Identification and Characterization of the cis-Acting Elements of the Human CD155 Gene Core Promoter*
- From the ‡Department of Molecular Genetics and Microbiology, School of Medicine, State University of New York at Stony Brook, Stony Brook, New York 11794 and the¶Max-Delbrück-Centrum für Molekulare Medizin, Robert-Rossle-Strasse 10, 13122 Berlin, Germany
Abstract
The CD155 protein is the founding member of a new group of related molecules 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 CD155 core promoter, designated FPI, II, and III. The results of linker scanning mutagenesis suggest that all three of these cis-elements are required in varying degrees for the promoter activity of the core promoter fragment. The 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-2α indicated that this transcription factor bound 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 extracts to FPI and to FPII. Lastly, cotransfection of the CD155promoter with an AP-2α expression vector indicates that overexpression of AP-2α modulated the promoter activity of aCD155 promoter construct.
Footnotes
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↵* This work was supported in part by National Institutes of Health Grant AI39485.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵§ Member of the graduate program in Molecular and Cellular Biology, SUNY at Stony Brook, and recipient of a grant from the Deutscher Akademischer Austauschdienst.
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↵‖ Supported by Grant BE1886/1-1 from the Deutsche Forschungsgemeinschaft. To whom correspondence should be addressed. Tel.: 49-30-9406-3330; Fax: 49-30-9406-2887.
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↵2 D. Solecki, E. Wimmer, M. Lipp, and G. Bernhardt, unpublished observations.
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↵3 M. Gromeier, D. Solecki, and E. Wimmer, submitted for publication.
- Abbreviations:
- PV
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poliovirus
- bp
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base pair(s)
- EMSA
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electrophoretic mobility shift assay
- PCR
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polymerase chain reaction
- RSV
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Rous sarcoma virus
- RT
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reverse transcription
- BL
-
Burkitt’s lymphoma
- NF
-
nuclear factor.
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- Received June 26, 1998.
- Revision received September 24, 1998.
- The American Society for Biochemistry and Molecular Biology, Inc.
