scholarly journals Regulation of the human involucrin gene promoter by co-activator proteins

2004 ◽  
Vol 381 (1) ◽  
pp. 267-273 ◽  
Author(s):  
Nhu Q. TRAN ◽  
David L. CROWE
Keyword(s):  
Protein Kinase ◽  
Epithelial Cells ◽  
Promoter Activity ◽  
Binding Protein ◽  
Regulatory Region ◽  
Terminal Differentiation ◽  
Dominant Negative ◽  
Mitogen Activated Protein Kinase ◽  
Activator Proteins ◽  
Involucrin Promoter

Stratified squamous epithelial cells undergo an orderly process of terminal differentiation that is characterized by specific molecular and morphological changes, including expression of the cornified envelope protein involucrin. Significant progress has been made in characterizing the upstream regulatory region of the involucrin gene. Binding sites for AP-1 (activator protein 1) and Sp1 transcription factors were shown to be important for involucrin promoter activity and tissue-specific expression. Defective terminal differentiation is often characterized by decreased or lack of involucrin expression. Recently, a dominant-negative construct of the transcriptional co-activator P/CAF [p300/CBP-associated factor, where CBP stands for CREB (cAMP-response-element-binding protein)-binding protein] was shown to inhibit involucrin expression in immortalized keratinocytes [Kawabata, Kawahara, Kanekura, Araya, Daitoku, Hata, Miura, Fukamizu, Kanzaki, Maruyama and Nakajima (2002) J. Biol. Chem. 277, 8099–8105]. Loss of expression or inactivation of other co-activators has also been demonstrated [Suganuma, Kawabata, Ohshima, and Ikeda (2002) Proc. Natl. Acad. Sci. U.S.A. 99, 13073–13078]. In the present study, we re-expressed CBP and P/CAF in immortalized keratinocyte lines that had lost expression of these co-activator proteins. Re-expression of these proteins restored calcium- and RA (retinoic acid)-responsive involucrin expression in these cells. RA and calcium signalling induced exchange of CBP and P/CAF occupancy at the AP-1 sites of the involucrin promoter. CBP and P/CAF inductions of the involucrin expression were not dependent on MEK (mitogen-activated protein kinase/extracellular-signal-regulated kinase kinase), p38, protein kinase C or CaM kinase (calcium/calmodulin-dependent kinase) signalling. Kinase-induced changes in involucrin promoter activity directly resulted from changes in AP-1 protein expression. We concluded that CBP and P/CAF are important regulators of involucrin expression in stratified squamous epithelial cells.

scholarly journals Lung Krüppel-like factor (LKLF) is a transcriptional activator of the cytosolic phospholipase A2 α promoter

2005 ◽  
Vol 387 (1) ◽  
pp. 239-246 ◽  
Author(s):  
Marilee J. WICK ◽  
Stacy BLAINE ◽  
Vicki VAN PUTTEN ◽  
Milene SAAVEDRA ◽  
Raphael A. NEMENOFF
Keyword(s):  
Epithelial Cells ◽  
Phospholipase A2 ◽  
Promoter Activity ◽  
Cytosolic Phospholipase A2 ◽  
Dominant Negative ◽  
Lung Epithelial Cells ◽  
Mitogen Activated Protein Kinase ◽  
Normal Lung ◽  
Cytosolic Phospholipase ◽  
Lung Epithelial

