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)

The enzyme activities of the major kinases found within the cytosolic and microsomal fractions of embryonic avian calvaria osteoblasts were assayed for their specificity for various noncollagenous extracellular matrix (ECM) proteins of bone. At least 6 proteins with M(r)'s of 66, 58, 50, 36, 30, and 22 kD out of more than 30 of the noncollagenous proteins of the bone ECM were phosphorylated by the kinase(s) found in both osteoblast cellular fractions. The purification and N-terminal sequence analysis of three of the above proteins, M(r)'s 66 and 58 kD (+50 kD), identified them as chicken bone sialoprotein (BSP) and osteopontin (OPN), respectively. Heparin, a specific inhibitor of factor-independent protein kinase (FIPK) activity, blocked the phosphorylation of all six ECM proteins by the microsomal kinase(s) but only inhibited the phosphorylation of the 66, 50, and 36 kD by the cytosolic enzyme(s). Casein kinase II (a known FIPK) showed a similar phosphorylation pattern of the same bone ECM proteins as the FIPK(s) found in osteoblast cell extracts, while purified cyclic adenosine monophosphate (cAMP)-dependent protein kinase did not phosphorylate any of the ECM proteins. Use of dephosphorylated casein showed that in comparison with casein kinase II, casein was a poor substrate for the FIPK found in the osteoblast cellular extracts. Further studies, using FIPK(s) of osteoblasts and purified chicken OPN or bacterially produced recombinant murine OPN as a substrate, showed that both species of OPN were excellent substrates for the FIPK(s) found in osteoblasts. The phosphorylation of the purified chicken and recombinant mouse OPNs were evaluated by quantitative analysis using commercially available protein kinases. cAMP-dependent kinase showed no phosphorylation of either protein, and cyclic guanodine monophosphate (cGMP)-dependent kinase and protein kinase C incorporated 1.2 and 0.5 mol phosphate/mol OPN, respectively. However, both chicken and mouse OPNs were significantly phosphorylated by casein kinase II (9.3 and 9.0 mol of phosphate/mol of OPN, respectively). These results demonstrate that the noncollagenous proteins of the bone ECM, and in particular OPN, are predominantly phosphorylated by FIPK(s), and this class of kinase is the major enzyme found within the microsomal fraction of osteoblasts.
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PMID:Protein kinases of cultured osteoblasts: selectivity for the extracellular matrix proteins of bone and their catalytic competence for osteopontin. 888 46

Prolactin induces milk protein gene expression in rabbit primary mammary cells without any concomitant cell multiplication. Prolactin or other lactogenic hormones is the major inducer of cell division in the rat lymphoid Nb2 cells. In Nb2 cells, prolactin also rapidly induces the expression of the c-myc gene, and beta-actin and stathmin gene expression is induced more slowly. The possible involvement of casein kinase II (CKII), mitogen-activated protein kinase (MAPK) and protein kinase C (PKC) in these process is not well known. The present work was undertaken to evaluate the effect of prolactin on these protein kinases and to determine the possible involvement of these enzymes in the activity of several genes under the control of the hormone. In rabbit mammary cells, prolactin did not alter CKII activity but did transiently stimulate MAP kinase activity. Prolactin also stimulated Ca(2+)-independent PKC. This effect was visible after 10 min and was maintained for at least 24 hr. Staurosporine, an inhibitor of PKC and of several tyrosine kinases altered Ca(2+)-independent PKC only moderately. In contrast, GF 109203X, a potent and specific inhibitor of PKC, abrogated almost all PKC activity. Staurosporine, but not GF 109203X, prevented the induction of the casein gene by prolactin. In Nb2 cells, prolactin induced a slow stimulation of CKII activity. The hormone did not induce MAP kinase activity. Prolactin stimulated Ca(2+)-independent PKC over periods of 24 hr. GF 109203X, but not staurosporine, inhibited PKC activity, whereas staurosporine but not GF 109203X, inhibited the induction of Nb2 cell multiplication and the accumulation of c-myc, beta-actin and stathmin mRNAs. From these data, it can be concluded that (1) the stimulation of CKII by prolactin in Nb2 cells is concomitant with cell multiplication: (2) MAPK stimulation is not necessary for prolactin to induce Nb2 cell multiplication; and (3) PKC is stimulated in mammary and Nb2 cells, but this stimulation is not required for prolactin to stimulate casein, c-myc, beta-actin and stathmin gene expression and Nb2 cell division.
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PMID:The effect of prolactin on casein kinase II, MAP kinase and PKC in rabbit mammary cells and Nb2 rat lymphoid cells. 898 34

