activity in electroplax membranes, nor by partially purified Ca2+/calmodulin-dependent protein kinases from soluble or particulate fractions from the electroplax. Of the four acetylcholine receptor subunits, termed α, β, γ and δ, only the γ- and δ-subunits were phosphorylated by the cAMP-dependent protein kinase (+cAMP), or by its purified catalytic subunits.  相似文献   

12.
Summary Studies were performed to determine the effect of protein phosphorylation mediated by calcium-calmodulin-dependent multifunctional protein kinase II and calcium-phospholipid-dependent protein kinase on Na+/H+ exchange activity. Proteins from the apical membrane of the proximal tubule of the rabbit kidney were solubilized in octyl glucoside and incubated in phosphorylating solutions containing the protein kinase.22Na+ uptake was determined subsequently after reconstitution of the proteins into proteoliposomes. Calcium-calmodulin-dependent multifunction protein kinase II inhibited the amiloride-sensitive component of proton gradient-stimulated Na+ uptake in a dose-dependent manner. The inhibitory effect of this kinase had an absolute requirement for calmodulin, Ca2+, and ATP. Calcium-phospholipid-dependent protein kinase stimulated the amiloride-sensitive component of proton gradient-stimulated Na+ uptake in a dose-dependent manner. The stimulating effect of this kinase had an absolute requirement for ATP, Ca2+, and an active phorbol ester. These experiments indicate that Na+/H+ exchange activity of proteoliposomes reconstituted with proteins from renal brush-border membranes are inhibited by protein phosphorylation mediated by calcium-calmodulin-dependent multifunctional protein kinase II and stimulated by that mediated by calcium-calmodulin-dependent protein kinase.  相似文献   

13.
The efficient retrieval of synaptic vesicle membrane and cargo in central nerve terminals is dependent on the efficient recruitment of a series of endocytosis modes by different patterns of neuronal activity. During intense neuronal activity the dominant endocytosis mode is activity-dependent endocytosis (ADBE). Triggering of ADBE is linked to calcineurin-mediated dynamin I dephosphorylation since the same stimulation intensities trigger both. Dynamin I dephosphorylation is maximised by a simultaneous inhibition of its kinase glycogen synthase kinase 3 (GSK3) by the protein kinase Akt, however it is unknown how increased neuronal activity is transduced into Akt activation. To address this question we determined how the activity-dependent increases in intracellular free calcium ([Ca2+]i) control activation of Akt. This was achieved using either trains of high frequency action potentials to evoke localised [Ca2+]i increases at active zones, or a calcium ionophore to raise [Ca2+]i uniformly across the nerve terminal. Through the use of either non-specific calcium channel antagonists or intracellular calcium chelators we found that Akt phosphorylation (and subsequent GSK3 phosphorylation) was dependent on localised [Ca2+]i increases at the active zone. In an attempt to determine mechanism, we antagonised either phosphatidylinositol 3-kinase (PI3K) or calmodulin. Activity-dependent phosphorylation of both Akt and GSK3 was arrested on inhibition of PI3K, but not calmodulin. Thus localised calcium influx in central nerve terminals activates PI3K via an unknown calcium sensor to trigger the activity-dependent phosphorylation of Akt and GSK3.  相似文献   

14.
Ca2+/calmodulin-dependent protein kinase II (CaMKII), the most abundant kinase at the postsynaptic density (PSD), is expected to be involved in activity-induced regulation of synaptic properties. CaMKII is activated when it binds calmodulin in the presence of Ca2+ and, once autophosphorylated on T-286/7, remains active in the absence of Ca2+ (autonomous form). In the present study we used a quantitative mass spectrometric strategy (iTRAQ) to identify sites on PSD components phosphorylated upon CaMKII activation. Phosphorylation in isolated PSDs was monitored under conditions where CaMKII is: (1) mostly inactive (basal state), (2) active in the presence of Ca2+, and (3) active in the absence of Ca2+. The quantification strategy was validated through confirmation of previously described autophosphorylation characteristics of CaMKII. The effectiveness of phosphorylation of major PSD components by the activated CaMKII in the presence and absence of Ca2+ varied. Most notably, autonomous activity in the absence of Ca2+ was more effective in the phosphorylation of three residues on SynGAP. Several PSD scaffold proteins were phosphorylated upon activation of CaMKII. The strategy adopted allowed the identification, for the first time, of CaMKII-regulated sites on SAPAPs and Shanks, including three conserved serine residues near the C-termini of SAPAP1, SAPAP2, and SAPAP3. Involvement of CaMKII in the phosphorylation of PSD scaffold proteins suggests a role in activity-induced structural re-organization of the PSD.  相似文献   

15.
Ca2+/Calmodulin-dependent protein kinase II (CaMKII) has been shown to play a major role in establishing memories through complex molecular interactions including phosphorylation of multiple synaptic targets. However, it is still controversial whether CaMKII itself serves as a molecular memory because of a lack of direct evidence. Here, we show that a single holoenzyme of CaMKII per se serves as an erasable molecular memory switch. We reconstituted Ca2+/Calmodulin-dependent CaMKII autophosphorylation in the presence of protein phosphatase 1 in vitro, and found that CaMKII phosphorylation shows a switch-like response with history dependence (hysteresis) only in the presence of an N-methyl-D-aspartate receptor-derived peptide. This hysteresis is Ca2+ and protein phosphatase 1 concentration-dependent, indicating that the CaMKII memory switch is not simply caused by an N-methyl-D-aspartate receptor-derived peptide lock of CaMKII in an active conformation. Mutation of a phosphorylation site of the peptide shifted the Ca2+ range of hysteresis. These functions may be crucial for induction and maintenance of long-term synaptic plasticity at hippocampal synapses.  相似文献   

