Regulation of cell homeostasis by fbx4

描述（由申请人提供）：我的研究的长期目标是阐明细胞周期机制感知细胞外信号并调节正常细胞与肿瘤细胞的细胞周期进程的机制。这些信息将提供必要的框架，以阐明如何生长调节途径被颠覆的肿瘤。我们目前的研究集中在生长信号通路如何调节有丝分裂反应的D型细胞周期蛋白，更具体地说，这些途径如何调节正常细胞与癌细胞中活性的核细胞周期蛋白D1依赖性激酶的积累。细胞周期蛋白D1在多种人类癌症中的过度表达突出了阐明调节细胞周期蛋白D1活性的机制的重要性。虽然细胞周期蛋白D1过表达是一个结果的基因扩增和染色体易位在一个子集的癌症，减少细胞周期蛋白D1蛋白降解，这取决于其磷酸化的Thr 286，被认为是在大多数癌症的关键因素。由于缺乏关于指导细胞周期蛋白D1蛋白水解的E3泛素连接酶的性质的信息，我们在表征癌症中细胞周期蛋白D1过表达的机制方面的进展受到阻碍。我们最近鉴定了SCF（Fbx 4-（B crystallin）），其控制细胞周期蛋白D1的泛素化和降解。这一发现提供了重要的信息和工具，以评估机制，有助于正常组织发育过程中的细胞周期蛋白D1的积累和潜在的失调细胞周期蛋白D1在恶性转化过程中的调节。这种E3连接酶的鉴定以及我们最近的初步研究导致了一个总体假设，即SCF（Fbx 4-（B晶状体蛋白）连接酶通过Fbx 4和（B晶状体蛋白）对磷酸化细胞周期蛋白D1的协调识别，在维持细胞生长和组织维持中起关键作用。本基金中提出的实验将确定Fbx 4对体内组织稳态的贡献（目的1）。我们将利用fbx 4-/-细胞来最终确定Fbx 4在细胞周期蛋白D1和细胞增殖调节中的作用（目的2）。将通过表征在黑色素瘤中鉴定的突变Fbx 4等位基因来鉴定Fbx 4中底物识别所必需的残基。

DESCRIPTION (provided by applicant): The long-term objective of my research centers on elucidation of the mechanisms whereby extra-cellular signals are sensed by the cell cycle machinery and regulate cell cycle progression in normal versus tumor cells. This information will provide the necessary framework to elucidate how growth regulatory pathways are subverted in neoplasia. Our current studies focus on how growth-signaling pathways regulate the mitogenically responsive D-type cyclins and more specifically, how these pathways regulate accumulation of an active, nuclear cyclin D1-dependent kinase in normal versus cancerous cells. The noted overexpression of cyclin D1 in multiple human cancers highlights the importance of elucidating the mechanisms that regulate cyclin D1 activity. While cyclin D1 overexpression is a consequence of gene amplification and chromosome translocation in a subset of cancers, decreased cyclin D1 protein degradation, which depends on its phosphorylation on Thr286, is thought to be the key factor in a majority of cancers. Our progress in the characterization of this mechanism of cyclin D1 overexpression in cancer has been hindered by lack of information regarding the nature of the E3 ubiquitin ligase that directs cyclin D1 proteolysis. We have recently identified the SCF(Fbx4-(B crystallin) that controls cyclin D1 ubiquitination and degradation. This discovery has provided essential information and tools to assess the mechanisms that contribute to regulation of cyclin D1 accumulation during normal tissue development and the potential deregulation of cyclin D1 during malignant transformation. The identification of this E3 ligase as well as our recent preliminary studies lead to the overarching hypothesis that the SCF(Fbx4 -(B crystallin) ligase, via coordinated recognition of phospho- cyclin D1 by Fbx4 and (B crystallin, plays a critical role in the maintenance of cell growth and tissue maintenance. Experiments proposed in this grant will determine the contribution of Fbx4 to tissue homeostasis in vivo (Aim 1). We will utilize fbx4-/- cells to conclusively define the role of Fbx4 in the regulation of cyclin D1 and cell proliferation (Aim 2). The identification of residues in Fbx4 necessary for substrate recognition will be identified through characterization of mutant Fbx4 alleles identified in melanoma.