G protein-coupled receptor kinases (GRKs) specifically recognize and phosphorylate the hormone-occupied form of numerous G protein-coupled receptors, ultimately resulting in termination of receptor signaling. While little is presently known about the regulation of GRK function, recent studies suggest a role for protein kinase C (PKC) phosphorylation of the beta-adrenergic receptor kinase in membrane association and activation of the kinase. To assess a potential general role for PKC in regulating GRK function, we characterized the ability of PKC to phosphorylate GRK5, a recently identified member of the GRK family. We demonstrate that GRK5 can be rapidly and stoichiometrically phosphorylated by PKC in vitro. Intact cell studies reveal that GRK5 is also phosphorylated when transiently expressed in COS-1 cells following treatment with the PKC activator, phorbol 12-myristate 13-acetate. In vitro analysis reveals two major sites of PKC phosphorylation within the C-terminal 26 amino acids of GRK5. GRK5 phosphorylation by PKC dramatically reduces its ability to phosphorylate both receptor (light-activated rhodopsin) and non-receptor (casein and phosvitin) substrates. Kinetic analysis reveals an approximately 5-fold increased Km and approximately 3-fold decreased Vmax for rhodopsin, with no change in the Km for ATP. The reduced affinity of PKC-phosphorylated GRK5 for rhodopsin was also evident in a decreased ability to bind to rhodopsin-containing membranes, while direct binding of GRK5 to phospholipids appeared unaltered. These results suggest that PKC might play an important role in modulating the ability of GRK5 to regulate receptor signaling and that GRK phosphorylation by PKC may serve as a disparate mechanism for regulating GRK activity.
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PMID:Regulation of the G protein-coupled receptor kinase GRK5 by protein kinase C. 901 39

The majority of familial Alzheimer disease mutations are linked to the recently cloned presenilin (PS) genes, which encode two highly homologous proteins (PS-1 and PS-2). It was shown that the full-length PS-2 protein is phosphorylated constitutively within its N-terminal domain by casein kinases, whereas the PS-1 protein is not. Full-length PS proteins undergo endoproteolytic cleavage within their hydrophilic loop domain resulting in the formation of approximately 20-kDa C-terminal fragments (CTF) and approximately 30-kDa N-terminal fragments [Thinakaran, G., et al. (1996) Neuron 17, 181-190]. Here we describe the surprising finding that the CTF of PS-1 is phosphorylated by protein kinase C (PKC). Stimulation of PKC causes a 4- to 5-fold increase of the phosphorylation of the approximately 20-kDa CTF of PS-1 resulting in reduced mobility in SDS gels. PKC-stimulated phosphorylation occurs predominantly on serine residues and can be induced either by direct stimulation of PKC with phorbol-12,13-dibutyrate or by activation of the m1 acetylcholine receptor-signaling pathway with the muscarinic agonist carbachol. However, phosphorylation of full-length PS-1 and PS-2 is not altered upon PKC stimulation. In addition, a mutant form of PS-1 lacking exon 10, which does not undergo endoproteolytic cleavage [Thinakaran, G., et al. (1996) Neuron 17, 181-190] is not phosphorylated by PKC, although it still contains all PKC phosphorylation sites conserved between different species. These results show that PKC phosphorylates the PS-1 CTF. Therefore, endoproteolytic cleavage of full-length PS-1 results in the generation of an in vivo substrate for PKC. The selective phosphorylation of the PS-1 CTF indicates that the physiological and/or pathological properties of the CTF are regulated by PKC activity.
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PMID:Proteolytic processing of the Alzheimer disease-associated presenilin-1 generates an in vivo substrate for protein kinase C. 914 40

