scholarly journals The lung enriched transcription factor TTF-1 and the ubiquitously expressed proteins Sp1 and Sp3 interact with elements located in the minimal promoter of the rat Clara cell secretory protein gene

1996 ◽  
Vol 316 (2) ◽  
pp. 467-473 ◽  
Author(s):  
Ruud F. G. TOONEN ◽  
Sharon GOWAN ◽  
Colin D. BINGLE
Keyword(s):  
Promoter Region ◽  
Secretory Protein ◽  
Minimal Promoter ◽  
Clara Cell ◽  
Start Site ◽  
Transcription Start ◽  
Mobility Shift ◽  
Clara Cell Secretory Protein ◽  
Gel Mobility Shift ◽  
Minimal Promoter Region

The mechanisms that direct expression of the Clara cell secretory protein (CCSP) gene to the bronchiolar epithelial cells of the lung remain to be elucidated. Previous studies have identified a number of proteins which bind to a functionally important region (Region 1) located -132 to -76 bp from the transcription start site in the rat CCSP gene. Subsequently we have shown that while Region 1 is an important positive regulator of CCSP gene expression, sequences 3′ of this region (-75 to +38) are sufficient to confer tissue-specific expression of a reporter gene. In the present study we have used transient transfections with a deletion series of CCSP–CAT reporter plasmids (where CAT is chloramphenicol acetyltransferase) and gel mobility shift assays with a series of overlapping oligonucleotides covering the whole minimal promoter region to study protein–DNA interactions within this region. These studies have identified a conserved functional binding site for the lung and thyroid enriched homeodomain transcription factor TTF-1, located between positions -51 and -42 from the transcription start site. CCSP–CAT chimaeric reporters containing this region are specifically activated by TTF-1 in co-transfection assays, and nuclear extracts from cells which express TTF-1 bind to this region, as does in vitro translated rat TTF-1. Three additional conserved regions were identified, and in further gel mobility shift studies with an oligonucleotide spanning the conserved region immediately 5′ to the TTF-1 site we identified a binding site for the ubiquitously expressed zinc-finger-containing proteins Sp1 and Sp3. These studies suggest that cell-type-restricted and ubiquitous nuclear proteins may play a combined role in the regulation of the CCSP gene within the bronchiolar epithelium by interacting with the minimal promoter region.

scholarly journals Identification of hepatocyte nuclear factor-3 binding sites in the Clara cell secretory protein gene

1993 ◽  
Vol 295 (1) ◽  
pp. 227-232 ◽  
Author(s):  
C D Bingle ◽  
J D Gitlin
Keyword(s):  
Gene Expression ◽  
Binding Sites ◽  
Secretory Protein ◽  
Clara Cell ◽  
Mouse Lung ◽  
Specific Gene ◽  
Nuclear Factor ◽  
Hepatocyte Nuclear Factor ◽  
Mobility Shift ◽  
Clara Cell Secretory Protein

To determine the mechanisms of cell-specific gene expression in the developing pulmonary epithelium the Clara cell secretory protein (CCSP) gene promoter was analysed by DNAase I footprinting. A prominent site of protein-DNA interaction was detected from nucleotides -132 to -76 using nuclear extract from mouse lung and human H441 cells. Mobility shift analysis revealed that an oligonucleotide corresponding to this region interacted with multiple proteins from lung and H441 cell nuclear extracts. Analysis of the nucleotide sequence of this region identified two potential binding sites for hepatocyte nuclear factor 3 (HNF-3), and consistent with this finding binding to this CCSP oligonucleotide was specifically competed for by an oligonucleotide corresponding to the HNF-3-binding site from the mouse transthyretin gene. Mobility shift of the CCSP oligonucleotide was supershifted using antisera specific to HNF-3 alpha and HNF-3 beta, and HNF-3 alpha and HNF-3 beta translated in vitro were found to bind specifically to this same oligonucleotide. Co-transfection of HNF-3 alpha- and HNF-3 beta-expression plasmids increased cell-specific reporter gene activity in H441 cells transfected with a CCSP-CAT gene chimeric construct containing this -132 to -76 region. Taken together, these results suggest a role for HNF-3 in mediating cell-specific CCSP gene expression within the bronchiolar epithelium. These findings support the hypothesis that members of the HNF-3 ‘forkhead’ family of transcription factors determine gene expression and cell fate in multiple cell lineages derived from the primitive gut endoderm.

