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)

Normal human epidermal keratinocytes (NHEK) grown in serum-free medium on a plastic substrate spontaneously differentiate at high cell densities in vitro. Because protein kinase C (PKC) regulates murine keratinocyte differentiation triggered by a variety of stimuli, we examined the role of this signaling pathway in density-dependent activation of NHEK differentiation. Relative to subconfluent cultures, confluent NHEK expressed markedly higher levels of multiple differentiation markers assayed by immunoblotting, including keratin 1, loricrin, filaggrin, involucrin, TGK, and SPR-1. Expression of several of these markers continued to increase for several days after cells reached confluency. The total level of several PKC isoforms was not substantially altered in NHEK harvested at different cell densities, based on immunoblotting; however, subcellular fractionation revealed that PKCalpha underwent a redistribution to the particulate fraction in confluent and postconfluent NHEK cultures, suggesting that this isozyme was activated under these conditions and may be involved in triggering the terminal differentiation program. Supporting this concept, inhibition of PKC function using bryostatin 1 or GF 109203X blocked the induction of keratinocyte differentiation markers at high cell densities. These data suggest that endogenous activation of PKC is responsible for cell density-mediated stimulation of NHEK differentiation, establishing a critical role for this pathway in regulating human as well as murine keratinocyte differentiation.
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PMID:Differentiation of cultured human epidermal keratinocytes at high cell densities is mediated by endogenous activation of the protein kinase C signaling pathway. 980 35

Human involucrin (hINV) mRNA level and promoter activity increase when keratinocytes are treated with the differentiating agent, 12-O-tetradecanoylphorbol-13-acetate (TPA). This response is mediated via a p38 mitogen-activated protein kinase-dependent pathway that targets activator protein 1 (Efimova, T., LaCelle, P. T. , Welter, J. F., and Eckert, R. L. (1998) J. Biol. Chem. 273, 24387-24395). In the present study we examine the role of various PKC isoforms in this regulation. Transfection of expression plasmids encoding the novel PKC isoforms delta, epsilon, and eta increase hINV promoter activity. In contrast, neither conventional PKC isoforms (alpha, beta, and gamma) nor the atypical isoform (zeta) regulate promoter activity. Consistent with these observations, promoter activity is inhibited by the PKCdelta-selective inhibitor, rottlerin, but not by Go-6976, an inhibitor of conventional PKC isoforms, and novel PKC isoform-dependent promoter activation is inhibited by dominant-negative PKCdelta. This regulation appears to be physiologically important, as transfection of keratinocytes with PKCdelta, -epsilon, or -eta increases expression of the endogenous hINV gene. Synergistic promoter activation (>/=100-fold) is observed when PKCepsilon- or -eta-transfected cells are treated with TPA. In contrast, the PKCdelta-dependent response is more complex as either activation or inhibition is observed, depending upon PKCdelta concentration.
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PMID:Regulation of human involucrin promoter activity by novel protein kinase C isoforms. 1063 51

Previous studies suggest that a PKC/Ras/MEKK1 cascade regulates involucrin (hINV) gene expression in human epidermal keratinocytes. MEK7, which is expressed in epidermis, has been identified as a member of this cascade (Efimova, T., LaCelle, P., Welter, J. F., and Eckert, R. L. (1998) J. Biol. Chem. 273, 24387-24395 and Efimova, T., and Eckert, R. L. (2000) J. Biol. Chem. 275, 1601-1607). However, the kinase that functions downstream of MEK7 has not been identified. Our present studies show that MEK7 expression in keratinocytes markedly activates p38alpha and modestly activates JNK. Activation of p38 MAPK by MEK7 is a novel finding, as previous reports have assigned MEK7 as a JNK regulator. We also demonstrate that this regulation is physiologically important, as the p38alpha- and JNK-dependent activities regulate hINV promoter activity and expression of the endogenous hINV gene.
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PMID:MEK7-dependent activation of p38 MAP kinase in keratinocytes. 1124 91

The hairless (hr) gene is a putative transcriptional factor whose mutations lead to hair loss in animals and humans. As a step toward understanding the role of the hr gene, we investigated the expression of hr mRNA in mouse keratinocyte differentiation. Treatment of mouse primary keratinocyte cultures with phorbol-12-myristate-13-acetate (PMA) reduced DNA synthesis and sequentially induced an up-regulation of p21Cip1/WAF1 (p21), hr and involucrin (inv) mRNAs in a time-dependent fashion, suggesting that an increase in hr gene expression is associated with keratinocyte differentiation. This up-regulation was blocked by the RNA synthesis inhibitor actinomycin D. However, an increase in hr mRNA, but not in inv mRNA, was seen in cells treated with the protein synthesis inhibitor cycloheximide, suggesting that new protein synthesis is involved in the suppression of hr transcription or in the degradation of hr mRNA in the steady state. The up-regulation of hr mRNA expression by PMA was blocked by the protein kinase C (PKC) inhibitor, GF109203X. These data indicate that PKC activation is involved in the up-regulation of hr mRNA expression during mouse keratinocyte differentiation.
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PMID:Protein kinase C is involved in the regulation of hairless mRNA expression during mouse keratinocyte differentiation. 1137 77

