Fig. 8. Induction of MMP-9 promoter activity and gelatinolytic expression by synergistic co-operation of oncogenic RasVal12 and Snail. (A) MDCK-CMV cells were transiently co-transfected with the pMMP9-392luc reporter construct and the indicated combinations of oncogenic Ras (RasVal12), Snail, a dominant negative Ras (RasN17) or the pCDNA3 expression vector (Mock), as well as TK renilla. After transfection, the cells were treated for an additional 5 hours with UO126 inhibitor (10 µM) or DMSO, prior to determining luciferase activity. The data are expressed as the mean±s.d. of the relative normalised luciferase values from three independent experiments. (B) Schematic representation of the proximal 5' region examined in these experiments. The pMMP9-389luc (construct III) contains 389 bp from the initiation transcription site and the positions of potential regulatory control elements are indicated. (C) The effects of Snail and Ras oncogene were analysed in the conditioned media by zymography in gelatin-embedded SDS polyacrylamide gels. MDCK-Snail cells were transiently transfected with oncogenic Ras (RasV12), a dominant negative Ras (RasN17) or the empty plasmid and media were collected after 24 hours. Conditioned medium from MDCK-CMV cells was also included as control.