IDENTIFICATION OF REGULATORY SEQUENCES IN THE PROMOTER OF THE PDGF B-CHAIN GENE IN MALIGNANT MESOTHELIOMA CELL-LINES

Platelet-derived growth factor (PDGF) B-chain mRNA is readily detectab le in malignant mesothelioma (MM) cell lines, but not in normal mesoth elial (NM) cell lines. The high affinity receptor for PDGF B-chain dim ers, the PDGF beta-receptor, is expressed in Mh? cell lines. NM cell l ines predominantly express the PDGF alpha-receptor. Coexpression of th e PDGF beta-receptor and its ligand may lead to an autocrine growth st imulating loop in the malignant cell type. In nuclear run off experime nts, PDGF B-chain mRNA was detectable in MM cells only, indicating an increased level of transcription in this cell type. The proximal promo ter of the PDGF B-chain gene contains DNaseI hypersensitive (DH) sites and mediates reporter gene activation in both normal and malignant ce lls. Nuclear proteins, extracted from both cell types, interact with D NA sequences within the proximal promoter around bp -64 to -61 relativ e to the transcription start site. Electrophoretic mobility shift assa ys (EMSAs) indicate that these factors are more abundantly present in the malignant than in the normal cell type. A DH site around -9.9 kb w as found in both cell types. When tested in CAT assays, this region ex erted a stimulatory effect on transcription in malignant cells. The el evated level of transcription of the PDGF B-chain gene in malignant ce lls may well be the result of interaction of regulatory sites in the p roximal promoter and an enhancing element located at -9.9 kb from the transcription start site.