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J Biol Chem, Vol. 273, Issue 19, 11737-11744, May 8, 1998
From the Immunology Research Center, St Vincent's Hospital,
Fitzroy 3065, Victoria, Australia
The expression of intercellular adhesion molecule
2 (ICAM-2) in adult tissues is restricted to vascular endothelial cells and megakaryocytes. We have previously shown that the
endothelial-specific in vivo activity of the human ICAM-2
promoter is contained within a small (0.33-kilobase (kbp)) 5'-flanking
region of the gene. Here we describe the in vitro
characterization of this region. The ICAM-2 promoter is TATA-less, and
transcription in endothelial cells initiates at four sites. Reporter
gene expression directed by the promoter was 125-fold greater than
vector alone in bovine aortic endothelial cells but less than 2-fold
vector alone in non-endothelial (COS) cells, confirming that
specificity in vivo was paralleled in vitro.
The addition of 2.7 kbp of 5'-flanking region to the 0.33-kbp fragment
had no effect on promoter activity or specificity. The mutation of an
Sp1 motif centered at base pair
The Human ICAM-2 Promoter is Endothelial Cell-specific in
Vitro and in Vivo and Contains Critical Sp1 and GATA
Binding Sites
194 or an eight-base pair palindrome
at 268 each reduced promoter activity by 70%. Mutation of GATA
motifs at 145 and 53 reduced promoter activity by 78 and 61%,
respectively. Specific binding of bovine aortic endothelial cells
nuclear proteins to the Sp1 and GATA sites was demonstrated by gel
shift analysis. Promoter activity in COS cells was transactivated
3-4-fold by overexpression of GATA-2. The results presented here
suggest that transcription from the ICAM-2 promoter in endothelial
cells is regulated by the interplay of several positive-acting factors and provide the basis for further analysis of endothelial-specific gene
expression.
Copyright © 1998 by The American Society for Biochemistry and Molecular Biology, Inc.
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