Lysyl oxidase (LO) functions as a suppressor of the tumorigenicity of the ras oncogene. In NIH 3T3 cells transformed by multiple copies of LTR-C-H-ras (RS485), the transcription of LO is markedly decreased. Long term treatment with interferon (INF) beta created revertants that still overexpressed ras but had restored LO expression. Transfection of persistent revertant cells with antisense lysly oxidase expression constructs led to retransformation and loss of LO expression. Because the ras oncogene is involved in several human cancers, it might be possible to prevent or treat such cancers by maintaining or restoring LO expression. Although LO is known as a secreted enzyme involved in extracellular collagen maturation, the gene is expressed in normal epithelial cells of breast, prostate, and colon. In human tumors derived from breast and prostate epithelium, LO expression is reduced or lacking. The mechanism(s) by which ras transformation down regulates LO expression will be studied. Preliminary studies of bisulfite modified genomic DNA suggest that the CpG island in the LO promoter is differentially methylated between NIH 3T3 and TS485. Methylation patterns in the LO promoter of NIH 3T3, RS485, and persistent revertant cells will be determined and compared to elucidate the possible contribution of methylation to the down regulation of LO expression. The mechanism by which restoration of LO expression suppressed the tumorigenic ras phenotype will also be investigated. In addition to its extracellular function in the maturation of collagen and elastin, LO was recently shown to be present and active in nuclei, suggesting that LO does have an intracellular function. LO deletion mutants will be used to determine which protein domains contribute to the functionality of reversion of FS485. Studies with antibody to IRF1 showed that in RS485 cells there was a protein smaller than IRF-1 that also bound IRF-1 antibody. The relationship of these two proteins and the contribution of the smaller species to LO transcription will be investigated. Treatment of RS485 cells with a combination of IFN beta and retinoic acid gave rise to a high percentage of revertants that had lost all of the multiple copies of the transforming LTR-c-H-ras oncogence. The mechanism involved in this deletion will be investigated as it might be useful for the treatment of HTLV or HIV induced diseases, which involve LTR-linked viruses.
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