Keratinocyte proliferation and differentiation result from expression of specific groups of genes regulated by unique combinations of transcription factors. To better understand these regulatory processes, we studied HOXA7 expression and its regulation of differentiation-specific keratinocyte genes. We isolated the homeobox transcription factor HOXA7 from keratinocytes through binding to a differentiation-dependent viral enhancer and analyzed its effect on endogenous differentiation-dependent genes, primarily transglutaminase 1. HOXA7 overexpression repressed transglutaminase 1-reporter activity. HOXA7 message markedly decreased, and transglutaminase RNA increased, upon phorbol ester-induced differentiation, in a protein kinase C-dependent manner. Overexpression of HOXA7 attenuated the transglutaminase 1 induction by phorbol ester, demonstrating that HOXA7 expression is inversely related to keratinocyte differentiation, and to transglutaminase 1 expression. Antisense HOXA7 expression activated transglutaminase 1, involucrin, and keratin 10 message and protein levels, demonstrating that endogenous HOXA7 down-regulates multiple differentiation-specific keratinocyte genes. In keeping with these observations, epidermal growth factor receptor activation stimulated HOXA7 expression. HOX genes function in groups, and we found that HOXA5 and HOXB7 were also down-regulated by phorbol ester. These results provide the first example of protein kinase C-mediated homeobox gene regulation in keratinocytes, and new evidence that HOXA7, potentially in conjunction with HOXA5 and HOXAB7, silences differentiation-specific genes during keratinocyte proliferation, that are then released from inhibition in response to differentiation signals.
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PMID:Human homeobox HOXA7 regulates keratinocyte transglutaminase type 1 and inhibits differentiation. 1143 35

A signaling cascade that includes protein kinase C (PKC), Ras, and MEKK1 regulates involucrin (hINV) gene expression in epidermal keratinocytes (Efimova, T., LaCelle, P., Welter, J. F., and Eckert, R. L. (1998) J. Biol. Chem. 273, 24387-24395 and Efimova, T., and Eckert, R. L. (2000) J. Biol. Chem. 275, 1601-1607). Because signal transfer downstream of MEKK1 may involve several MAPK kinases (MEKs), it is important to evaluate the regulatory role of each MEK isoform. In the present study we evaluate the role of MEK6 in transmitting this signal. Constitutively active MEK6 (caMEK6) increases hINV promoter activity and increases endogenous hINV levels. The caMEK6-dependent increase in gene expression is inhibited by the p38 MAPK inhibitor, SB203580, and is associated with a marked increase in p38alpha MAPK activity; JNK and ERK kinases are not activated. In addition, hINV gene expression is inhibited by dominant-negative p38alpha and increased when caMEK6 and p38alpha are co-expressed. caMEK6 also activates p38delta, but p38delta inhibits the caMEK6-dependent activation. These results suggest that MEK6 increases hINV gene expression by regulating the balance between activation of p38alpha, which increases gene expression, and p38delta, which decreases gene expression.
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PMID:MEK6 regulates human involucrin gene expression via a p38alpha - and p38delta -dependent mechanism. 1145 75

Culture models of target cells are anticipated to help elucidate the mechanism by which inorganic arsenic acts as a carcinogen in humans. Present work characterizes the response of human keratinocytes, a target cell type, to arsenic suppression of their differentiation program. Four representative differentiation marker mRNAs (involucrin, keratinocyte transglutaminase, small proline-rich protein 1, and filaggrin) were suppressed by both arsenate and arsenite in normal, spontaneously immortalized (premalignant), and malignant keratinocytes with EC50 values in the low micromolar range. The suppression was almost completely reversed 9 days after removal of arsenate from the culture medium. In the case of the involucrin gene, suppression was mediated primarily by two functional AP1 response elements in the gene promoter. Both glucocorticoid and serum stimulation of differentiation occurred to a similar extent in the presence and absence of arsenic, indicating neither stimulation was a specific target of arsenic action and neither agent could overcome arsenic suppression. In contrast, 12-O-tetradecanoylphorbol-13-acetate prevented the suppression of keratinocyte transglutaminase, suggesting that arsenic acts upstream of protein kinase C.
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PMID:Keratinocyte differentiation marker suppression by arsenic: mediation by AP1 response elements and antagonism by tetradecanoylphorbol acetate. 1148 91

