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

The phosphorylation of cytokeratin was investigated in primary cultures of hepatocytes. The two hepatocyte cytokeratins CK8 and CK18 (55,000 and 49,000 M(r) respectively) were phosphorylated, CK8 being more phosphorylated than CK18. Treatment of the hepatocytes with 150 nM 12-O-tetradecanoyl-phorbol-13-acetate (TPA) an activator of protein kinase C induced a transient increase in the level of phosphorylation of CK8 but not CK18. This effect was maximal after 15 min of TPA treatment and was maintained for up to 3 h. After 22 h of treatment with TPA, which down-regulates protein kinase C, CK8 phosphorylation was returned to the basal level. Further addition of TPA to the 22-h treated cells did not cause an increase in CK8 phosphorylation. Indirect immunofluorescence microscopy with a monoclonal antibody to CK8 indicated that while the addition of TPA induced the formation of granular cytokeratin aggregates in some hepatocytes, in most hepatocytes no major changes in the intermediate filament network were observed. Staining for actin showed that actin microfilaments were rapidly reorganized after the treatment and a loss of stress fibres were observed. We propose that CK8 is an in vivo substrate for protein kinase C and that the specific phosphorylation of CK8 plays a role in protein kinase C signal transduction.
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PMID:Differential phosphorylation of CK8 and CK18 by 12-O-tetradecanoyl-phorbol-13-acetate in primary cultures of mouse hepatocytes. 128 12

A 40-kD protein kinase C (PKC)epsilon related activity was found to associate with human epithelial specific cytokeratin (CK) polypeptides 8 and 18. The kinase activity coimmunoprecipitated with CK8 and 18 and phosphorylated immunoprecipitates of the CK. Immunoblot analysis of CK8/18 immunoprecipitates using an anti-PKC epsilon specific antibody showed that the 40-kD species, and not native PKC epsilon (90 kD) associated with the cytokeratins. Reconstitution experiments demonstrated that purified CK8 or CK18 associated with a 40-kD tryptic fragment of purified PKC epsilon, or with a similar species obtained from cells that express the fragment constitutively but do not express CK8/18. A peptide pseudosubstrate specific for PKC epsilon inhibited phosphorylation of CK8/18 in intact cells or in a kinase assay with CK8/18 immunoprecipitates. Tryptic peptide map analysis of the cytokeratins that were phosphorylated by purified rat brain PKC epsilon or as immunoprecipitates by the associated kinase showed similar phosphopeptides. Furthermore, PKC epsilon immunoreactive species and CK8/18 colocalized using immunofluorescent double staining. We propose that a kinase related to the catalytic fragment of PKC epsilon physically associates with and phosphorylates cytokeratins 8 and 18.
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PMID:PKC epsilon-related kinase associates with and phosphorylates cytokeratin 8 and 18. 137 67

Continuous epithelial cell lines from individuals with cystic fibrosis (CF) and normal controls are required to understand the genetic and cellular defects in CF. We used retroviruses to transduce SV40 large T antigen into nasal epithelial cells. Transformed continuous cell lines were isolated that expressed epithelial markers, cytokeratin, and tight junctions. Northern blot analysis shows that all of the cell lines express the putative CF gene mRNA. Studies of transepithelial electrolyte transport show that CF and normal cell lines develop a transepithelial electrical resistance. Normal but not CF cell lines secreted Cl- in response to agonists that increase cellular levels of adenosine 3',5'-cyclic monophosphate (cAMP) (isoproterenol, forskolin, and a membrane-permeant analogue of cAMP) or in response to a tumor-promoting phorbol ester that activates protein kinase C. In contrast, the Ca2(+)-elevating agonist bradykinin and the Ca2+ ionophore A23187 stimulated secretion in both normal and CF cell lines. The continuous cell lines we have produced maintain their proper phenotypes and will serve as useful tools in understanding the pathophysiology of CF.
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PMID:Expression of normal and cystic fibrosis phenotypes by continuous airway epithelial cell lines. 170 80

PLC/PRF/5 human hepatoma cells cultured with teleocidin reduced the rate of cell proliferation and were transformed into large cells with many vacuole-like subcellular structures. In these vacuolated cells, the protein content per cell increased without changing the total cellular protein synthesis. Cytokeratin was one of the proteins which increased quantitatively. This intermediate filament formed fibrous network structures throughout the enlarged cytoplasm. The assembly of other cytoskeletal proteins such as actin, tubulin, and vimentin was not altered remarkably, suggesting that teleocidin morphologically transformed the hepatoma cells by changing the assembly of cytokeratin protein selectively. On the other hand, the alterations of cell proliferation, cell morphology, and cytokeratin assembly induced by teleocidin were not associated with either down-regulation of protein kinase C or reduced number of epidermal growth factor receptors. In addition, these teleocidin effects were not mimicked by the protein kinase C agonist 1-oleoyl-2-acetylglycerol or inhibited by the protein kinase C inhibitor 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine. From these results it can be speculated that the morphological transformation and reduced cell proliferation induced by teleocidin may be mediated by still unknown mechanisms unrelated to protein kinase C.
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PMID:Vacuole formation and cytokeratin rearrangement of hepatoma cells induced by teleocidin are not associated with down-regulation of protein kinase C. 170 98

