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Functional Analysis of Human MUC7 Mucin Gene 5'-Flanking Region in Lung Epithelial Cells

Department of Oral Biology, School of Dental Medicine, University at Buffalo, The State University of New York, Buffalo, New York

Correspondence and requests for reprints should be addressed to Libuse Anna Bobek, Ph.D., Department of Oral Biology, University at Buffalo, The State University of New York, 109 Foster Hall, 3435 Main Street, Buffalo, NY 14214. E-mail: lbobek{at}buffalo.edu

The human MUC7 gene encodes a low-molecular-mass mucin glycoprotein that functions in modulation of microbial flora in the oral cavity and respiratory tracts. MUC7 gene expression is tissue- and cell-specific, with dominant expression in salivary gland acinar cells. To begin to understand the molecular mechanisms responsible for controlling MUC7 gene expression, we analyzed the promoter activity of MUC7 5'-flanking region in a human lung epithelial cell line A549. We demonstrated that MUC7 gene is expressed constitutively in this cell line and is upregulated by TNF- stimulation. The promoter activities of a 2,762-bp fragment of the human genomic DNA (–2,732/+30 bp) and its deletion series, subcloned into a luciferase reporter vector, were characterized at the basal level and under stimulation by TNF-. The results indicated that the minimal functional MUC7 promoter is in the region of –138/+30 bp. This region also revealed the greatest increase in the promoter activity upon TNF- stimulation. Two putative AP1-binding elements and one NF-B–binding element were identified within the proximal promoter. Further analyses demonstrated that mutations of these elements dramatically reduced specific DNA-protein binding ability and reporter gene expression. AP1 elements played an essential role in the constitutive expression, while the NF-B element was crucially important in the response to TNF- stimulation, demonstrating that TNF- activates MUC7 transcription via NF-B signaling pathway.

Key Words: activator protein-1 • human lung epithelial cell line • human TNF- • MUC7 mucin gene promoter • NF-B