Nucleic Acids Research, 1992, Vol. 20, No. 18 4765-4772
© 1992
MOLECULAR BIOLOGY |
Cooperative effects of C/EBP-like and NFxB-like binding sites on rat serum amyloid A1 gene expression in liver cells
Department of Biochemistry and Molecular Biology, The University of Texas M.D.Anderson Cancer Center 1515 Holcombe Blvd., Houston, TX 77030, USA
*To whom correspondence should be addressed
Received June 1, 1992. Accepted August 11, 1992.
Serum amyloid A (SAA) is a major acute-phase protein synthesized and secreted mainly by the liver. In response to inflammation, its expression is increased by 1000-fold, primarily because of a 200-fold increase in the rates of SAA gene transcription. We have shown that when 304 bp of 5' flanking region of the rat SAA1 gene is fused to a reporter gene, the chloramphenicol acetyltransferase (CAT) gene, CAT activity is induced in a cell-specific manner in response to conditioned media prepared from activated mixed lymphocyte cultures and recombinant interleukin-1. In this study, deletion of the SAA1 promoter to –120 bp with respect to the transcriptional start site did not diminish promoter activity; however, deletion to –94 bp renders the promoter completely inactive. Functional analysis have demonstrated that a 66–bp DNA fragment spanning –138 bp to –73 bp could confer cytokine responsiveness to a heterologous thymidine kinase promoter. Within this 66–bp responsive element resided an NFxB-llke-binding site and a C/EBP-like-binding site. Although each binding site alone could confer responsiveness when stimulated with conditioned media and TPA, the response was much weaker than that observed when both sites were present. Moreover, site-specific mutations of either binding site completely abolished SAA1 promoter activity. Taken together, these results suggest a functional importance for and cooperative interaction of these two nuclear-factor binding sites in the cytokine-induced expression of the rat SAA1 gene.
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