Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.13 (protein kinase C)
 49,245 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Dynamin I is a nerve terminal phosphoprotein with intrinsic guanosine triphosphatase (GTPase) activity that is required for endocytosis. Upon depolarization and synaptic vesicle recycling, dynamin I undergoes a rapid dephosphorylation. Dynamin I was found to be a specific high-affinity substrate for calcineurin in vitro. At low concentrations, calcineurin dephosphorylated dynamin I that had been phosphorylated by protein kinase C. The dephosphorylation inhibited dynamin I GTPase activity in vitro and after depolarization of nerve terminals. The effect in nerve terminals was prevented by the calcineurin inhibitor cyclosporin A. This suggests that in nerve terminals, calcineurin serves as a Ca(2+)-sensitive switch for depolarization-evoked synaptic vesicle recycling.
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PMID:Calcineurin inhibition of dynamin I GTPase activity coupled to nerve terminal depolarization. 805 58

Depolarization-induced Ca2+ influx into rat brain synaptosomes induces dephosphorylation of dephosphin, a 96-94-kDa protein kinase C (PKC) substrate recently identified as dynamin I, a protein associated with endocytosis. We characterized purified dynamin I to better understand regulation of its phosphorylation in nerve terminals. Purified dynamin I possessed a very high affinity for PKC but did not fit Michaelis-Menten kinetics. It had an optimum phosphorylation rate of 1.42 +/- 0.02 mumol/mg/min and a concentration giving half-maximal activity (S0.5) of 0.14 +/- 0.02 microM, the highest affinity reported for a PKC substrate protein. Concentrations of dynamin greater than 0.5 microM inhibited phosphorylation. The stoichiometry was 1.5, indicating more than one phosphorylation site. Dynamin was predominantly associated with the brain particulate fraction under conditions of low ionic strength, and this prevented its phosphorylation by PKC until released by moderate increases in ionic strength (Na+, K+, and Mg2+) or by GTP or ATP. In intact synaptosomes the largest dynamin pool was associated with the particulate fraction, while a smaller pool was cytosolic or extracted with 150 nM NaCl and contained all the phosphorylated protein. Purified dynamin also bound to phospholipid-coated controlled-pore glass beads, but poorly in the presence of NaCl, Mg2+, GTP, or ATP. Ca2+ induced a reversible translocation from the cytosol to the particulate fraction (50% at 183 microM Ca2+) in brain homogenates, and the purified protein also underwent Ca(2+)-sensitive translocation to phospholipid-coated controlled-pore glass beads. We conclude that dynamin I is a nerve terminal Ca(2+)-sensitive phospholipid-binding protein with very high substrate affinity for PKC. We propose that phosphorylation by PKC occurs in the nerve terminal soluble compartment and that Ca2+ may mediate its binding to the particulate fraction, thereby blocking the PKC phosphorylation sites. These properties may contribute to the lack of PKC phosphorylation during depolarization, despite the presence of activated PKC.
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PMID:Dynamin I is a Ca(2+)-sensitive phospholipid-binding protein with very high affinity for protein kinase C. 806 23

Dynamin is a GTP-, microtubule-, and phospholipid-binding protein that is expressed primarily in brain. In Drosophila, the shibire gene encodes a homologue of dynamin; mutations in this gene result in a defect in endocytosis, suggesting a function for dynamin in endocytic membrane traffic. In the present study we show that there are at least two distinct dynamin genes in mammals whose products are referred to as dynamins I and II. The two dynamins are similar to each other (79% identity) and are both equally homologous to the Drosophila shibire gene product (66% identity). The highest degree of identity between dynamins is observed in their N-terminal halves, whereas their C termini exhibit little homology. Transcripts of both dynamin genes are subject to at least two alternative splicing events, the first of which is identically found in both dynamins, whereas the second site of alternative splicing is different between the two types of dynamins. The first alternatively spliced sequence of the dynamins consists of an interior region that is present in two distinct but homologous forms in both dynamins, suggesting alternative use of exons in both genes at identical positions. The second site of alternative splicing results in the generation of different C termini in dynamin I and in the inclusion or exclusion of an interior four-amino acid sequence in dynamin II. The two dynamins exhibit remarkable differences in their tissue distribution and regulation. Dynamin I is almost exclusively expressed in the central nervous system. Conversely, dynamin II is expressed ubiquitously in all tissues tested. Previous studies revealed that the GTPase activity of dynamin I is regulated by phosphorylation by protein kinase C in nerve terminals. Expression of dynamins I and II by transfection in COS cells demonstrates that only dynamin I but not dynamin II is a substrate for protein kinase C. Our data suggest a specialization in the endocytic functions and the regulation of dynamins between neural and non-neural tissues in mammals.
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PMID:Differential expression and regulation of multiple dynamins. 830 25

