V-SRC INDUCES PROSTAGLANDIN SYNTHASE-2 GENE-EXPRESSION BY ACTIVATION OF THE C-JUN N-TERMINAL KINASE AND THE C-JUN TRANSCRIPTION FACTOR

A consensus cyclic AMP response element (CRE) in the murine prostaglandin synthase-2 (PGS2) promoter is essential for pgs2 gene expression induced by pp60(v-src) the v-src oncogene product. In this study, we investigate (i) the transcription factors active at the PGS2 ''CRE site'' in response to v-src activation and (ii) the signal transduction pathways by which pp60(v-src) activates these transcription factors. Transient transfection assays with pgs2 promoter/luciferase reporter chimeric genes suggest that c-Jun mediates v-src-induced pgs2 gene expression. Antibody supershift experiments demonstrate that c-Jun can participate in a complex with the pgs2 promoter CRE site. Moreover, in vitro immunocomplex assays demonstrate that pp60(v-src) expression strongly activates c-Jun N-terminal kinase (JNK1) enzyme activity, Serines 63 and 73, the sites of c-Jun phosphorylation by JNK, are essential for v-src-induced, pgs2 promoter mediated luciferase expression. Cotransfection studies with plasmids expressing wild-type JNK, dominant-negative JNK, and dominant-negative MEKK-1 confirm that activation of the Ras/MEKK-1/JNK/c-Jun pathway is required for v-src-induced pgs2 gene expression. Overexpression of either wild type ERK-1 or ERK-2 proteins also potentiate v-src-mediated luciferase expression driven by the pgs2 promoter, and expression of dominant-negative mutants of ERK-1, ERK-2, or Raf-1 attenuate this response. Thus, in response to v-src expression, a Ras/MEKK-1/JNK signal transduction pathway activating c-Jun and a Ras/Raf-1/ERK pathway converge to mediate pgs2 gene expression via the CRE site in the pgs2 promoter.