Increased expression of cPLA2 (cytosolic phospholipase A2) has been shown to be the cause of tumorigenesis of NSCLC (non-small-cell lung cancer). Our laboratory has previously demonstrated that oncogenic forms of Ras increase transcription of cPLA2 in normal lung epithelial cells and NSCLC lines through activation of the ERK (extracellular-signal-regulated kinase) and JNK (c-Jun N-terminal kinase) MAPK (mitogen-activated protein kinase) family. We have also defined a minimal region of the cPLA2 promoter that is critical for this induction. To identify potential transcription factors that bind to this region and regulate expression, a yeast one-hybrid screen was performed with a rat lung cDNA library. Multiple members of the Krüppel family were identified, with LKLF (lung Krüppel-like factor) being isolated a number of times. Overexpression of LKLF in lung epithelial cells or Drosophila SL-2 cells increased cPLA2 promoter activity. Conversely, expression of a dominant negative form of LKLF inhibited induction of cPLA2 promoter activity by oncogenic Ras in normal lung epithelial cells and NSCLC. By electrophoretic mobility-shift assay analysis, it was found that LKLF bound to a GC-rich region of the cPLA2 promoter located between −37 and −30 upstream from the transcription start site. Expression of siRNA (small interfering RNA) directed against LKLF inhibited basal expression of cPLA2 in lung epithelial cells and blocked induction by H-Ras. In NSCLC, siRNA against LKLF co-operated with siRNA against Sp1 (stimulatory protein 1) to inhibit cPLA2 promoter activity. Finally, recombinant LKLF was a substrate for ERKs. These results indicate that LKLF is an important regulator of cPLA2 expression and participates in the induction of this protein, which is critical for increased eicosanoid production associated with lung tumorigenesis.

Involvement of the MAP kinase ERK2 in MUC1 mucin signaling

2001 ◽  
Vol 281 (1) ◽  
pp. L86-L91 ◽  
Author(s):  
Daoud Meerzaman ◽  
Paul S. Shapiro ◽  
K. Chul Kim
Keyword(s):  
Protein Kinase ◽  
Epithelial Cells ◽  
Tyrosine Phosphorylation ◽  
Dominant Negative ◽  
Mitogen Activated Protein Kinase ◽  
Kinase Assay ◽  
Stable Cell Line ◽  
Signaling Mechanism ◽  
Muc1 Mucin ◽  
Mitogen Activated Protein

MUC1 mucin is a receptor-like glycoprotein expressed abundantly in various cancer cell lines as well as in glandular secretory epithelial cells, including airway surface epithelial cells. The role of this cell surface mucin in the airway is not known. In an attempt to understand the signaling mechanism of MUC1 mucin, we established a stable cell line from COS-7 cells expressing a chimeric receptor consisting of the extracellular and transmembrane domains of CD8 and the cytoplasmic (CT) domain of MUC1 mucin (CD8/MUC1 cells). We previously observed that treatment of these cells with anti-CD8 antibody resulted in tyrosine phosphorylation of the CT domain of the chimera. Here we report that treatment of CD8/MUC1 cells with anti-CD8 resulted in activation of extracellular signal-regulated kinase (ERK) 2 as assessed by immunoblotting, kinase assay, and immunocytochemistry. The activation of ERK2 was completely blocked either by a dominant negative Ras mutant or in the presence of a mitogen-activated protein kinase kinase (MEK) inhibitor. We conclude that tyrosine phosphorylation of the CT domain of MUC1 mucin leads to activation of a mitogen-activated protein kinase pathway through the Ras-MEK-ERK2 pathway. Combined with the existing data by others, it is suggested that one of the roles of MUC1 mucin may be regulation of cell growth and differentiation via a common signaling pathway, namely the Grb2-Sos-Ras-MEK-ERK2 pathway.

scholarly journals Regulation of the gene promoter for extracellular signal-regulated protein kinase 2 by transcription factors NF-Y and Sp3

2000 ◽  
Vol 347 (1) ◽  
pp. 155-161 ◽  
Author(s):  
Naoaki SUGIURA ◽  
Kunio TAKISHIMA
Keyword(s):  
Transcription Factors ◽  
Protein Kinase ◽  
Promoter Activity ◽  
Dominant Negative ◽  
Mitogen Activated Protein Kinase ◽  
Flanking Sequence ◽  
Mobility Shift ◽  
Extracellular Signal ◽  
Promoter Deletion Analysis ◽  
Ccaat Box