Dynamin is a microtubule-binding protein with a microtubule-activated GTPase activity. The gene encoding dynamin is mutated in shibire, a Drosophila mutant defective in endocytosis in nerve terminals and other cells. These observations place dynamin into two distinct functional contexts, suggesting roles in microtubule-based motility or in endocytosis. We report here that dynamin is identical to the neuronal phosphoprotein dephosphin (P96), originally identified by its stimulus-dependent dephosphorylation in nerve terminals. Dynamin is a protein doublet of M(r) 94 and 96K arising by alternative splicing of its primary transcript. In the nerve terminal, both forms of dynamin are phosphorylated by protein kinase C (PKC) and are quantitatively dephosphorylated on excitation. In vitro, dynamin is also phosphorylated by casein kinase II which inhibits PKC phosphorylation. Phosphorylation by PKC but not by casein kinase II enhances the GTPase activity of dynamin 12-fold. The dynamins are therefore a group of nerve terminal phosphoproteins whose GTPase is regulated by phosphorylation in parallel with synaptic vesicle recycling. The regulation of dynamin GTPase could serve as the trigger for the rapid endocytosis of synaptic vesicles after exocytosis.
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PMID:Dynamin GTPase regulated by protein kinase C phosphorylation in nerve terminals. 837 52

Synaptic vesicle recycling is a neuronal specialization of endocytosis that requires the GTPase activity of dynamin I and is triggered by membrane depolarization and Ca2+ entry. To establish the relationship between dynamin I GTPase activity and Ca2+, we used purified dynamin I and analyzed its interaction with Ca2+ in vitro. We report that Ca2+ bound to dynamin I and this was abolished by deletion of dynamin's C-terminal tail. Phosphorylation of dynamin I by protein kinase C promoted formation of a dynamin I tetramer and increased Ca2+ binding to the protein. Moreover, Ca2+ inhibited dynamin I GTPase activity after stimulation by phosphorylation or by phospholipids but not after stimulation with a GST-SH3 fusion protein containing the SH3 domain of phosphoinositide 3-kinase. These results suggest that in resting nerve terminals, phosphorylation of dynamin I by protein kinase C converts it to a tetramer that functions as a Ca(2+)-sensing protein. By binding to Ca2+, dynamin I GTPase activity is specifically decreased, possibly to regulate synaptic vesicle recycling.
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PMID:Calcium binds dynamin I and inhibits its GTPase activity. 878 38

Dynamin is a neuronal phosphoprotein and a GTPase enzyme which mediates late stages of endocytosis in both neural and non-neural cells. Current knowledge about dynamin is reviewed with particular emphasis on its structure and regulation with respect to phosphorylation, protein-protein interactions and phospholipid binding. The major themes are the biochemical regulation of dynamin, its effects on dynamin's GTPase activity and how this might relate to assembling the 'fission ring' that brings about vesicle retrieval. Dynamin I is an isoform of the enzyme primarily located in the central and peripheral nervous systems, where it is enriched in areas of abundant synaptic contacts. Dynamin I undergoes protein-protein interactions via its proline-rich domain at the C-terminus and these can elevate its N-terminal GTPase activity. Dynamin I interacts with multiple proteins in the nerve terminal, including SH3 domain-containing proteins such as amphiphysin and potentially with other proteins such as betagamma subunits. These regulate its role in endocytosis by targeting dynamin I to specific subcellular locations of retrieval. Dynamin I is phosphorylated in vivo by PKC and dephosphorylated on depolarization and calcium influx into nerve terminals in parallel with the coupled events of exocytosis and endocytosis. In late stages of synaptic vesicle retrieval dynamin I undergoes stimulated assembly into a collar, or fission ring, that surrounds the neck of recycling synaptic vesicles. Activation of GTP hydrolysis probably then generates the free synaptic vesicle, which can be refilled with neurotransmitters. This targeting and assembly may involve sequential steps including recruitment of AP-2 to synaptotagmin on the synaptic vesicle, and recruitment of amphiphysin, dynamin I, and synaptojanin. In addition to synaptic vesicle retrieval, dynamin has been associated with intracellular events mediated by growth factor receptors, insulin receptors and the beta-adrenergic receptor. This is likely to reflect targeting of these receptors for endocytosis soon after their activation. However, does it also suggest a broader role for dynamin in other aspects of intracellular signalling pathways?
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PMID:Dynamin, endocytosis and intracellular signalling (review). 911 59

