Interplay between 14-3-3gamma and MDMX in regulating the p53 pathway

描述（由申请人提供）：本提案主要研究14-3-3？参与调节MDM2-MDMX-p53反馈通路以应对缺氧引起的细胞应激。缺氧在肿瘤发生中起着至关重要的作用，特别是在实体瘤的进展中，因为在快速生长的肿瘤中，由于血液供应不足，所有实体瘤的缺氧区都显示0%-8%的氧浓度。在这一病理阶段，肿瘤抑制因子p53被诱导和激活，通过诱导细胞凋亡和细胞生长停滞以及减缓血管生成来阻止肿瘤的生长。尽管p53在低氧条件下抑制肿瘤进展中的作用已经得到了很好的证实，但低氧诱发p53的确切机制仍然知之甚少。生化和遗传学研究表明，泛素介导的蛋白酶体转换和p53肿瘤抑制因子的活性受到两种称为MDM2和MDMX的癌蛋白的严格控制。这两种蛋白一起作为p53的负反馈调节因子，因为MDM2也是p53的转录靶点，而MDMX是MDM2的合作伙伴。因此，解开这个反馈回路对于激活p53至关重要，以防止细胞在各种应激反应中发生转化和肿瘤。然而，尚不清楚缺氧是否通过影响这一环诱导p53。我们最近的研究表明14-3-3？可能在缺氧p53通路中起作用。根据我们最近的工作以及其他人的研究，我假设缺氧可能会导致14-3-3？通过ATR-ChK1级联磷酸化MDMX，从而抑制MDMX活性并导致p53激活。因此，本提案将通过解决三个具体目标来检验这一假设：为了确定ATR和ChK1是否在调节14-3-3？-MDMX相互作用对缺氧的反应；2. 阐明缺氧诱导14-3-3？-MDMX相互作用和p53激活；3. 以确定是否损失14-3-3？在细胞培养和动物实验中减轻p53对缺氧的反应。这些研究的成功实施不仅将阐明p53在缺氧反应中被诱导的新机制，而且还将确定14-3-3？作为细胞生长控制和肿瘤发生的另一个重要角色。

DESCRIPTION (provided by applicant): This proposal focuses on studying the role of 14-3-3? in regulating the MDM2-MDMX-p53 feedback pathway in response to cellular stress caused by hypoxia. Hypoxia plays a crucial role in tumorigenesis and particularly in the progression of solid tumors, as all solid tumors display 0%-8% oxygen concentrations in their hypoxic regions due to insufficient blood supply in rapidly growing tumors. During this pathological period, the tumor suppressor p53 is induced and activated to prevent the growth of the tumors by inducing apoptosis and cell growth arrest as well as slowing down angiogenesis. Although the role of p53 in suppressing tumor progression in response to hypoxia has been well established, the precise mechanism underlying hypoxia-triggered induction of p53 remains poorly understood. Biochemical and genetic studies have demonstrated that the ubiquitin-mediated proteasomal turnover and activity of the p53 tumor suppressor are tightly controlled by two oncoproteins called MDM2 and MDMX. Both of the proteins work together as p53 negative feedback regulators, because MDM2 is also a transcriptional target for p53 and MDMX acts as an MDM2 partner. Thus, untying this feedback loop is essential for activating p53 in order to prevent cells from undergoing transformation and neoplasia in response to various stresses. However, it is still unclear whether hypoxia induces p53 by affecting this loop. Clues from our recent studies suggest that 14-3-3? may play a role in the hypoxia-p53 pathway. In light of our recent work as well as studies by others, I hypothesize that hypoxia may induce the association of 14-3-3? with MDMX that is phosphorylated by the ATR-ChK1 cascade, consequently suppressing MDMX activity and leading to p53 activation. Therefore, this proposal will examine this hypothesis by addressing three specific aims: 1. To determine if ATR and ChK1 play a role in regulating the 14-3-3?-MDMX interaction in response to hypoxia; 2. To elucidate the mechanisms underlying hypoxia-induced 14-3-3?-MDMX interaction and p53 activation; 3. To determine if loss of 14-3-3? alleviates p53 response to hypoxia in cell culture and in animals. Successful implementation of these proposed studies will not only elucidate a novel mechanism by which p53 is induced in response to hypoxia, but would also identify 14-3-3? as another important player in cell growth control and tumorigenesis.