Structure and Regulation of the Murine Clara Cell Secretory Protein Gene

Genomics ◽  
1994 ◽  
Vol 20 (1) ◽  
pp. 27-35 ◽  
Author(s):  
Barry R. Stripp ◽  
Jacquelyn A. Huffman ◽  
Robert J. Bohinski
Keyword(s):  
Protein Gene ◽  
Secretory Protein ◽  
Clara Cell ◽  
Clara Cell Secretory Protein

Identification of a cell-type-specific transcriptional repressor in the promoter region of the mouse hepatocyte growth factor gene

1994 ◽  
Vol 14 (11) ◽  
pp. 7046-7058
Author(s):  
Y Liu ◽  
A B Beedle ◽  
L Lin ◽  
A W Bell ◽  
R Zarnegar
Keyword(s):  
Epithelial Cells ◽  
Promoter Region ◽  
Nuclear Protein ◽  
Transcriptional Repressor ◽  
Cell Type ◽  
Mobility Shift ◽  
A Cell ◽  
Cell Type Specific ◽  
Gel Mobility Shift ◽  
Hepatocyte Growth

Hepatocyte growth factor (HGF), a cytokine with multiple functions, exhibits cell-type-specific as well as cytokine- and steroid hormone-regulated expression. The HGF gene is known to be expressed predominately in mesenchymal but not in epithelial cells. In this study, we report the identification of a cell-type-specific transcriptional repressor in the promoter region of the mouse HGF gene, which is evidently responsible for the suppression of HGF expression in epithelial cells. Gel mobility shift assays and DNase I footprinting studies revealed that a 27-bp element (-16 to +11) around the transcription initiation site is responsible for the binding of a nuclear protein which is present in epithelial but not in mesenchymally derived cells. Further analysis of the binding activity of the DNA region with nuclear protein revealed that an approximately 19-bp sequence containing a unique palindromic structure (5'-AACCGACCGGTT-3') overlapped by a CAP box is essential for binding. Substitution of a single base (the contact site) within this region by site-directed mutagenesis resulted in total abrogation of the binding of the nuclear protein and a concomitant increase in the transcriptional activity of various lengths of HGF-chloramphenicol acetyltransferase fused genes when transfected into the epithelial cell line RL95-2 but not the mesenchymal cell line NIH 3T3. Southwestern (DNA-protein) analyses revealed that the nuclear protein which binds to this repressor element is a single polypeptide of approximately 70 kDa. Analysis of the nuclear extract prepared from regenerating mouse liver at various times after two-thirds partial hepatectomy by gel mobility shift assay revealed a substantial reduction (more than 75% within 3 h) in the binding of the repressor to its cognate binding site. Our results suggest that a cis-acting transcriptional repressor in the promoter region of the mouse HGF gene is involved in cell-type-specific regulation through binding to its cognate trans-acting protein which exists in epithelial cells but is absent in fibroblast cells.

The Expression of Clara Cell Secretory Protein in BAL Fluid of Patients with Idiopathic Interstitial Pneumonia

2002 ◽  
Vol 53 (2) ◽  
pp. 127
Author(s):  
Sang Won Um ◽  
Seon Jin Han ◽  
Chang Min Choi ◽  
Chang Hoon Lee ◽  
Chul Gyu Yoo ◽  
...  
Keyword(s):  
Interstitial Pneumonia ◽  
Secretory Protein ◽  
Idiopathic Interstitial Pneumonia ◽  
Clara Cell ◽  
Clara Cell Secretory Protein

IL-13-induced Clara cell secretory protein expression in airway epithelium: role of EGFR signaling pathway

2002 ◽  
Vol 283 (1) ◽  
pp. L67-L75 ◽  
Author(s):  
Suil Kim ◽  
Jae Jeong Shim ◽  
Pierre-Regis Burgel ◽  
Iris F. Ueki ◽  
Trang Dao-Pick ◽  
...  
Keyword(s):  
Growth Factor ◽  
Protein Expression ◽  
Tyrosine Kinase ◽  
Tyrosine Kinase Inhibitor ◽  
Kinase Inhibitor ◽  
Growth Factor Receptor ◽  
Secretory Protein ◽  
Clara Cell ◽  
Clara Cell Secretory Protein ◽  
Gene And Protein Expression

Previous work showed that the Th2 cytokine interleukin (IL)-13 induces goblet cell metaplasia via an indirect mechanism involving the expression and subsequent activation of epidermal growth factor receptor (EGFR). Because Clara cell secretory protein (CCSP) expression has been reported in cells that express mucins, we examined the effect of IL-13 on CCSP gene and protein expression in pathogen-free rat airways and in pulmonary mucoepidermoid NCI-H292 cells. Intratracheal instillation of IL-13 induced CCSP mRNA in epithelial cells without cilia within 8–16 h, maximal between 24 and 48 h; CCSP immunostaining increased in a time-dependent fashion, maximal at 48 h. The CCSP immunostaining was localized in nongranulated secretory cells and goblet cells and in the lumen. Pretreatment with the selective EGFR tyrosine kinase inhibitor BIBX1522, cyclophosphamide (an inhibitor of bone marrow leukocyte mobilization), or a blocking antibody to IL-8 prevented CCSP staining. Treatment of NCI-H292 cells with the EGFR ligand transforming growth factor-α, but not with IL-13 alone, induced CCSP gene and protein expression. Selective EGFR tyrosine kinase inhibitors, BIBX1522 and AG1478, prevented CCSP expression in NCI-H292 cells, but the platelet-derived growth factor receptor tyrosine kinase inhibitor AG1295 had no effect. These findings indicate that IL-13 induces CCSP expression via an EGFR- and leukocyte-dependent pathway.