We have previously shown that the maximal promoter activity of the gene for extracellular signal-regulated protein kinase 2 (ERK2; also known as p42 mitogen-activated protein kinase) resides in the 371 bp 5ʹ-flanking sequence. In the present study we defined roles for a CCAAT box and two adjacent GC boxes in the activity of this promoter. Deletion analysis and DNase I footprinting of this 371 bp region indicated that the CCAAT box at -64 and GC boxes at -86 and -39 are crucial for promoter activity. Electrophoretic mobility-shift assays showed that transcription factor NF-Y/CBF binds to the CCAAT box. Sp1 and Sp3, members of the Sp family of transcription factors, bind to the GC boxes of the ERK2 promoter. The binding of Sp3 was predominant over that of Sp1. Disruption by mutation of any of the CCAAT box and GC boxes similarly decreased promoter activity. These three cis elements exhibited a moderate synergy in promoter function. The transactivating role of NF-Y was corroborated by the finding that a dominant-negative form of NF-YA diminished the promoter activity. These results provide clues for refining our understanding of not only the regulation of expression of the gene for ERK2 but also mechanisms by which NF-Y and Sp1/Sp3 regulate transcription.

Leukotriene D4 activates MAPK through a Ras-independent but PKCϵ-dependent pathway in intestinal epithelial cells

2002 ◽  
Vol 115 (9) ◽  
pp. 1883-1893
Author(s):  
Sailaja Paruchuri ◽  
Bengt Hallberg ◽  
Maria Juhas ◽  
Christer Larsson ◽  
Anita Sjölander
Keyword(s):  
Protein Kinase ◽  
Epithelial Cells ◽  
Proliferative Response ◽  
Egf Receptor ◽  
Intestinal Epithelial Cells ◽  
Dominant Negative ◽  
Mitogen Activated Protein Kinase ◽  
Intestinal Epithelial ◽  
Leukotriene D4 ◽  
Mitogen Activated Protein

We have recently shown that leukotriene D4 (LTD4)increases cell survival in intestinal epithelial cells. Here we report and explore the complementary finding that LTD4 also enhances proliferation in these cells. This proliferative response was approximately half of that induced by epidermal growth factor (EGF) and its required activation of protein kinase C (PKC), Ras and the mitogen-activated protein kinase (MAPK) Erk-1/2. EGF also activated Erk-1/2 in these cells; however the EGF-receptor inhibitor PD153035 did not affect the LTD4-induced activation of Erk-1/2. In addition, LTD4 did not induce phosphorylation of the EGF receptor, nor did pertussis toxin (PTX) block EGF-induced activation of Erk-1/2, thus refuting a possible crosstalk between the receptors. Furthermore, LTD4-induced, but not EGF-induced,activation of Erk-1/2 was sensitive to PTX, PKC inhibitors and downregulation of PKCϵ. A definite role for PKCϵ in LTD4-induced stimulation of Erk-1/2 was documented by the inability of LTD4 to activate Erk-1/2 in cells transfected with either the regulatory domain of PKCϵ (an isoform specific dominant-negative inhibitor) or a kinase-dead PKCϵ. Although Ras and Raf-1 were both transiently activated by LTD4, only Raf-1 activation was abolished by abrogation of the PKC signal. Furthermore, the LTD4-induced activation of Erk-1/2 was unaffected by transfection with dominant-negative N17 Ras but blocked by transfection with kinase-dead Raf-1. Consequently, LTD4 regulates the proliferative response by a distinct Ras-independent, PKCϵ-dependent activation of Erk-1/2 and a parallel Ras-dependent signaling pathway.

scholarly journals Prostaglandin E2 Increases Transforming Growth Factor-β Type III Receptor Expression through CCAAT Enhancer-Binding Protein δ in Osteoblasts

2007 ◽  
Vol 21 (11) ◽  
pp. 2713-2724 ◽  
Author(s):  
Thomas L. McCarthy ◽  
Tony H. Pham ◽  
Bianca I. Knoll ◽  
Michael Centrella
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Promoter Activity ◽  
Binding Protein ◽  
Gene Promoter ◽  
Dominant Negative ◽  
Type Iii ◽  
Kinase C ◽  
Enhancer Binding Protein ◽  
Ccaat Enhancer Binding Protein

