Utilizing alternative reading frames, the mammalian ARF-INK4a locus encodes two unrelated proteins that both function in tumor suppression, ARF and the cell cycle inhibitor, p16. ARF binds with MDM2 and blocks MDM2-directed ubiquitination of p53 and p53 nuclear export, thus preventing cytoplasmic degradation of p53. Previously, we, and others, found that human ARF binds the MDM2 oncoprotein leading to stabilization and transcriptional activation of p53 with a resultant proliferative arrest. ARF levels are increased through transcription following exposure to oncogenic stimuli, but post-translational regulation of ARF is largely unexplored. Recently, we have found a novel protein, LZAP, (leucine zipper containing ARF-binding protein) that binds ARF in the nucleus and regulates ARF biochemical activity toward MDM2. In addition to its regulation of MDM2 activity, ARF has recently been found to have p53-independent effects including S-phase growth delay, regulation of rRNA processing and inhibition of the nuclear factor kappa B (NF-kB), that may contribute to p53-independent effects of ARF observed in tumorigenesis. In addition to its regulation of ARF, we initially explored the role of LZAP in regulation of NF-kB activity and found that expression of LZAP decreased both basal and cytokine stimulated NF-kB activity, while inhibition of endogenous LZAP expression resulted in increased basal NF-kB activity. Remarkably, LZAP was found to bind directly to NFkB within cells. We have identified a novel protein that regulates ARF activity toward MDM2 and decreases NF-kB activity. Given the importance of ARF and NF-kB in tumorigenesis, the biochemical activities of LZAP suggest that it may serve as a tumor suppressor. We will further evaluate the role of LZAP in regulation of ARF and NF-kB activity and determine if LZAP has tumor suppressor activity.
