NOVEL PROTEIN REGULATOR OF TUMOR SUPPRESSOR ARF & NF-kB

描述(申请人提供)：哺乳动物的ARF-INK4a基因座利用不同的阅读框架，编码两种在肿瘤抑制中都起作用的无关蛋白质：ARF和细胞周期抑制因子p16。ARF与MDM2结合，阻断MDM2诱导的P53泛素化和P53核输出，从而阻止P53的胞质降解。在此之前，我们和其他人发现，人类ARF与MDM2癌蛋白结合，导致p53的稳定和转录激活，从而导致增殖停止。ARF水平在暴露于致癌刺激后通过转录增加，但ARF的翻译后调控在很大程度上尚未被探索。最近，我们发现了一种新的蛋白质LZAP(亮氨酸拉链包含ARF结合蛋白)，它能在细胞核内与ARF结合，并调节ARF对MDM2的生化活性。除了对MDM2活性的调节外，最近发现ARF还具有非p53依赖的作用，包括S期生长延迟、调节rRNA加工和抑制核因子kappaB(NF-kB)，这可能与ARF在肿瘤发生中观察到的P53非依赖性作用有关。除了对ARF的调节外，我们还初步探讨了LZAP在调节NF-kB活性中的作用，发现LZAP的表达降低了基础和细胞因子刺激的NF-kB的活性，而抑制内源性LZAP的表达会导致基础的NF-kB活性增加。值得注意的是，LZAP被发现在细胞内直接与NFkB结合。我们已经确定了一种新的蛋白质，它调节ARF对MDM2的活性并降低NF-kB的活性。鉴于ARF和NF-kB在肿瘤发生中的重要性，LZAP的生化活性表明它可能是一种肿瘤抑制因子。我们将进一步评估LZAP在调节ARF和NF-kB活性中的作用，并确定LZAP是否具有肿瘤抑制活性。

DESCRIPTION (provided by applicant): 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.