Differential display of mRNA has been used to identify genes whose expression may drive or prevent progression to tumor cell phenotype. One such gene, tissue inhibitor of metalloproteinases TIMP-3), was expressed in preneoplastic but not neoplastic JB6 cells (Sun et al., Cancer Res, 1994; Sun et al., J Biol Chem, 1995) and was found to act as a tumor suppressor when expressed in human DLD colon carcinoma cells (Bian et al., Carcinogenesis, 1996). The transcriptional silencing of TIMP-3 is attributable to sequence specific methylation of its promoter (Pennie et al., Cell Growth and Diff, 1999). Expression of antisense DNA methyl transferase restored expression of TIMP-3 to tumor cells and recapitulated the unmethylated status of the same sites associated with TIMP-3 expression in preneoplastic JB6 cells. A separate differential display analysis was carried out to identify gene expression that may mediate or inhibit tumor promoter dependent stages of progression. Full-length cDNA clones have been isolated for two genes preferentially expressed in promotion resistant mouse JB6 cells. One designated pdcd4 (Cmarik et al., PNAS 1999, is novel; the other is plekstrin (Cmarik et al Genomics 2000). Antisense expression of the novel pdcd4 gene converts P- to P+ cells and pdcd4 sense expression (Yang et al Oncogene 2001) converts P+ to P- cells, thus establishing a causal relationship to prevention of tumor promoter induced transformation. Furthermore pdcd4 expression suppresses tumor phenotype in transformed JB6 cells (Yang et al Oncogene 2003). Current studies focus on the molecular function and localization of Pdcd4 protein. Examination of the possible inhibitory effect of pdcd4 on molecular events known to be required for tumor promotion revealed that pdcd4 expression inhibited the activation of transcription factor AP-1 but not of NFkappa B or of ornithine decarboxylase(Yang et al Oncogene 2001). The AP-1 inhibiting activity of Pdcd4 appears to be attributable to blocking the transactivation of cJun and cFos (Yang et al Oncogene 2003). Although expression of Pdcd4 protein blocks AP-1 activation, Pdcd4 does not interact directly with Jun or Fos proteins. Analysis of Pdcd4 binding partners by a yeast two-hybrid assay revealed the translation initiation factor eIF4A as a major binding partner (Yang et al Molec Cell Biol 2003). Binding of Pdcd4 to eIF4A is required for Pdcd4 to inhibit eIF4a's RNA helicase, to inhibit translation, and to inhibit the activation of AP-1 dependent transcription required for neoplastic transformation. Current research is focused on identifying specific transformation relevant mRNAs whose translation may be inhibited by Pdcd4 expression. Finally, we have discovered that Pdcd4 expression in human cancer cell lines is predictive for sensitivity to tamoxifen and geldanamycin. Moreover, expression of Pdcd4 actually confers sensitivity to these drugs (Jansen et al Molec Cancer Ther 2003 in press).
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