Several nuclear factors, called coactivators, such as CREB (cAMP response element binding protein)-binding protein (CBP) and p300/CBP associated factor (P/CAF), have intrinsic histone acetyltransferase (HAT) activity. Recent studies have shown that, in addition to histones, transcriptional regulatory molecules are also targets of HATs, and nuclear acetylation is thought to be involved in several biological events. We observed that a high concentration of calcium induced HAT activity in the keratinocyte cell line, HaCaT. The steady-state level of specific acetylated nuclear proteins changed in a dynamic fashion in HaCaT cells induced with 1.2 mm calcium. One (approximately 97-kDa acetylated protein designated as ap97) was transiently induced, one (ap78) was induced and then continuously expressed, and one (ap70) disappeared with time. Although the up-regulation of ap70 and ap78 was not influenced by GF109203X, a specific inhibitor of protein kinase C (PKC), the calcium-induced accumulation of ap97 and the induction of P/CAF HAT activity were similarly attenuated by GF109203X. Notably, mutant P/CAF lacking HAT activity repressed the expression of ap97 and involucrin, a keratinocyte differentiation marker. Our results suggest that P/CAF HAT activity and induction of ap97 are involved in calcium-dependent keratinocyte differentiation.
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PMID:Possible role of transcriptional coactivator P/CAF and nuclear acetylation in calcium-induced keratinocyte differentiation. 1174 39

Calcium is an important physiologic regulator of keratinocyte function that may regulate keratinocyte differentiation via modulation of protein kinase C (PKC) activity. PKCalpha and PKCdelta are two PKC isoforms that are expressed at high levels in keratinocytes. In the present study, we examine the effect of PKCdelta and PKCalpha on calcium-dependent keratinocyte differentiation as measured by effects on involucrin (hINV) gene expression. Our studies indicate that calcium increases hINV promoter activity and endogenous hINV gene expression. This response requires PKCdelta, as evidenced by the observation that treatment with dominant-negative PKCdelta inhibits calcium-dependent hINV promoter activity, whereas wild type PKCdelta increases activity. PKCalpha, in contrast, inhibits calcium-dependent hINV promoter activation, a finding that is consistent with the ability of dominant-negative PKCalpha and the PKCalpha inhibitor, Go6976, to increase hINV gene expression. The calcium-dependent regulatory response is mediated by an AP1 transcription factor-binding site located within the hINV promoter distal regulatory region that is also required for PKCdelta-dependent regulation; moreover, both calcium and PKCdelta produce similar, but not identical, changes in AP1 factor expression. A key question is whether calcium directly influences PKC isoform function. Our studies show that calcium does not regulate PKCalpha or delta levels or cause a marked redistribution to membranes. However, tyrosine phosphorylation of PKCdelta is markedly increased following calcium treatment. These findings suggest that PKCalpha and PKCdelta are required for, and modulate, calcium-dependent keratinocyte differentiation in opposing directions.
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PMID:Calcium-dependent involucrin expression is inversely regulated by protein kinase C (PKC)alpha and PKCdelta. 1186 71

The novel protein kinase C (nPKC) isoforms are important regulators of human involucrin (hINV) gene expression during keratinocyte differentiation (Efimova, T., and Eckert, R. L. (2000) J. Biol. Chem. 275, 1601-1607). Although the regulatory mechanism involves mitogen-activated protein kinase (MAPK) activation, the role of individual MAPK isoforms has not been elucidated. We therefore examined the effects of individual nPKCs on MAPK activation. We observe unique changes whereby nPKC expression simultaneously increases p38 activity and decreases ERK1 and ERK2 activity. Although p38 alpha, p38 beta, and p38 delta are expressed in keratinocytes, only a single isoform, p38 delta, accounts for the increased p38 activity. Parallel studies indicate that this isoform is also activated by treatment with the keratinocyte regulatory agents, 12-O-tetradecanoylphorbol-13-acetate, calcium, and okadaic acid. These changes in MAPK activity are associated with increased C/EBP alpha transcription factor expression and DNA binding to the hINV promoter and increased hINV gene expression. Expression of PKC delta, PKC epsilon, or PKC eta causes a 10-fold increase in hINV promoter activity, whereas C/EBP alpha expression produces a 25-fold increase. However, simultaneous expression of both proteins causes a synergistic 100-fold increase in promoter activity. These responses are eliminated by the dominant-negative C/EBP isoform, GADD153, and are also inhibited by dominant-negative forms of Ras, MEKK1, MEK3, and p38. These results suggest that the nPKC isoforms produce a unique shift in MAPK activity via a Ras, MEKK1, MEK3 pathway, to increase p38 delta and inhibit ERK1/2 and ultimately increase C/EBP alpha binding to the hINV promoter and hINV gene expression.
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PMID:Novel protein kinase C isoforms regulate human keratinocyte differentiation by activating a p38 delta mitogen-activated protein kinase cascade that targets CCAAT/enhancer-binding protein alpha. 1208 77


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