16.
Incubation of synaptosomes under conditions which result in complete phosphorylation of membrane bound accepter proteins does not affect the permeability to Na+ or K+ as measured by a spectrophotometric method. This technique was not, however, sensitive enough to determine permeability to Ca2+ which was thus estimated using 45Ca2+. It was found that although phosphorylation did not affect the equilibrium binding of 45Ca it did lower the rate of both Ca2+ uptake and efflux. The most likely interpretation of these results is that phosphorylation of proteins in the synaptic membrane lowers the permeability of the membrane to Ca2+. This could have a role in the regulation of synaptic transmission.  相似文献   

17.
Several plasma-membrane proteins from beet root (Beta vulgaris L.) have been functionally incorporated into reconstituted proteoliposomes. These showed H+-ATPase activity, measured both as ATP hydrolysis and H+ transport. The proton-transport specific activity was 10 times higher than in plasma membranes, and was greatly stimulated by potassium and valinomycin. These proteoliposomes also showed calcium-regulated protein kinase activity. This kinase activity is probably due to a calmodulin-like domain protein kinase (CDPK), since two protein bands were recognized by antibodies against soybean and Arabidopsis CDPK. This kinase phosphorylated histone and syntide-2 in a Ca2+-dependent manner. Among the plasma-membrane proteins phosphorylated by this kinase, was the H+-ATPase. When the H+-ATPase was either prephosphorylated or assayed in the presence of Ca2+, both the ATP-hydrolysis and the proton-transport activities were slower. This inhibition was reversed by an alkaline-phosphatase treatment. A trypsin treatment (that has been reported to remove the C-terminal autoinhibitory domain from the H+-ATPase) also reversed the inhibition caused by phosphorylation. These results indicate that a Ca2+-dependent phosphorylation, probably caused by a CDPK, inhibits the H+-ATPase activities. The substrate of this regulatory phosphorylation could be the H+-ATPase itself, or a different protein influencing the ATPase activities. Received: 1 May 1997 / Accepted: 25 June 1997  相似文献   

18.
A novel Mr 17,000 Ca2+-binding protein isolated from bovine brain was found to be a potent inhibitor of the Ca2+- and phospholipid-dependent protein kinase (protein kinase C), also isolated from bovine brain. Halfmaximal inhibition by this calciprotein of the initial rate of phosphorylation of histone III-S by protein kinase C occurred at a calciprotein concentration of 2.2 μM under standard conditions. Comparison of the effects of a number of Ca2+-binding proteins on protein kinase C activity indicated that the Mr 17,000 Ca2+-binding protein was the most potent inhibitor, followed by the intestinal Ca2+-binding protein and calcineurin. Calmodulin, troponin C, S-100 protein and a Mr 21,000 Ca2+-binding protein of bovine brain were relatively weak inhibitors of protein kinase C. The inhibitory effect of the Mr 17,000 Ca2+-binding protein was apparently not due to its interaction with phospholipid or the basic protein substrate and therefore appears to be due to a direct effect on the protein kinase C. These observations suggest that the novel Mr 17,000 Ca2+-binding protein, and possibly other Ca2+-binding proteins, may play a physiological role in regulating the activity of protein kinase C.  相似文献   

19.
Activation of Ca2+-calmodulin- and cyclic AMP-dependent protein kinases has been suggested to be involved in stimulus-secretion coupling in the pancreatic β-cell. To study the properties of such kinases and their endogenous protein substrates homogenates of rat islets of Langerhans were incubated with [γ-32P]ATP. Phosphorylated proteins were separated by sodium dodecyl sulphate polyacrylamide gel electrophoresis and detected by autoradiography. The phosphorylation of certain proteins could be enhanced by Ca2+ plus calmodulin or by cyclic AMP. The major effect of Ca2+ and calmodulin was to stimulate the phosphorylation of a protein (P53) of molecular weight 53 100±500 (n = 15). Maximum phosphorylation of protein P53 occurred within 2 min with 2 μM free Ca2+ and 0.7 μM calmodulin. Incorporation of label into protein P53 was inhibited by trifluoperazine or W7 but not by cyclic AMP-dependent protein kinase inhibitor. Phosphorylation of a protein of similar molecular weight could be enhanced to a lesser extent in the absence of Ca2+ but in the presence of cyclic AMP and 3-isobutylmethylxanthine: this phosphorylation was blocked by cyclic AMP-dependent protein kinase inhibitor. Cyclic AMP also stimulated incorporation of label into polypeptides of molecular weights 55 000 and 70–80 000. The results are consistent with the hypothesis that protein phosphorylation mechanisms may play a role in the regulation of insulin secretion.  相似文献   

20.
Ca2+/Calmodulin-dependent protein kinase II (CaMKII) has been shown to play a major role in establishing memories through complex molecular interactions including phosphorylation of multiple synaptic targets. However, it is still controversial whether CaMKII itself serves as a molecular memory because of a lack of direct evidence. Here, we show that a single holoenzyme of CaMKII per se serves as an erasable molecular memory switch. We reconstituted Ca2+/Calmodulin-dependent CaMKII autophosphorylation in the presence of protein phosphatase 1 in vitro, and found that CaMKII phosphorylation shows a switch-like response with history dependence (hysteresis) only in the presence of an N-methyl-D-aspartate receptor-derived peptide. This hysteresis is Ca2+ and protein phosphatase 1 concentration-dependent, indicating that the CaMKII memory switch is not simply caused by an N-methyl-D-aspartate receptor-derived peptide lock of CaMKII in an active conformation. Mutation of a phosphorylation site of the peptide shifted the Ca2+ range of hysteresis. These functions may be crucial for induction and maintenance of long-term synaptic plasticity at hippocampal synapses.  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号