The role of FBXW2 as a novel lung tumor suppressor that cross-talks with oncogenic beta-TrCP and SKP2

产品描述（申请人提供）：SCF（SKP 1-Cullins-F box protein）E3泛素连接酶是E3连接酶中最大的家族，可促进多种关键调控蛋白的泛素化和降解，从而控制许多重要的生物学过程，如凋亡、细胞周期进程、DNA复制、胚胎发生和肿瘤发生。SCF由4个组分组成，包括支架蛋白cullin-1、接头蛋白SKP 1、RING蛋白RBX 1或RBX 2和F-box蛋白。在哺乳动物细胞中具有69个成员的F-box蛋白是决定SCF E3的特异性的底物识别亚基。虽然各种蛋白质底物已被鉴定的一个良好表征的F-盒蛋白β TrCP，FBXW 7，或SKP 2，非常少的是已知的F-盒蛋白是否和如何相互调节。我们的初步研究结果表明：1）致癌F-box蛋白FBXW 2与致癌F-box蛋白FBXW 2结合，促进其泛素化和降解; 2）FBXW 2与致癌F-box蛋白SKP 2结合，促进其泛素化和降解; 3）FBXW 2对肺癌细胞具有肿瘤抑制作用; 4）在人非小细胞肺癌组织中发现了大量功能获得性FBXW 2突变; 5）β-TrCP、FBXW 2和SKP 2在肺癌组织中的表达存在反向相关性，与患者的生存相关。虽然β-TrCP和SKP 2是具有致癌活性的F-box蛋白，但它们是否以及如何相互调节细胞增殖和存活是完全未知的。这项拟议研究的目的是了解FBXW 2如何介导和调节两种众所周知的致癌F-box蛋白β-TrCP和SKP 2的活性，以及FBXW 2如何在体外细胞培养和体内遗传修饰小鼠模型中作为一种新型肿瘤抑制因子在肺肿瘤发生中发挥作用，并阐明其作用机制。中心假设是三种F盒蛋白β TrCP-FBXW 2-SKP 2形成癌基因-肿瘤抑制因子-癌基因轴，其相互调节以控制细胞增殖和肺肿瘤发生。提出了三个具体目标来验证我们的中心假设：1）将FBXW 2表征为β-TRCP的新型底物; 2）将FBXW 2表征为SKP 2降解的新型E3连接酶; 3）将Fbxw 2表征为针对肺肿瘤发生的新型肿瘤抑制剂。相关性：这项拟议研究的成功完成将从机制上阐明三种F-box蛋白如何通过相互靶向降解来调节肺癌细胞的增殖，以及两种致癌F-box蛋白FBXW 2-TrCP和SKP 2如何通过肿瘤抑制性F-box蛋白FBXW 2相互交谈，以及FBXW 2如何作为肺中的肿瘤抑制因子，从而阐明了肺肿瘤发生的新机制，并为肺癌管理中的未来生物标志物开发提供了有吸引力的靶点。

DESCRIPTION (provided by applicant): SCF (SKP1-Cullins-F box protein) E3 ubiquitin ligase is the largest family of E3 ligases that promote the ubiquitylation and degradation of various key regulatory proteins, thus controlling many important biological processes, such as apoptosis, cell cycle progression, DNA replication, embryogenesis and tumorigenesis. SCF consists of 4 components, including a scaffold protein cullin-1, an adaptor protein SKP1, a RING protein RBX1 or RBX2 and an F-box protein. The F-box protein with 69 members in mammalian cells is the substrate- recognizing subunit that determines the specificity of SCF E3s. While a variety of protein substrates have been identified by one of well-characterized F-box proteins ßTrCP, FBXW7, or SKP2, very little is known as to whether and how F-box proteins regulate each other. Our preliminary data showed that 1) oncogenic F-box protein TrCP binds to yet a poorly characterized F-box protein FBXW2 to promote its ubiquitylation and degradation; 2) FBXW2 in turn binds to oncogenic F-box protein SKP2 to promote its ubiquitylation and degradation; 3) FBXW2 has tumor suppressive function against lung cancer cells; 4) A number of gain-of- function FBXW2 mutations were found in human non-small cell lung carcinoma tissues; and 5) reversed correlation in expression among β-TrCP, FBXW2 and SKP2 exists in lung cancer tissues, which is associated with patient survival. Although β-TrCP and SKP2 are well-studied F-box proteins with oncogenic activity, whether and how they regulate each other with regard to cell proliferation and survival is totally unknown. The objectives of this proposed study is to understand how FBXW2 mediates and modulates the activity of two well-known oncogenic F-box proteins, β-TrCP and SKP2, and how FBXW2 acts as a novel tumor suppressor in lung tumorigenesis using both in vitro cell culture and in vivo genetically modified mouse models with elucidation of its mechanism of action. The central hypothesis is that three F- box proteins, βTrCP-FBXW2-SKP2 form an oncogene-tumor suppressor-oncogene axis that regulates each other to control cell proliferation and lung tumorigenesis. Three specific aims are proposed to test our central hypothesis by 1) characterizing FBXW2 as a novel substrate of β-TRCP; 2) characterizing FBXW2 as a novel E3 ligase for SKP2 degradation; and 3) characterizing Fbxw2 as a novel tumor suppressor against lung tumorigenesis. Relevance: Successful completion of this proposed study will mechanistically elucidate how three F-box proteins regulate proliferation of lung cancer cells by targeting each other for degradation, and how two oncogenic F-box proteins ß-TrCP and SKP2 talk with each other via a tumor suppressive F-box protein FBXW2, and how FBXW2 acts as a tumor suppressor in the lung, thus elucidating a novel mechanism of lung tumorigenesis and providing attractive targets for future biomarker development in lung cancer management.