Plk3, A New Player In The Hypoxia Regulatory Networ.

描述(申请人提供)：HIF-1控制许多基因的转录，这些基因涉及癌症生物学的关键方面。HIF-11是HIF-1在低氧条件下的一个可诱导亚单位，其过度表达在许多癌症中是一个预后因素。PI3K、PDK1和Akt(又称蛋白激酶B)介导的主要信号轴在HIF-11的表达调控中起重要作用。众所周知，GSK32和MAPK都能直接磷酸化HIF-11，从而影响其稳定性和/或核定位。为了了解抑制肿瘤发展和肿瘤血管生成的调控网络，我们最近发现HIF-11是Polo-like kinase3(Plk3)的一种新的体外底物。Plk3在体外强烈磷酸化丝氨酸576和657上的HIF-11，这两个残基分别位于氧依赖降解区域和核输出信号附近。通过对原代等基因小鼠胚胎成纤维细胞(MEF)的研究，我们发现PLK3-/-MEF对缺氧或镍(一种缺氧模拟物)诱导的HIF-11高度敏感。与野生型MEF相比，PLK3-/-MEF具有高水平的Akt1/PKB活性，这与GSK32的抑制磷酸化密切相关。与Plk3在调节缺氧信号网络中的潜在作用一致，PLK3-/-小鼠在高龄时会发生各种器官的肿瘤，并且PLK3-/-肿瘤体积大，血管丰富，表明肿瘤血管生成活跃。根据这些关于Plk3和HIF-11之间的物理和功能相互作用的观察，我们假设Plk3负调控缺氧调节网络和HIF-11依赖的肿瘤血管生成。为了检验这一假设的有效性，我们将(I)研究Plk3与已知信号分子之间的功能相互作用，包括磷酸化HIF-11的GSK32和MAPKs，并确定Akt1上游的额外Plk3靶点(S)，(Ii)确定plk3-/-小鼠是否在缺氧条件下容易发生肿瘤，以及(Iii)研究携带Plk3耐磷酸化HIF-11突变等位基因的小鼠在镍暴露后是否更容易发生肿瘤。体外和体内的联合研究将极大地促进阐明Plk3在缺氧反应或暴露于环境致癌物(如镍化合物)后调节HIF-11的新机制。对HIF-11分子调控的详细了解将大大增加对肿瘤血管生成和肿瘤细胞对抗癌治疗的耐药性的现有知识。 与公共健康相关：类马球蛋白的功能调节细胞生长和细胞死亡的各个方面。我们最近的生化和小鼠遗传学研究表明，Plk3在调节HIF-11信号通路以及抑制肿瘤发生方面发挥着重要作用。本项目的主要目的是阐明Plk3在低氧应激反应中的信号转导途径，以及遗传毒性金属毒物在低氧时如何篡改调节HIF-11的细胞机制。

DESCRIPTION (provided by applicant): HIF-1 controls the transcription of many genes that are involved in key aspects of cancer biology. Overexpression of HIF-11, an inducible subunit of HIF-1 in response to hypoxia, is a prognostic factor in many cancers. The major signaling axis mediated by PI3K, PDK1, and Akt (also referred as protein kinase B) plays a significant role in the regulation of HIF-11 expression. Both GSK32 and MAPKs are also known to directly phosphorylate HIF-11, thereby affecting its stability and/or nuclear localization. To understand the regulatory network that suppresses tumor development and tumor angiogenesis, we have recently identified HIF-11 as a new in vitro substrate of Polo-like kinase 3 (Plk3). Plk3 strongly phosphorylates HIF-11 on serines 576 and 657 in vitro, two residues that lie in the oxygen-dependent degradation domain and near the nuclear export signal, respectively. By studying primary isogenic murine embryonic fibroblasts (MEFs), we have shown that PLK3-/- MEFs are hyper-sensitive to the induction of HIF-11 under hypoxia or treated with nickel, a hypoxia mimetic. Compared with that of wild- type MEFs, PLK3-/- MEFs contain a high level of Akt1/PKB activities, which is tightly associated with the inhibitory phosphorylation of GSK32. Consistent with the potential role of Plk3 in regulating the hypoxia signaling network, PLK3-/- mice develop tumors in various organs at an advanced age and PLK3-/- tumors are large in size and highly vascularized, suggesting active tumor angiogenesis. On the basis of these observations regarding physical and functional interactions between Plk3 and HIF-11, we hypothesize that Plk3 negatively regulates the hypoxia regulatory network and HIF-11-dependent tumor angiogenesis. To test the validity of this hypothesis, we will (i) study functional interaction between Plk3 and known signaling molecules, including GSK32 and MAPKs that phosphorylate HIF-11, and identify additional Plk3 target(s) upstream of Akt1, (ii) determine whether PLK3-/- mice are prone to tumorigenesis under hypoxia, and (iii) investigate whether mice harboring Plk3 phosphorylation-resistant mutant alleles of HIF-11 are more susceptible to tumorigenesis after nickel exposure. The combined in vitro and in vivo studies will greatly facilitate the elucidation of a new mechanism by which HIF-11 is regulated by Plk3 during hypoxic responses or after exposure to environmental carcinogens such as nickel compounds. A detailed understanding of the molecular regulation of HIF-11 will add significantly to the existing knowledge of tumor angiogenesis and tumor cell resistance to anti-cancer therapies. PUBLIC HEALTH RELEVANCE: Polo-like kinases function to regulate various aspects of cell growth and cell death. Our recent biochemical and mouse genetic studies show that Plk3 plays an important role in the regulation of the HIF- 11 signaling pathway, as well as in suppressing tumorigenesis. The major goal of this project is to elucidate what signaling pathways mediate Plk3 function in hypoxic stress responses and how genotoxic metal toxicants usurp cellular mechanisms that regulate HIF-11 during hypoxia.