Cloning and characterization of the 5' flanking region of the sialomucin complex/rat Muc4 gene: Promoter activity in cultured cells

Shari A. Price-Schiavi, Aymee Perez, Roy Barco, Kermit L. Carraway

Research output: Contribution to journalArticle

16 Citations (Scopus)

Abstract

Sialomucin complex (SMC/Muc4) is a heterodimeric glycoprotein complex consisting of a mucin subunit ascites sialoglycoprotein-1 (ASGP-1) and a transmembrane subunit (ASGP-2), which is aberrantly expressed on the surfaces of a variety of tumour cells. SMC is transcribed from a single gene, translated into a large polypeptide precursor, and further processed to yield the mature ASGP-1/ASGP-2 complex. SMC has complex spatial and temporal expression patterns in the normal rat, suggesting that it has complex regulatory mechanisms. A crude exon/intron map of the 5' regions of the SMC/Muc4 gene generated from clones isolated from a normal rat liver genomic DNA library reveals that this gene has a small first exon comprising the 5' untranslated region and signal peptide, followed by a large intron. The second exon appears to be large, comprising the 5' unique region and a large part (probably all) of the tandem repeat domain. This structure is strikingly similar to that reported for the human MUC4 gene. Using PCR-based DNA walking, 2.4 kb of the 5'-flanking region of the SMC/Muc4 gene was cloned and characterized. Promoter-pattern searches yielded multiple motifs commonly found in tissue-specific promoters. Reporter constructs generated from this 2.4 kb fragment demonstrate promoter activity in primary rat mammary epithelial cells (MEC), the human colon tumour cell line HCT-116, and the human lung carcinoma cell line NCI-H292, but not in COS-7 cells, suggesting epithelial cell specificity. Deletion constructs of this sequence transfected into rat MEC or HCT-116 cells demonstrate greatly varying levels of activity, suggesting that there are positive and negative, as well as tissue-specific, regulatory elements in this sequence. Taken together, these data suggest that the rat SMC/Muc4 promoter has been identified, that it is tissue- (epithelial cell-) specific, and that there are both positive and negative, as well as tissue-specific, regulatory elements in the sequence.

Original languageEnglish
Pages (from-to)641-649
Number of pages9
JournalBiochemical Journal
Volume349
Issue number2
DOIs
StatePublished - Jul 15 2000

Fingerprint

Sialomucins
Mucin-4
5' Flanking Region
Cloning
Rats
Organism Cloning
Cultured Cells
Genes
Cells
Epithelial Cells
Tissue
Exons
Introns
Tumors
Breast
HCT116 Cells
Tandem Repeat Sequences
Genomic Library
Sequence Deletion
5' Untranslated Regions

Keywords

  • Glycoprotein
  • Mucin
  • Regulatory elements
  • Transcriptional regulation

ASJC Scopus subject areas

  • Biochemistry

Cite this

Cloning and characterization of the 5' flanking region of the sialomucin complex/rat Muc4 gene : Promoter activity in cultured cells. / Price-Schiavi, Shari A.; Perez, Aymee; Barco, Roy; Carraway, Kermit L.

In: Biochemical Journal, Vol. 349, No. 2, 15.07.2000, p. 641-649.

Research output: Contribution to journalArticle

Price-Schiavi, Shari A. ; Perez, Aymee ; Barco, Roy ; Carraway, Kermit L. / Cloning and characterization of the 5' flanking region of the sialomucin complex/rat Muc4 gene : Promoter activity in cultured cells. In: Biochemical Journal. 2000 ; Vol. 349, No. 2. pp. 641-649.
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