The phosphorylation of epithelial-specific cytokeratin (CK) 8 and 18 was studied in the human colonic cell line HT29. Metabolic labelling of cells with orthophosphate resulted in phosphorylation of cytokeratins 8/18 on serine residues. When phorbol acetate was added to labelled cells, a 2.2-fold increase in CK8/18 phosphate labelling was noted, whereas increasing intracellular cAMP levels using forskolin or 8-Br-cAMP showed no significant change in CK phosphorylation. CKs8/18 were also phosphorylated by added PKC in the presence of [gamma-32P]ATP. Tryptic peptide map analysis of the phosphorylated CK8 species showed that treatment of cells with 8-Br-cAMP or phorbol acetate generated a phosphopeptide not seen in control cells. In contrast, tryptic peptide maps of phosphorylated CK18 showed no discernable differences. Our results support a role for PKC in the phosphorylation of epithelial cytokeratins, with some phosphorylation sites being modulated by cAMP dependent protein kinase.
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PMID:Phorbol acetate enhances the phosphorylation of cytokeratins 8 and 18 in human colonic epithelial cells. 170 19

Differentiation of bone-marrow-derived precursor cells into mature mouse T lymphocytes occurs in the thymus and involves sequential interactions with MHC-positive hemopoietic and epithelial stromal cells. To study the in vitro molecular mechanisms at play during the lympho-epithelial cell adhesion, we derived thymic stromal cell lines which were shown to possess cytokeratin filaments and tight junctions. These mouse thymic epithelial (MTE) cell lines did not express the classical hemopoietic stromal cell surface markers (i.e. LFA-1, Mac-1, and CD45) but expressed ICAM-1, NCAM, J11d, CD44, and MHC molecules. A quantitative cell adhesion assay was used to evaluate the interaction of various lymphoid cell subsets with MTE cells. Two cell interaction patterns could be defined: first, a rapid adhesion of a fraction of CD4+CD8+ and of a few CD4-CD8- immature thymocytes to MTE cells was observed at 4 degrees C. The CD8 molecule was shown to be partially involved in this initial contact. The strength of adhesion between MTE cells and distinct thymocyte subsets was evaluated and found to be maximal with neonatal thymocytes. Second, a temperature-dependent adhesion step characterized by a rapid and active stabilization of the interaction of MTE cells with 20% of CD4+CD8+CD3low thymocytes was seen, followed by a more progressive de-adhesion step. This active process of engagement was highly LFA-1-dependent, involved the CD4 and CD8 molecules, and required protein kinase C activation and cytoskeletal integrity. The results are consistent with the involvement of LFA-1 in a transient and regulated cell adhesion under the control of the TCR-CD3 complex that progressively appears on maturing cells. This phenomenon might contribute to the selection of a subset of immature thymocytes by epithelial cells occurring during the process of maturation of these cells.
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PMID:Mouse thymic epithelial cell lines interact with and select a CD3lowCD4+CD8+ thymocyte subset through an LFA-1-dependent adhesion--de-adhesion mechanism. 215 May 94

A human epithelial cell line, WISH, and a mouse cell line, LB6-uPAR, transfected with the human urokinase receptor (uPAR), both expressed high affinity uPAR but undetectable levels of urokinase (uPA). In two independent assays, binding of exogenous pro-uPA produced an up to threefold enhancement of migration. The migration was time and concentration dependent and did not involve extracellular proteolysis. This biologic response suggested that uPAR can trigger an intracellular signal. Since this receptor is a glycosyl-phosphatidylinositol-linked protein, we postulated that it must do so by interacting with other proteins, among which, by analogy to other systems, would be a kinase. To test this hypothesis, we carried out a solid phase capture of uPAR from WISH cell lysates using either antibodies against uPAR or pro-uPA adsorbed to plastic wells, followed by in vitro phosphorylation of the immobilized proteins. SDS-PAGE and autoradiography revealed two phosphorylated protein bands of 47 and 55 kD. Both proteins were phosphorylated on serine residues. Partial sequence of the two proteins showed a 100% homology to cytokeratin 18 (CK18) and 8 (CK8), respectively. A similar pattern of phosphorylation was obtained with lysates from A459 cells, a lung carcinoma, but not HL60, LB6-uPAR or HEp3 cell lysates, suggesting that the identified multiprotein uPAR-complex may be specific for simple epithelia. Moreover, immunocapture with antibody to another glycosyl-phosphatidylinositol-linked protein, CD55, which is highly expressed in WISH cells, was ineffective. The kinase was tentatively identified as protein kinase C, because it was inhibited by an analogue of staurosporine more specific for PKC and not by a PKA or tyrosine kinase inhibitors. The kinase was tentatively identified as PKC epsilon because of its resistance to PMA down-modulation, independence of Ca2+ for activity, and reaction with a specific anti-PKC epsilon antibody in Western blots. Cell fractionation into cytosolic and particulate fractions revealed that all four proteins, the kinase, uPAR, CK18, and CK8, were present in the particulate fraction. In vivo, CK8, and to a lesser degree CK18, were found to be phosphorylated on serine residues. Occupation of uPAR elicited a time-dependent increase in the phosphorylation intensity of CK8, a cell shape change and a redistribution of the cytokeratin filaments. These results strongly suggest that uPAR serves not only as an anchor for uPA but participates in a signal transduction pathway resulting in a pronounced biological response.
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PMID:Induction of cell migration by pro-urokinase binding to its receptor: possible mechanism for signal transduction in human epithelial cells. 751 43

