Identification of targetgenes of E1AF using two hybrid system

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E1AF is a member of the ets-oncogene family transcription factor, and E1AF was shown to bind to the promoter region of MMPs that induce transcription from multiple MMP genes.In the present study, we demonstrated that Ad type 12 E1A (Ad12E1A), a highly tumorgenic adenovirus product is able to suppress the invasion ability of a cancer cell line.12-12S cell lines were established by transfection with Ad12E1A expression plasmid into HSC3, a highly invasive oral squamous cell carcinoma cell line. Expression of E1AF mRNA decreased in 12-12S cells compared to that of the parental HSC3 cells. Reduced expression of MMPs in 12-12S cells was identified both in mRNA levels and in protein levels. Furthermore, in vitro invasion assay using Matrigel revealed that the invasion ability of 12-12S cells was suppressed by half compared to that of HSC3 cells.These results suggest that Ad12E1A downregulates E1AF which, in turn, restrains invasion ability of human cancer cells by way of downregulaton of MMP expression. This may suggest that a negative-feedback circuit exists between Ad 12 E1A and E1AF.We examined the effect of HGF on the E1AF and MMP gene expression in terms of the invasive potential of the HSC3. HGF stimulated expression of the E1AF gene. The levels of MMP-1, -3 and -9 mRNAs increased in cells treated with HGF and correlated with E1AF upregulation. In contrast, no obvious upregulation of MMP-1 and MMP-9 mRNA was observed in ASE1AFHSC3 cells that was transfected with antisense E1AF expression vector into parental HSC3 cells.These results suggest that HGF induces expression of the Ets-related E1AF transcription factor gene whose product in turn activates MMP genes and leads to oral cancer cell invasion.