A Negative Regulatory Element of the Macrophage-Specific Human Mannose Receptor Gene Represses Its Expression in Nonmyeloid Cells

Abstract

We have cloned the putative promoter of the human mannose receptor gene using the ligation-mediated polymerase chain reaction. This modified polymerase chain reaction method depends upon the ligation of restricted genomic DNA fragments to a sequence of DNA containing a generic primer site. Approximately 400 bp of genomic DNA sequence immediately upstream from the 5′ end of the lectin gene was amplified with this strategy. Primer-extended reverse transcription identified several 5′ ends of the mannose receptor mRNA corresponding to differential use of initiation transcription sites. DNA sequence analysis of the 5′ flanking sequence of the mannose receptor gene indicated the presence of a TATA box and various putative binding sites for several transcription activators. The insertion of the sequence into a plasmid containing a promoterless luciferase reporter gene reveals a promoter activity with a high cell type specificity, efficient expression upon transfection into macrophage type cells, and lack of efficiency upon transfection into nonmyeloid cells. A series of deletion mutants reveals that this cell-type-specific promoter activity is mediated by a negative regulatory element located at the 5′ end of the isolated promoter.