Identification and characterization of the cis-acting elements of human CD155 gene core promoter

The CD155 protein is the founding member of a new group or related molecule s within the immunoglobulin superfamily sharing a V-C2-C2 domain structure and significant amino acid identity, We have recently isolated the promoter of the CD155 gene so that we may determine the transcription factors that regulate its expression and possibly gain insight into the cell biology of this gene. Here we report the mapping of three cis-elements within the CD15 5 core promoter, designated FPI, II, and III. The results of linker scannin g mutagenesis suggest that all three of these cis elements are required in varying degrees for the promoter activity of the core promoter fragment. Th e relative contribution of each region ranked in the following order: III > II > I. Interestingly, footprint and electrophoretic mobility shift assays show that FPIII binding activity is much reduced in a human cell line that does not express CD155, Additionally, protein binding to FPI and FPII was also investigated. DNase I footprinting using recombinant hAP-2 alpha indic ated that this transcription factor hound to both the FPI and FPII regions of the CD155 core promoter fragment. Electrophoretic mobility shift assays and supershift analysis confirmed the binding of AP-2 from crude nuclear ex tracts to FPI and to FPII. Lastly, cotransfection of the CD155 promoter wit h an AP-2 alpha expression vector indicates that overexpression of AP-2 alp ha modulated the promoter activity of a CD155 promoter construct.