Nonsegmented negative strand RNA viruses package an RNA-dependent RNA polymerase composed of two subunits, a large protein L and a phosphoprotein P, for transcription and replication of their genome RNAs. The RNA polymerase activity resides within the L protein, while the P protein acts as a transcription factor or transactivator of the polymerase. Since P protein is heavily phosphorylated and phosphorylation is known to regulate function of many viral as well as cellular proteins, the role of phosphorylation of P protein in the gene expression of this group of RNA viruses has recently been investigated. Through expression in bacteria the P protein was produced in large quantity in the nonphosphorylated form and involvement of cellular kinase(s) in its phosphorylation was studied. Casein kinase II and/or protein kinase C have been shown to play a critical role in the activation of P protein in transcription. These findings have opened up a new avenue for studying an important regulatory step in virus gene expression that may lead to the development of an effective antiviral agent.
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PMID:Role of cellular kinases in the gene expression of nonsegmented negative strand RNA viruses. 922 28

The human DNA repair protein apurinic/apyrimidinic endonuclease (APE) is a dual-function protein that has important roles in both the repair of baseless sites that arise in DNA and in regulating the redox state of a number of proteins (Ref-1). Although previous attention has been focused on how the human APE/Ref-1 gene may be regulated at the DNA level, we have instead examined if APE/Ref-1 is phosphorylated, and if so how it may affect DNA repair activity. We demonstrate here that APE/Ref-1 is indeed a substrate for phosphorylation by the serine/threonine casein kinases (CK) I and II and protein kinase C. Notably, although phosphorylation by CKI and protein kinase C had no effect whatsoever on the ability of APE/Ref-1 to act at abasic sites in DNA, phosphorylation by CKII completely abolished DNA repair activity. That phosphorylation was responsible for the loss of abasic repair activity was concluded from experiments showing that inactive APE/Ref-1 could be reversed to an active DNA repair protein with phosphatase treatment. These results may help to explain the mechanism by which APE/Ref-1 switches from one unrelated function to another.
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PMID:The DNA repair activity of human redox/repair protein APE/Ref-1 is inactivated by phosphorylation. 940 49

Dietary protein deficiency is known to alter the protein kinase C activity in various tissues of rats. Protein kinase C activity is influenced by the metabolism of membrane phosphoinositides and phosphatidyl choline (PC). For metabolic studies, hepatocytes have been the cells of choice of various workers. Therefore, studies on protein kinase C and these phospholipids were conducted in hepatocytes of rats maintained on three different diets viz. casein (20% protein) deficient (4% protein, rice flour as source of protein) and supplemented (deficient diet supplemented with L-lysine and DL-threonine) diet for 28 days. The protein deficiency in diet led to a decline in protein kinase C activity (P < 0.01) without effecting its translocation, an increase in phosphatidyl inositol 4,5-bisphosphate (P < 0.001) and a decrease in phosphatidyl inositol 4-monophosphate and phosphatidyl inositol (P < 0.01) but did not alter the PC contents, as compared to the casein group. Supplementation of deficient diet with L-lysine and DL-threonine could considerably reverse the effect of deficiency of protein in diet. The results suggest that quality of dietary protein is mainly relevant for maintaining phospholipid metabolism and physiology of hepatocytes and thus the signalling mechanism in these cells.
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PMID:Dietary protein deficiency induced changes in protein kinase C activity and phospholipid metabolism in rat hepatocytes. 947 78

Casein kinase II (CKII) phosphorylates the rat neuronal growth-associated protein B-50 (GAP-43) at serines 191/192 and threonines 88, 89 and/or 95 both in vitro and in neuronal growth cones. Since little is known concerning regulation of the phosphorylation of these sites, these studies were undertaken to characterize the factors which determine the degree of B-50 phosphorylation by CKII in vitro. Phosphorylation of rat B-50 on serine and threonine residues by recombinant human CKII is stimulated by polylysine. Maximal stimulation occurs at 10 microg/ml of polylysine, a concentration which has no effect on protein kinase C (PKC)-mediated phosphorylation of B-50. Digestion with Staphylococcus aureus V8 protease demonstrates CKII-mediated phosphorylation of B-501-132 and the C-terminal fragment S3/S4. Phosphorylation of B-50 by either CKII or PKC is inhibited by the N-terminal monoclonal antibody NM2, while the C-terminal antibody NM6 has no effect on phosphorylation by either protein kinase. Protein phosphatase 2A dephosphorylates both the CKII and PKC sites, while protein phosphatases 2B and 1 are more selective for the PKC site. These results indicate that the phosphorylations of B-50 by CKII and PKC are determined by distinct regulatory signals in vivo.
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PMID:Regulation of in vitro phosphorylation of the casein kinase II sites in B-50 (GAP-43). 950 76