Vertebrate m-calpain, calpastatin, constitutive nitric oxide synthase, myelin basic protein, and dynamin I are substrates of protein kinase C (PKC). The presence/absence of similar/related protein in nonvertebrate was investigated by immunological methods, including (1) affinity chromatography on agarose-secondary antibodies and agarose IgG for removal of nonspecific immunoreactivities from crude extracts; (2) omitting beta-mercaptoethanol treatment and boiling prior to SDS-PAGE to increase the immunoreactivity; (3) immunoreactivity comparisons of nonspecific IgG as controls with specific anti-(vertebrate PKC-substrates/related proteins) in Western blots. It was found that (a) m-calpain and dynamin I were absent in baker's yeast, wheat germ and lobster tail muscle, (b) m-calpain, nitric oxide synthase, myelin basic protein and dynamin II were present in all three samples, and (c) calpastatin was present in baker's yeast and lobster tail muscle. The presence and absence of these proteins suggest evolutionary conservation and divergence, respectively, of these PKC substrates.
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PMID:Immunoreactivities of m-calpain, calpastatin, nitric oxide synthase, myelin basic protein and dynamin II in baker's yeast, wheat germ and lobster tail muscle. 921 26

Dynamin, a 100 kD GTPase, is necessary for the normal development and function of mammalian neural tissue. In neurons, it is necessary for the biogenesis of synaptic vesicles, and in other cell types dynamin has a general and important role in clathrin mediated receptor endocytosis. Different isoforms function as molecular scissors either during the formation of coated vesicles from plasma membrane coated pits, or during the release of intracellular vesicles from donor membranes. The mechanism entails the formation of a horseshoe-shaped dynamin polymer at the neck of the budding vesicle, followed by neck scission through a GTP hydrolysis dependent activity. The primary sequence of dynamin contains several C-terminal SH3 binding proline motifs, a central pleckstrin homology (PH) domain, and an N-terminal GTPase domain. Each of these domains appears to play a distinct role in dynamin function. Dynamin is activated by stimulus coupled PKC phosphorylation in brain, possibly mediated through PKC interactions with the PH domain. Further, SH3 domain interactions with the C-terminal sequences and phophatidylinositol/G beta gamma interactions with the PH domain also increase dynamin GTPase activity. Each of these various regulatory mechanisms is important in dynamin function during vesicle budding, although the means by which these mechanisms integrate in the overall function of dynamin remains to be elucidated.
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PMID:The role of the PH domain and SH3 binding domains in dynamin function. 937 20

Protein kinase C (PKC) has been implicated in integrin-mediated spreading and migration. In mammary epithelial cells there is a partial co-localization between beta1 integrin and PKCalpha. This reflects complexes between these proteins as demonstrated by fluorescense resonance energy transfer (FRET) monitored by fluorescence lifetime imaging microscopy and also by coprecipitation. Constitutive complexes are observed for the intact PKCalpha and also form with the regulatory domain in an activation-dependent manner. Expression of PKCalpha causes upregulation of beta1 integrin on the cell surface, whereas stimulation of PKC induces internalization of beta1 integrin. The integrin initially traffics to an endosomal compartment in a Ca(2+)/PI 3-kinase/dynamin I-dependent manner and subsequently enters an endocytic recycling pathway. This induction of endocytosis by PKCalpha is a function of activity and is not observed for the regulatory domain. PKCalpha, but not PKCalpha regulatory domain expression stimulates migration on beta1 integrin substrates. This PKCalpha-enhanced migratory response is inhibited by blockade of endocytosis.
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PMID:PKCalpha regulates beta1 integrin-dependent cell motility through association and control of integrin traffic. 1040 96

The pleckstrin homology (PH) domain, identified in numerous signaling proteins including the beta-adrenergic receptor kinase (betaARK), was found to bind to various phospholipids as well as the beta subunit of heterotrimeric G proteins (Gbeta) [Touhara, K., et al. (1994) J. Biol. Chem. 269, 10217-10220]. Several PH domain-containing proteins are also substrates of protein kinase C (PKC). Because RACK1, an anchoring protein for activated PKC, is homologous to Gbeta (both contain seven repeats of the WD-40 motif), we determined (i) whether a direct interaction between various PH domains and RACK1 occurs and (ii) the effect of PKC on this interaction. We found that recombinant PH domains of several proteins exhibited differential binding to RACK1. Activated PKC and the PH domain of beta-spectrin or dynamin-1 concomitantly bound to RACK1. Although PH domains bind acidic phospholipids, the interaction between various PH domains and RACK1 was not dependent on the phospholipid activators of PKC, phosphatidylserine and 1, 2-diacylglycerol. Binding of these PH domains to RACK1 was also not affected by either inositol 1,4,5-triphosphate (IP(3)) or phosphatidylinositol 4,5-bisphosphate (PIP(2)). Our in vitro data suggest that RACK1 binds selective PH domains, and that PKC regulates this interaction. We propose that, in vivo, RACK1 may colocalize the kinase with its PH domain-containing substrates.
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PMID:RACK1, a protein kinase C anchoring protein, coordinates the binding of activated protein kinase C and select pleckstrin homology domains in vitro. 1052 23


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