Abstract Variations in individual TGF-β receptors (TβRs) may modify TGF-β activity and significantly alter its effects on connective tissue growth or repair. Differences in the amount of TβR type III (TβRIII) relative to signal transducing TβRI occur on bone cells during differentiation or in response to other growth regulators. Here we investigated prostaglandin (PG) E2, a potent effector during trauma, inflammation, or mechanical load, on TβR expression in primary osteoblast-enriched cultures. PGE2 rapidly increased TβRIII mRNA and protein expression and enhanced TβRIII gene promoter activity through a discrete region within 0.4 kb of the transcription start site. PGE2 alters osteoblast function through multiple signal-inducing pathways. In this regard, protein kinase A (PKA) activators, PGE1 and forskolin, also enhanced gene expression through the TβRIII gene promoter, whereas protein kinase C activators, PGF2α and phorbol myristate acetate, did not. The stimulatory effect of PGE2 on TβRIII promoter activity was suppressed by a dominant negative PKA-regulatory subunit, but not by dominant negative protein kinase C. PGE2 specifically increased nuclear factor CCAAT enhancer-binding protein δ (C/EBPδ) binding to a half-binding site upstream of the basal TβRIII promoter region, and promoter activity was sensitive to C/EBPδ overexpression and to dominant-negative C/EBPδ competition. In parallel with their effect on TβRIII expression, activators of PKA decreased TGF-β-induced activity. In summary, high levels of PGE2 that occur with inflammation or trauma may, through PKA-activated C/EBPδ, preferentially increase TβRIII expression and in this way delay TGF-β-dependent activation of osteoblasts during the early stabilization phase of bone repair.

scholarly journals Transcriptional Regulators of theSchizosaccharomyces pombe fbp1Gene Include Two Redundant Tup1p-like Corepressors and the CCAAT Binding Factor Activation Complex

Genetics ◽  
2001 ◽  
Vol 157 (3) ◽  
pp. 1205-1215 ◽  
Author(s):  
Rozmin T K Janoo ◽  
Lori A Neely ◽  
Burkhard R Braun ◽  
Simon K Whitehall ◽  
Charles S Hoffman
Keyword(s):  
Protein Kinase ◽  
Fold Increase ◽  
Transcriptional Regulators ◽  
Dominant Negative ◽  
Mitogen Activated Protein Kinase ◽  
Lacz Expression ◽  
Protein Subunit ◽  
Plasmid Library ◽  
Binding Factor ◽  
Mitogen Activated Protein

AbstractThe Schizosaccharomyces pombe fbp1 gene, which encodes fructose-1,6-bis-phosphatase, is transcriptionally repressed by glucose through the activation of the cAMP-dependent protein kinase A (PKA) and transcriptionally activated by glucose starvation through the activation of a mitogen-activated protein kinase (MAPK). To identify transcriptional regulators acting downstream from or in parallel to PKA, we screened an adh-driven cDNA plasmid library for genes that increase fbp1 transcription in a strain with elevated PKA activity. Two such clones express amino-terminally truncated forms of the S. pombe tup12 protein that resembles the Saccharomyces cerevisiae Tup1p global corepressor. These clones appear to act as dominant negative alleles. Deletion of both tup12 and the closely related tup11 gene causes a 100-fold increase in fbp1-lacZ expression, indicating that tup11 and tup12 are redundant negative regulators of fbp1 transcription. In strains lacking tup11 and tup12, the atf1-pcr1 transcriptional activator continues to play a central role in fbp1-lacZ expression; however, spc1 MAPK phosphorylation of atf1 is no longer essential for its activation. We discuss possible models for the role of tup11- and tup12-mediated repression with respect to signaling from the MAPK and PKA pathways. A third clone identified in our screen expresses the php5 protein subunit of the CCAAT-binding factor (CBF). Deletion of php5 reduces fbp1 expression under both repressed and derepressed conditions. The CBF appears to act in parallel to atf1-pcr1, although it is unclear whether or not CBF activity is regulated by PKA.

scholarly journals Attenuation of CHOP-mediated Myocardial Apoptosis in Pressure-overloaded Dominant Negative p38α Mitogen-activated Protein Kinase Mice

2011 ◽  
Vol 27 (5) ◽  
pp. 487-496 ◽  
Author(s):  
Flori R. Sari ◽  
Bambang Widyantoro ◽  
Rajarajan A. Thandavarayan ◽  
Meilei Harima ◽  
Arun Prasath Lakshmanan ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Dominant Negative ◽  
Mitogen Activated Protein Kinase ◽  
Myocardial Apoptosis ◽  
Mitogen Activated Protein
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