The human tissue-type plasminogen activator gene expression is regulated in a celltype specific manner by the tumor promoting phorbol ester, phorbol 12-myristate 13-acetate (PMA): t-PA expression is down-regulated by PMA in HT-1080 cells but up-regulated in HeLa cells. A cAMP response element, TGACATCA (referred to as the tPACRE) and an AP-2 like site significantly influence transactivation of the t-PA promoter in both cell types. Supershift experiments have demonstrated a cell-type specific pattern of expression of tPACRE binding proteins in these two cell lines. Here we demonstrate that tPACRE and AP-2 elements are also functionally involved in t-PA promoter transactivation in transfected human umbilical vein endothelial cell line (C11STH cells). Using a panel of antibodies directed against various members of the CREB/ATF and fos/jun families of transcription factors in a supershift assay, we demonstrate that the major tPACRE binding nuclear proteins in C11STH cells are ATF-2, CREB, CREM, and c-jun We found no evidence for ATF-3, ATF-4 or c-fos in these cells. PMA treatment did not significantly alter the tPACRE binding activity of these factors, but did result in the selective recruitment of jun-D The identical tPACRE binding pattern was observed using nuclear proteins extracted from primary human umbilical vein endothelial cells (HUVEC's). Taken together, these results underscore the global significance of the tPACRE and AP-2-like sites in the transactivation of the t-PA gene promoter. Moreover, the identification of ATF-2, CREB, CREM, c-jun and jun-D as endothelial derived proteins which interact with the tPACRE suggests an integral role for these factors in the regulation of t-PA gene expression. We are presently overexpressing dominant negative forms of CREB and ATF-2 to determine the functional roles of these transcription factors in the regulation of the t-PA gene.
|Number of pages||1|
|Journal||Fibrinolysis and Proteolysis|
|Issue number||SUPPL. 3|
|Publication status||Published - 1 Dec 1997|