The catalytic domain of p72(syk) kinase (CDp72(syk)) was purified from a 30000 g particulate fraction of rat spleen. The purification procedure employed sequential chromatography on columns of DEAE-Sephacel and Superdex-200, and elution from HA-Ultrogel by chloride. The analysis of the final CDp72(syk) preparation by SDS/PAGE revealed a major silver-stained 40 kDa protein. The kinase was identified by covalent modification of its ATP-binding site with [14C]5'-fluorosulphonylbenzoyladenosine and by immunoblotting with a polyclonal antibody against the 'linker' region of p72(syk). By using poly(Glu4, Tyr1) as a substrate, the specific activity of the enzyme was determined as 18.5 nmol Pi/min per mg. Casein, histones H1 and H2B and myelin basic protein were efficiently phosphorylated by CDp72(syk). The kinase exhibited a limited ability to phosphorylate random polymers containing tyrosine residues. CDp72(syk) autophosphorylation activity was associated with an activation of the kinase towards exogenous substrates. The extent of activation was dependent on the substrates added. CDp72(syk) was phosphorylated by protein kinase C (PKC) on serine and threonine residues. With a newly developed assay method, we demonstrated that the PKC-mediated phosphorylation had a strong activating effect on the tyrosine kinase activity of CDp72(syk). Studies extended to conventional PKC isoforms revealed an isoform-dependent manner (alpha > betaI = betaII > gamma) of CDp72(syk) phosphorylation. The different phosphorylation efficiencies of the PKC isoforms closely correlated with the ability to enhance the tyrosine kinase activity.
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PMID:Purification of catalytic domain of rat spleen p72syk kinase and its phosphorylation and activation by protein kinase C. 953 9

Osteopontin (OP) is a highly phosphorylated bone matrix protein and contains the RGD cell-binding motif, which mediates cell adhesion through integrin receptors that include alpha(v)beta3. Casein kinase 2 (CK2) is a factor-independent serine/threonine kinase, which may be the predominant physiologically relevant kinase for OP phosphorylation. This study was designed to examine the effects of unphosphorylated recombinant rat OP, and CK2-phosphorylated OP (P-OP), on the adhesion and function of mouse osteoclasts (OC) and osteoblast-like cells (UMR 201-10B and UMR 106-06) in vitro. OP significantly increased OC adhesion compared to plastic alone, and cell attachment was further increased at least twofold on OP phosphorylated with CK2. Attachment was dependent on the integrity of the RGD domain and was completely abolished in the presence of 1 mM RGD peptide. Neither CK2 phosphorylation of mutant OP, in which the RGD was converted to RGE or RAD, nor protein kinase C (PKC) phosphorylation of wild-type OP enhanced OC attachment. An antibody to the beta3 integrin subunit, but not anti-mouse CD44 antibody, specifically blocked the proportion of attachment due to phosphorylation of OP. Actin ring formation in OC was increased by plating cells onto OP, with no further increase by phosphorylation. Both OP and CK2-phosphorylated OP enhanced attachment of the two osteoblastic cell lines, compared to plastic, but in contrast to OCs, there was no significant difference with phosphorylation. Osteoblast attachment was totally blocked by 1 mM RGD peptide, but was not influenced by the beta3 integrin antibody. Plating of UMR 201-10B cells onto OP further increased retinoic acid-induced alkaline phosphatase expression. The results suggest that specific phosphorylation of OP is important for interaction with OCs, compared with osteoblastic cells, and that alternative integrins may be important in the interaction between osteoblastic cells and OP compared with OCs.
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PMID:Casein kinase 2 phosphorylation of recombinant rat osteopontin enhances adhesion of osteoclasts but not osteoblasts. 961 57


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