Lysyl Oxidase (LO) is the key enzyme that controls collagen and elastin maturation. LO is synthesized as a 50 kDa pro-enzyme, secreted into the extracellular environment and is then processed by proteolytic cleavage, via procollagen C-proteinases, to a functional 32 kDa enzyme and an 18 kDa pro-peptide. The LO enzyme is localized mainly extracellularly, although processed 32 kDa LO has been seen intracellularly. Importantly, LO was identified as the ras-recision gene (rrg), with ability to reduce ras-induced transformed phenotype. For example, ras-transformed NIH 3T3 cells, reverted by interferon beta/gamma, returned to their transformed phenotype upon inhibition of LO. Interestingly, ras negatively regulates LO expression. Thus, LO is down regulated in many naturally occurring and oncogene-induced tumors, and is induced concomitantly with stable phenotypic reversion, suggesting this reduction plays a role in the transformation process. Recently, we showed that LO suppresses ras-mediated activation of NF-KappaB in NIH 3T3 cells, primarily via the phosphatidylinositol-3-kinase (PI3K) pathway. Further, we have recently demonstrated that the aberrant activation of NFkappaB factors that typifies breast cancer can play a causal role in mammary tumorigenesis. Thus, here we propose to use cell culture and mouse models to test our central hypothesis that LO functions as a tumor suppressor of ras-mediated transformation via inhibiting NFkappaB activation induced by the PI3K/Akt pathway. Furthermore, we will elucidate the domains of LO mediating its ability to suppress ras-mediated transformation, and the mechanisms of oncogene-mediated reduction of LO levels. Specifically, we propose to: 1) Identify the region(s) of LO necessary for its anti-oncogenic activity in ras-transformed cells; 2) Elucidate the mechanism of LO-mediated inhibition of ras activation of PI3K; 3) Elucidate the mechanism of repression of LO gene transcription as a function of transformation; 4) Elucidate the effects of LO on mammary tumorigenesis by Ha-ras. Overall, these studies will further our understanding of the signaling mechanisms involved in the anti-oncogenic effects of LO in cancer, i.e., its ability to function as a novel tumor suppressor gene. Given that decreased expression or mutation of LO typifies many human cancers, including colon, prostate, and kidney, identification of the regulation of LO and of the domains mediating its action, may provide additional tools to modulate the amount and activity of LO and help define novel therapeutic drugs for the treatment of ras-induced cancers of multiple origins.
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