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)

We investigated the effects of the non-phorbol tumor promoter okadaic acid on human leukemia K562 cells. It was found that okadaic acid potently and reversibly inhibited cell growth, with a nearly complete inhibition of thymidine uptake seen at about 10 nM. The cytotoxicity of okadaic acid was characterized by a marked mitotic arrest of the cells exhibiting scattered chromosomes and abnormal anaphase-like structures, a phenomenon distinct from the typical metaphase arrest caused by colchicine. Okadaic acid (10-1,000 nM) greatly stimulated phosphorylation of a number of nuclear proteins in K562 cells. Phosphorylation of many of the same proteins was also stimulated by 12-O-tetradecanoylphorbol-13-O-acetate, a protein kinase C activator. The present findings, consistent with recent reports that okadaic acid is a potent inhibitor of protein phosphatases 1 and 2A (PP1 and PP2A) shown to be essential for normal mitosis, provided evidence for the first time that okadaic acid inhibition of PP1/PP2A resulted in enhanced nuclear protein phosphorylation and subsequent mitotic arrest.
 ...
PMID:Mitotic arrest and enhanced nuclear protein phosphorylation in human leukemia K562 cells by okadaic acid, a potent protein phosphatase inhibitor and tumor promoter. 164 33

Arachidonate activation of the NADPH-oxidase in intact neutrophils and in a cell-free O2- generation system was compared to synergistic activation in response to arachidonate and agents that effect protein phosphorylation. In intact neutrophils, suboptimal doses of retinal which increase protein phosphorylation, or 4B-phorbol 12-myristate 13-acetate (PMA) an activator of protein kinase C, induced minimal O2- release, but primed neutrophils to release enhanced amounts of O2- in response to 2.5 microM arachidonate. In contrast to retinal or PMA, okadaic acid, a specific inhibitor of serine/threonine protein phosphatases, did not induce any release of O2-, but significantly increased the maximal rate and duration of O2- release in response to arachidonate. In the cell-free system, only arachidonate induced O2- generation. Consistent with previous findings, activation of the cell-free system was dependent of the presence of light membranes, cytosol, NADPH, Mg2+, and 82 microM arachidonate. Pretreatment of neutrophils with suboptimal doses of PMA or retinal had little effect on the arachidonate-stimulated release of O2- in cell-free preparations of these cells. However, cytosol (but not light membranes) from PMA or retinal-primed neutrophils was more effective in completing resting membrane NADPH-oxidase activity when compared to cytosol from resting cells. The addition of protein kinase C inhibitors staurosporine and 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine decreased the effectiveness of PMA-primed cytosol to complete the cell-free system, but had little effect on cytosol obtained from cells primed with retinal. The addition of protein phosphatase inhibitors, p-nitrophenyl phosphate or okadaic acid to neutrophil cavitates increased 3-fold the release of O2- in cell-free preparations of these cells. Okadaic acid and p-nitrophenyl phosphate also increased the effectiveness of both cytosol and light membranes to complete the cell-free system when combined with cytosol or light membranes from resting neutrophils, respectively, indicating that both fractions are affected by the inhibition of protein phosphatase activity. These data indicate that increases in protein phosphorylation alone do not lead to the activation of the NADPH-oxidase, but in addition to the requirement of an anionic amphiphile, the release of O2- from intact neutrophils or in the cell-free system is increased by stimulus activation of protein kinase C or more impressively by inhibition of protein phosphatase activity.
 ...
PMID:Arachidonate activation of the neutrophil NADPH-oxidase. Synergistic effects of protein phosphatase inhibitors compared with protein kinase activators. 165 30