Clara cell secretory protein deficiency increases oxidant stress response in conducting airways

1998 ◽  
Vol 275 (2) ◽  
pp. L348-L356 ◽  
Author(s):  
Gregory W. Mango ◽  
Carl J. Johnston ◽  
Susan D. Reynolds ◽  
Jacob N. Finkelstein ◽  
Charles G. Plopper ◽  
...  
Keyword(s):  
Stress Response ◽  
Mrna Expression ◽  
Molecular Basis ◽  
Oxidant Stress ◽  
Secretory Protein ◽  
Protective Role ◽  
Clara Cell ◽  
Clara Cells ◽  
Clara Cell Secretory Protein ◽  
Oxidant Sensitivity

Little is known about the molecular basis for differential pulmonary oxidant sensitivity observed between genetically disparate members of the same species. We have generated mice that are deficient in Clara cell secretory protein (CCSP −/−) and that exhibit an oxidant-sensitive phenotype. We characterized the kinetics and distribution of altered stress-response [interleukin-6 (IL-6) and metallothionein (MT)] and epithelial cell-specific [cytochrome P-450 2F2 (CYP2F2)] gene expression to further understand the cellular and molecular basis for altered oxidant sensitivity in 129 strain CCSP −/− mice. Increases in IL-6 and MT mRNA abundance were detected by 2 h of exposure to 1 part/million ozone and preceded reductions in Clara cell CYP2F2 mRNA expression. Despite being qualitatively similar, increases in IL-6 and MT mRNA expression were enhanced in CCSP −/− mice with respect to coexposed 129 strain wild-type mice. Increased MT mRNA expression, indicative of the stress response, localized to the airway epithelium, surrounding mesenchyme, and endothelium of blood vessels. These results demonstrate a protective role for Clara cells and their secretions and indicate potential genetic mechanisms that may influence susceptibility to oxidant stress.

scholarly journals Determination of the InvE Binding Site Required for Expression of IpaB of the Shigella sonnei Virulence Plasmid: Involvement of a ParB BoxA-Like Sequence

2003 ◽  
Vol 185 (17) ◽  
pp. 5158-5165 ◽  
Author(s):  
Takayuki Taniya ◽  
Jiro Mitobe ◽  
Shu-ichi Nakayama ◽  
Qi Mingshan ◽  
Kenji Okuda ◽  
...  
Keyword(s):  
Dna Binding ◽  
Transcription Start Site ◽  
Promoter Region ◽  
Binding Activity ◽  
Shigella Sonnei ◽  
Virulence Plasmid ◽  
Start Site ◽  
Transcription Start ◽  
Dna Binding Activity ◽  
K 12

ABSTRACT The InvE protein positively regulates the expression of virulence genes ipaBCD in Shigella sonnei. The InvE has significant homology with ParB of plasmid P1, which is known as a plasmid partitioning factor with DNA binding ability. Although the DNA binding activity of InvE has been predicted, it is not known whether the DNA binding activity is necessary for type III secretion system-associated gene expression. In this study, we determined the transcription start site of the icsB-ipaBCD operon (ipa operon) and constructed a series of deletions of the icsB promoter region in the Escherichia coli K-12 background. The deletion study revealed that an 86-bp region upstream of the icsB transcription start site was essential for expression of the ipa operon, where the ParB binding motif (ParB BoxA-like sequence) was observed. Purified glutathione S-transferase-InvE fusion protein bound directly to the −93 to −54 region (designating the icsB transcription start site as nucleotide +1) containing the ParB BoxA-like sequence. These results indicated that InvE bound directly to the promoter region.

Lung Hyperpermeability, Clara-Cell Secretory Protein (CC16), and Susceptibility to Ozone of Five Inbred Strains of Mice

2003 ◽  
Vol 15 (12) ◽  
pp. 1209-1230 ◽  
Author(s):  
Fabrice Broeckaert ◽  
André Clippe ◽  
Ruddy Wattiez ◽  
Paul Falmagne ◽  
Alfred Bernard
Keyword(s):  
Inbred Strains ◽  
Secretory Protein ◽  
Clara Cell ◽  
Clara Cell Secretory Protein ◽  
Inbred Strains Of Mice
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