Role of the AMPK-MDMX-p53 pathway in cancer

 描述(由申请人提供)：在过去的十年中，新陈代谢对癌症发展和进展的重要性已经得到越来越多的认识。癌细胞经历了剧烈的代谢变化，包括葡萄糖和脂肪的新陈代谢，以及氧化反应。这些过程受到AMPK和P53的高度调控。AMPK被LKB1和AMP/ADP激活，以响应生物能量短缺，通过磷酸化其靶蛋白来协调代谢、细胞生长、细胞凋亡和自噬。此外，肿瘤抑制基因P53可以被这种类型的应激激活，以限制有利于肿瘤生长的代谢途径，并通过诱导各种对这些功能重要的基因来启动导致细胞生长停滞和凋亡的细胞途径。P53的抗肿瘤功能受其两个生理拮抗剂MDM2和MDMX的严格调控，这两个拮抗剂形成一个复合体，作为P53的负反馈调节因子。因此，阻断这种反馈调节是细胞应激激活P53的关键。例如，核糖体应激通过诱导核糖体蛋白L11和L5与MDM2的相互作用并抑制后者对P53的活性而导致P53的激活，但在MDM2C305F小鼠中这种相互作用的缺陷损害了这一途径。有趣的是，AMPK可以通过磷酸化Ser342上的MDMX，通过诱导其与14-3-3结合而导致MDMX失活，从而激活P53，从而在代谢应激时作为P53的上游调节因子。因此，我们的研究表明，MDMX可能在代谢压力下发挥重要作用。此外，我们对MDM2C305F和MDMX3SA双重敲入小鼠的初步研究表明，MDM2-MDMX-P53通路的双重损伤导致禁食后肝脏中脂肪和葡萄糖代谢的显着缺陷。因此，我们推测代谢应激-AMPK-MDMX/RS-MDM2-P53通路可能在正常细胞和癌细胞对代谢应激的代谢调节中发挥重要作用。我们将通过解决三个具体目标来检验这一假设。1.阐明AMPK在代谢应激反应中抑制MDMX对P53活性的作用机制。A)AMPK对S342的磷酸化是否导致MDMX的核定位和降解？B)AMPK对S342的磷酸化是否影响MDMX-MDM2复合体的形成？2.确定AMPK-MDMX/RS-MDM2-P53信号通路是否参与细胞和小鼠的代谢调控。A.确定在生理和应激条件下，MDMX-MDM2-P53通路的双重缺陷是否会损害能量代谢。B.通过AMPK-MDMX/RS-MDM2-P53信号通路阐明新的P53应答基因对调节糖脂代谢具有重要意义。3.确定AMPK-MDMX/RS-MDM2-P53通路的双重缺陷是否加速了两种小鼠肿瘤模型的发生。完成这些研究将提供分子方面的洞察如何控制脂肪和葡萄糖代谢的这一途径，也揭示了新的分子靶点，为未来的抗癌药物发现。

 DESCRIPTION (provided by applicant): The importance of metabolism for cancer development and progression has been increasingly recognized over the past decade. Cancer cells undergo drastic alternations of metabolism, including metabolism of glucose and lipid, and oxidative responses. These processes are highly regulated by AMPK and p53. AMPK is activated by LKB1 and AMP/ADP in response to bio-energy shortage to coordinate metabolism, cell growth, apoptosis and autophagy by phosphorylating its target proteins. Also, the tumor suppressor p53 can be activated by this type of stress to confine metabolic pathways, which favor tumor growth, and to turn on cellular pathways that lead to cell growth arrest and apoptosis by inducing a variety of genes important for these functions. The anti-tumor function of p53 is tightly regulated by its two physiological antagonists MDM2 and MDMX, which form a complex to act as a negative feedback regulator of p53. Hence, blocking this feedback regulation is the key for cellular stresses to activate p53. For instance, ribosomal stress (RS) leads to p53 activation by inducing the interaction of ribosomal proteins L11 and L5 with MDM2 and restraining the latter's activity toward p53, but defect of this interaction in MDM2C305F mice impairs this pathway. Interestingly, AMPK can act as an upstream regulator of p53 upon metabolic stress by phosphorylating MDMX at Ser342, leading to inactivation of MDMX by inducing its binding to 14-3-3, consequently activating p53. Hence, our studies suggest that MDMX may function as a crucial player under metabolic stress. Also our preliminary studies using MDM2C305F and MDMX3SA double knock-in mice showed that the double impairments of the MDM2-MDMX-p53 pathways cause significant defects of lipid and glucose metabolisms in livers upon fasting. Thus, we hypothesize that the metabolic stress-AMPK-MDMX/RS-MDM2-p53 pathways may play an important role in regulation of metabolism in both normal and cancer cells upon metabolic stress. We will test this hypothesis by addressing three specific aims. 1. To elucidate mechanisms underlying the AMPK inhibition of MDMX activity toward p53 in response to metabolic stress. A) Does S342 phosphorylation by AMPK induce nuclear localization and degradation of MDMX? B) Does S342 phosphorylation by AMPK affect the formation of the MDMX-MDM2 complex? C) Does S342 phosphorylation by AMPK induce 14-3-3-binding to S367 of MDMX? 2. To determine if the AMPK-MDMX/RS-MDM2-p53 pathways play a role in regulation of metabolism in cells and mice. A. To determine if dual defects of the MDMX-MDM2-p53 pathway impair energy metabolism under physiological and stress conditions. B. To illustrate new p53 responsive genes important for regulation of lipid and glucose metabolism by the AMPK-MDMX/RS-MDM2-p53 pathways. 3. To determine if dual defects of the AMPK-MDMX/RS-MDM2-p53 pathways accelerate tumorigenesis in two mouse tumor model systems. Completing these studies would offer molecular insight into how lipid and glucose metabolism is controlled by this pathway, and also reveal new molecule targets for future anti-cancer drug discovery.