Okadaic acid, a potent tumor promoter and inhibitor of phosphoserine/threonine protein phosphatases 1 and 2A, produces a large increase in epidermal growth factor (EGF) receptor phosphorylation in several cell types. The increases are limited to phosphoserine and phosphothreonine residues. 12-O-Tetradecanoylphorbol-13-acetate (TPA), a distinct tumor promoter and protein kinase C activator, also induces serine/threonine phosphorylation of the EGF receptor and is known to modulate receptor functions. Comparison of okadaic acid and TPA influences on the EGF receptor show significant differences. Okadaic acid did not promote phosphorylation of Thr-654, a major site of TPA-induced phosphorylation. However, other sites of phosphorylation were similar for the two tumor promoters. In vitro experiments with purified protein phosphatase 2A demonstrate the insensitivity of Thr-654 phosphorylation, which regulates EGF receptor function, to dephosphorylation by this okadaic acid-sensitive protein phosphatase. In contrast to TPA, okadaic acid did not attenuate the tyrosine kinase activity or ligand binding capacity of the EGF receptor. However, okadaic acid did produce a decrease in EGF-stimulated inositol phosphate formation in a manner distinct from that of TPA.
 ...
PMID:Okadaic acid-induced hyperphosphorylation of the epidermal growth factor receptor. Comparison with receptor phosphorylation and functions affected by another tumor promoter, 12-O-tetradecanoylphorbol-13-acetate. 165 56

Okadaic acid, dinophysistoxin-1 (35-methylokadaic acid), and calyculin A are the okadaic acid class of non-12-O-tetradecanoylphorbol-13-acetate (TPA)-type tumor promoters, which do not bind to the phorbol ester receptors in cell membranes or activate protein kinase C in vitro. They have potent tumor-promoting activities on mouse skin, as strong as TPA-type tumor promoters, such as TPA, teleocidin, and aplysiatoxin. DNA samples isolated from tumors induced by dimethylbenz[alpha]anthracene and each of the okadaic acid class tumor promoters had the same mutation at the second nucleotide of codon 61 (CAA to CTA) in the c-H-ras gene. Okadaic acid receptors, protein phosphatases 1 and 2A, are present in the particulate as well as cytosolic fractions of various mouse tissues. The apparent "activation" of protein kinases by the okadaic acid class tumor promoters, after their incubation with 32P-ATP, protein kinases, and protein phosphatases, was observed. This activation was caused by inhibition of protein phosphatases 1 and 2A by the okadaic acid class tumor promoters. Treatment of primary human fibroblasts and human keratinocytes with the okadaic acid class tumor promoters induced the hyperphosphorylation of a 60-kDa protein in nuclear and cytosolic fractions, due to the inhibition of protein phosphatases. The 60-kDa protein is a proteolytic fragment of nucleolin, a major nonhistone protein and is designated as "N-60." The mechanisms of action of the okadaic acid class tumor promoters are discussed with emphasis on the inhibition of protein phosphatase activity.
 ...
PMID:Mechanisms of action of okadaic acid class tumor promoters on mouse skin. 166 50

Okadaic acid (OA), a potent inhibitor of types 1 and 2A protein phosphatases, was shown recently to induce chromatin condensation and germinal vesicle breakdown (GVBD) in mouse oocytes arrested at the dictyate stage by dibutyryl cAMP (dbcAMP), isobutyl methylxanthine (IBMX) and 12,13-phorbol dibutyrate (PDBu). We confirm these results using IBMX and another phorbol diester, 12-O-tetradecanoylphorbol-13-acetate (TPA) and show that OA also bypasses the inhibitory effect of 6-dimethylaminopurine (6-DMAP). It has been concluded that protein phosphatases 1 and/or 2A (PP1, 2A), involved in the negative control of MPF activation, are thus operating downstream from both the protein kinase A and protein kinase C catalysed phosphorylation steps that prevent the breakdown of GV. Similar enzymatic activities are also able to counteract the general inhibition of protein phosphorylation. However, PP1 and/or PP2A are positively involved in the activation of pericentriolar material (PCM) into microtubule organizing centres (MTOCs). This explains the inhibitory effect of OA on spindle assembly. Finally, OA interferes with the integrity and/or function of actomyosin filaments. This results in a dramatic ruffling of the plasma membrane leading to the internalization of large vacuoles, the inhibition of chromosome centrifugal displacement and, consequently, the prevention of polar body extrusion.
 ...
PMID:Pleiotropic effect of okadaic acid on maturing mouse oocytes. 171 79