To learn whether autophagy might be dependent on any of the major cytoskeletal elements, the effect of various cytoskeleton inhibitors on autophagy and cytoskeletal organization was studied in isolated rat hepatocytes. Autophagy, measured as the sequestration of endogenous lactate dehydrogenase, was completely inhibited in isolated rat hepatocytes by the protein phosphatase inhibitor okadaic acid (30 nM). Only small effects were seen with vinblastine (10 microM) or cytochalasin D (10 microM). Indirect immunofluorescence microscopy with antibody to a 55-kDa cytokeratin, corresponding to human cytokeratin 8 (CK8), revealed that whereas control cells contained a well-organized network of cytokeratin intermediate filaments, okadaic acid disrupted this network into small spherical aggregates. Treatment with cytochalasin D or vinblastine, which disrupt microfilaments and microtubules, respectively, had no detectable effect on the cytokeratin filament distribution. Neither the microtubule network (detected by indirect immunofluorescence with antibodies against alpha- and beta-tubulin) nor the actin microfilament network (detected by rhodamine-palloidin) was disrupted by okadaic acid. Naringin (100 microM), a putative protein kinase-inhibitory flavonoid, offered complete protection against the autophagy-inhibitory and cytokeratin-disruptive effects of okadaic acid. Two other flavonoids, genistein (100 microM) and prunin (100 microM), as well as KN-62 (10 microM), a specific inhibitor of Ca2+/calmodulin-dependent kinase II), likewise displayed a good ability to protect against the effect of okadaic acid upon cytokeratin organization, while no such protection was seen with H-89 (20 microM), an inhibitor of the cyclic nucleotide-dependent protein kinases, or with H-7 (100 microM), which in addition inhibits protein kinase C. The results suggest that the cytokeratin cytoskeleton of hepatocytes is subject to rapid control by phosphorylation and dephosphorylation and that cytokeratin filaments may somehow be involved in the autophagic process.
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PMID:Disruption of the cytokeratin cytoskeleton and inhibition of hepatocytic autophagy by okadaic acid. 754 Sep 86

The sheets serve as an maternal supply of assembled, cytokeratin, intermediate filaments. They are remodeled at each major developmental transition in mammalian early development, that is fertilization, embryonic compaction, blastocyst formation, and formation of the primitive ectoderm and primitive endoderm during implantation into the uterine wall. Our results indicate that the sheets exist as specialization for placental development as they have a major role in the maintenance of epithelial integrity at the time the embryo is implanting into the uterine wall. They also contribute intermediate filaments to the junctional complexes required for embryonic compaction. Our analyses demonstrate the they are regulated at the time of fertilization by the action of PKC/PKM, a kinase that acts as a cellular chronometer with both temporal and spatial precision that remodels the egg into the zygote.
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PMID:Remodeling of the specialized intermediate filament network in mammalian eggs and embryos during development: regulation by protein kinase C and protein kinase M. 874 68

The expression of cholesterol sulfate (CS) is known to increase during squamous differentiation of keratinocytes and to activate the epsilon, eta, and zeta forms of protein kinase C as a signal transduction molecule for the subsequent expression of transglutaminase-1 (TG-1) and cytokeratins. To gain further insight into the regulation of cellular differentiation and tumorigenesis by CS, we examined the concentration and the potential for synthesis of CS in seven and four surgical specimens from human ovarian and uterine cervical cancer patients, respectively, and eight cell lines established from human uterine cervical cancer patients and compared them for the rate of expression of cytokeratin. CS was present in all of the uterine cervical cancer tissue specimens but only in the mucinous type of cystadenocarcinoma among ovarian cancer tissue specimens, and cytokeratin was highly expressed in the tissues with a high concentration of CS, which were classified as well-differentiated on the basis of morphological examination. Similarly, cells derived from a keratinizing type of well-differentiated cervical carcinoma demonstrated strong potential for synthesis of CS, stained positive with anti-cytokeratin antibody, and exhibited a higher specific activity of TG-1, whereas the cells without CS did not stain positive with anti-cytokeratin antibody and exhibited a lower specific activity of TG-1. These findings indicate that CS is coexpressed with TG-1 and cytokeratin in the well-differentiated types of squamous cell cancers as a tumor marker.
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PMID:Coexpression of cholesterol sulfate and cytokeratin as tumor markers in well-differentiated squamous cell carcinoma of the human uterine cervix. 986 10


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