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Promoter of the canine tracheobronchial mucin gene

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Abstract

The mucin gene is up-regulated in diseases such as cystic fibrosis (CF) and asthma. To understand the mechanisms involved in transcriptional regulation of mucin gene expression we have characterized the region of the mucin gene up-stream of the transcriptional start site and analysed thecis-acting elements required for mucin promoter activity. We isolated clones from a dog genomic library containing the promoter region for the tracheobronchial mucin gene (TBM). The authenticity of the promoter was tested by nucleotide sequencing, primer extension analysis, electrophoretic mobility shift assay (EMSA) and reporter gene expression analysis. The canineTBM promoter is different from housekeeping gene promoters (as it is not rich in GC content and contains TATA- and CAAT-like sequences) and different from that of regulatory genes (because it contains many TATA- and CAAT-like sequences and multiple transcriptional initiation sites). Reporter gene analysis using canineTBM promoter-chloramphenicol acetyltransferase (CAT) fusion plasmids established the regions responsible for promoter activity and verified the positions of the major mucin transcriptional initiation sites. Reporter gene analysis also established that a region of the canineTBM promoter and first exon containing all of the transcriptional initiation sites is more active in mucin expressing cells (e.g. CT1 cells-immortalized canine tracheal epithelial cells, human CFT1 cells-immortalized tracheal epithelial cells from a CF subject, or HBE1 cells-immortalized tracheal epithelial cells from non-CF subject) than in mucin non-expressing cells (COS7, 3T3), suggesting cell specificity. The promoter region contained cAMP response element (CRE) sequences, and theTBM gene transcription was enhanced when cAMP analogs were added to transfected cells. EMSA indicated the presence of at least two DNA binding proteins in CT1 cells. This is the first report describing the characterization of aTBM gene promoter. The information obtained in the present studies will be valuable in understanding mucin gene regulation in normal and pathological conditions.

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Abbreviations

CF:

cystic fibrosis

CRE:

cAMP response element

CREB:

cAMP response element binding protein

CT:

cholera toxin

CTM:

canine tracheal mucin

EGF:

epidermal growth factor

EMSA:

electrophoretic mobility shift assay

GRE:

glucocorticoid response element

INS:

insulin

PBS:

phosphate buffered saline

RA:

retinoic acid

RT-PCR:

reverse transcriptase polymerase chain reaction

TF:

transferrin

TRE:

thyroid response element

VDRE:

vitamin D response element

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Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology, Basic Science Building, Georgetown University Medical Center, 3900 Reservoir Road, NW, 20007, Washington, DC, USA

    Mukesh Verma, Claudia Blass & Eugene A. Davidson

  2. Memorial Sloan-Kettering Cancer Center, 10021, New York, USA

    Vandavalli V. S. Murthy, Susan Mathew & Debabrata Banerji

  3. Graduate program in Pathobiology, Brown University, 02903-4923, Providence, RI, USA

    Rabinder N. Kurl & Matthew J. Olnes

  4. Department of Medicine, University of North Carolina at Chappel Hill, 27599-7020, NC, USA

    James R. Yankaskas

Authors
  1. Mukesh Verma
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  2. Vandavalli V. S. Murthy
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  3. Susan Mathew
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  4. Debabrata Banerji
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  5. Rabinder N. Kurl
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  6. Matthew J. Olnes
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  7. James R. Yankaskas
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  8. Claudia Blass
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  9. Eugene A. Davidson
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Accession number: L40433.

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Verma, M., Murthy, V.V.S., Mathew, S. et al. Promoter of the canine tracheobronchial mucin gene. Glycoconjugate J 13, 797–807 (1996). https://doi.org/10.1007/BF00702344

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  • DOI: https://doi.org/10.1007/BF00702344

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