We have recently reported a potent mitogenic stimulation of oligodendroglial (OL) progenitors by the protein kinase C (PKC) activating phorbol ester, i.e., phorbol 12-myristate 13-acetate (PMA) (Bhat NR, J Neurosci Res 22:20-27, 1989). The present study deals with PMA-induced protein phosphorylation reactions in cultured OL progenitors. The phorbol ester induced the phosphorylation of several cytosol and membrane-associated proteins, including a major protein with an apparent molecular weight of 80 kDa. In both control and PMA-treated cultures, phosphorylation level of the 80-kDa protein in cytosol was higher than that in the particulate fraction. Okadaic acid, an inhibitor of protein phosphatases, also increased the phosphorylation of several proteins and substantially enhanced protein phosphorylation induced by PMA. In vitro incubation of the cell membranes with phosphatidylserine and diacylglycerol (a physiological activator of PKC) in the presence of [gamma 32p]-ATP resulted in an increased phosphorylation of the 80-kDa protein. The induction of phosphorylation of the 80-kDa protein under both in situ and in vitro conditions was subject to inhibition by 1-[5[isoquinolinyl sulfonyl)-3-methylpiperazine (H-7), a potent inhibitor of PKC. The 80-kDa phosphoprotein was identified as the prominent PKC substrate, i.e., myristoylated alanine-rich C-kinase substrate (MARCKS) protein by immunoprecipitation with anti-MARCKS antibodies.
 ...
PMID:Phosphorylation of MARCKS (80-kDa) protein, a major substrate for protein kinase C in oligodendroglial progenitors. 179 60

Urokinase-type plasminogen activator (uPA) gene expression in LLC-PK1 cells is induced by activation of cAMP-dependent protein kinase (cAMP-PK) or protein kinase C (PK-C). To determine whether protein phosphatases can also modulate uPA gene expression, we tested okadaic acid, a potent specific inhibitor of protein phosphatases 1 and 2A, in the presence and absence of cAMP-PK and PK-C activators. Okadaic acid by itself induced uPA mRNA accumulation. This induction was strongly attenuated by the inhibition of protein synthesis. In contrast, the inhibition of protein synthesis enhanced induction by 8-bromo-cAMP and only delayed induction by 12-O-tetradecanoylphorbol-13-acetate (TPA). In addition, down-regulation of PK-C by chronic treatment with TPA did not abrogate the okadaic acid-dependent induction. These results provide evidence for a novel signal transduction pathway leading to gene regulation that involves protein phosphorylation but is independent of both cAMP-PK and PK-C.
 ...
PMID:Okadaic acid induction of the urokinase-type plasminogen activator gene occurs independently of cAMP-dependent protein kinase and protein kinase C and is sensitive to protein synthesis inhibition. 184 95

Most, if not all, signal transduction at cell surface receptors in animal cells appears to occur by one of four basic mechanisms, 1) cyclic nucleotide systems; 2) phosphoinositidase systems; 3) ion channel systems; 4) tyrosine kinase systems. The end effects of all four signal transduction systems are largely mediated by protein (serine/threonine) kinases and/or phosphatases. Thus, the only known high affinity intracellular receptors for cyclic nucleotides and diacylglycerol are cyclic nucleotide-dependent protein kinases and protein kinase C respectively, while activation of tyrosine kinase systems causes concomitant activation of several different protein (serine/threonine) kinases. Many, although not all, effects of elevated Ca2+ are also mediated by calmodulin-dependent protein kinases. Initial tests of the involvement of any of these kinases in control of a physiological system can be made using cell-permeable kinase activators or inhibitors, e.g. cyclic nucleotide analogues, phorbol esters or Ca2+ ionophores. A family of four protein (serine/threonine) phosphatases account for dephosphorylation of all known cytosolic or nuclear substrates phosphorylated by these protein kinases. Two of these (PP1 and PP2A) have a broad substrate specificity and appear to be highly conserved during evolution, being present in both animal and plant kingdoms. PP1 is potently inhibited by protein inhibitors-1 and -2, while the marine toxin and tumour promoter, okadaic acid, inhibits PP2A with extreme potency and PP1 less potently. Okadaic acid provides a novel cell-permeable probe for examining the role of protein phosphorylation, and PP1 and PP2A in particular, in any physiological process. Recent examples of its use are discussed. These approaches can provide initial evidence that a particular protein is regulated in response to an extracellular signal by protein phosphorylation. Confirmation of this hypothesis may be obtained by showing that the precise residue(s) phosphorylated by the protein kinase in a cell-free system are also phosphorylated in intact cells in response to the extracellular signal. Sensitive methods are now available for the analysis of phosphorylation sites by gas phase sequencing and Fast Atom Bombardment (FAB) mass spectrometry. Sequencing of phosphorylation sites also allows the development of synthetic peptide assays for the various kinases involved. These methods will be illustrated using the author's own studies on phosphorylation of enzymes of lipid metabolism.
 ...
PMID:Roles of protein kinases and phosphatases in signal transduction. 196 36

Phorbol esters, acting via activation of the protein kinase C family of protein serine/threonine kinases, are able to exert profound effects on various cellular functions. In this study, we used the EL4 thymoma cell line to study the potential role of "downstream" protein serine/threonine kinases in cellular responses to phorbol esters. In wild-type EL4 cells, addition of phorbol ester caused a rapid activation of kinase activity toward RRLSSLRA (S6P). This increased activity was maintained for at least 15 min but diminished to control levels by 60 min. Activation of a myelin basic protein (MBP) kinase was also seen in response to phorbol ester. In a variant EL4 cell line in which phorbol ester does not induce interleukin 2 transcription, phorbol ester failed to activate either the S6P kinase or MBP kinase. Partial purification of the activated S6P and MBP kinases from wild-type cells showed that they represent separate enzymes that are distinct from protein kinase C. Although the variant cells had reduced levels of protein kinase C as compared with the wild-type cells, the amount of membrane-bound enzyme increased in response to phorbol 12-myristate 13-acetate in both wild-type and variant cells. Treatment of intact cells with phorbol ester resulted in phosphorylation of some of the same protein substrates in both cell lines. Okadaic acid, a phosphatase inhibitor, increased S6P and MBP kinase activities in both wild-type and variant cells. Thus, phorbol ester failed to activate the S6P and MBP kinases in the variant cells even though these cells express activatable protein kinase C, S6P kinase, and MBP kinase. Two protein kinase inhibitors, staurosporine and H-7, inhibited the activity of all three kinases in vitro, while a peptide inhibitor (PKC 19-31) showed specificity for protein kinase C. In summary, these results suggest that activation of messenger-independent protein kinases may be critical for certain protein kinase C-dependent responses.
 ...
PMID:Activation of messenger-independent protein kinases in wild-type and phorbol ester-resistant EL4 thymoma cells. 198 54

The turnover and processing of the Alzheimer beta/A4 amyloid precursor protein (beta APP) has been studied in PC12 cells after treatment with agents that regulate protein phosphorylation. Phorbol 12,13-dibutyrate, an agent that stimulates protein kinase C, decreased the levels of mature beta APP and increased the levels of 15- and 19-kDa peptides. These peptides appeared to be COOH-terminal fragments of beta APP, which arose when phorbol 12,13-dibutyrate increased the rate of proteolytic processing of mature forms of beta APP. Okadaic acid, an inhibitor of protein phosphatases 1 and 2A, also led to decreased levels of mature beta APP and increased levels of the 15- and 19-kDa peptides. H-7, an inhibitor of protein kinase C and of several other protein kinases, apparently decreased the rate of proteolytic processing of mature beta APP. The sizes of the putative COOH-terminal fragments observed after treatment with either phorbol 12,13-dibutyrate or okadaic acid suggest that one or both may contain the entire beta/A4 region of beta APP and thus be amyloidogenic. Our results support the hypothesis that abnormal protein phosphorylation may play a role in the development of the cerebral amyloidosis that accompanies Alzheimer disease.
 ...
PMID:Processing of Alzheimer beta/A4 amyloid precursor protein: modulation by agents that regulate protein phosphorylation. 211 15


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>