Cyclin D1 and mammary carcinoma

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细胞周期进程的机制。这一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，这些途径如何调节积累的一个活跃的，核细胞周期蛋白D1依赖性激酶。阐明调节细胞周期蛋白D1的核积累的机制的重要性是强调我们的示范，细胞的失败，以消除活跃的D1/CDK复合物从细胞核在S期的结果在细胞转化。细胞周期蛋白D1在细胞周期的G1期响应促有丝分裂刺激而在细胞核中积累。在S期，细胞周期蛋白D1通过GSK-3 β磷酸化单个苏氨酰残基Thr-286而靶向细胞质。我们已经确定了一种天然存在的细胞周期蛋白D1亚型，D1 b，缺乏细胞周期蛋白核输出所需的关键残基。我们的初步数据表明，这种异构体是专门在癌细胞中表达，并可能代表一个致癌的经典细胞周期蛋白D1异构体的变体。此外，在子宫内膜癌中也有报道，细胞周期蛋白D1的C-末端突变会破坏细胞周期蛋白D1的核输出。我们假设，细胞周期蛋白D1 b和突变核细胞周期蛋白D1亚型是致癌的典型细胞周期蛋白D1（D1 a）的过度表达直接导致肿瘤恶性的变体。为了验证这一假设，我们建议：1）确定乳腺癌中细胞周期蛋白D1 b过表达的频率; 2）确定细胞周期蛋白D1突变体的组成性核驱动乳腺癌的能力;和3）表征癌症特异性细胞周期蛋白D1突变体。为了实现这些目标，我们将利用宾夕法尼亚大学的资源来确定核细胞周期蛋白D1变体D1 b在原发性人类乳腺癌中的频率，并评估其预后价值。此外，我们将使用新建立的小鼠模型来评估核细胞周期蛋白D1亚型在体内的致癌性。最后，我们建议表征新发现的细胞周期蛋白D1突变体的能力进行积极的，CRM 1依赖的核出口。虽然野生型细胞周期蛋白D1的表达在体外或动物模型中并不明显致癌，但我的实验室已经证明，组成型细胞周期蛋白D1亚型具有强烈的致癌性。 癌症相关性。这项工作的结果将支持一个模型，其中组成型核细胞周期蛋白D1作为一个启动癌基因的功能和机制，调节其核保留将在癌变过程中的目标。与此一致，在我们的初步数据中，我们提供了一种新的，组成型核细胞周期蛋白D1亚型，这是专门在癌症中表达的证据。本文提出的工作将建立细胞周期蛋白D1核滞留和细胞周期蛋白D1作为一种致癌蛋白在乳腺癌中的功能之间的关系。

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. This information will provide the framework necessary 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. The importance of elucidating the mechanisms that regulate nuclear accumulation of cyclin D1 is emphasized by our demonstration that the failure of the cell to remove active D1/CDK complexes from the nucleus during S-phase results in cell transformation. Cyclin D1 accumulates in the nucleus during G1 phase of the cell cycle in response to mitogenic stimulation. During S-phase, cyclin D1 is targeted to the cytoplasm via phosphorylation of the at a single threonyl residue, Thr-286, by GSK-3beta. We have identified a naturally occurring cyclin D1 isoform, D1b, which lacks critical residues necessary for cyclin nuclear export. Our preliminary data indicate that this isoform is specifically expressed in cancer cells and is likely to represent an oncogenic variant of the canonical cyclin D1 isoform. In addition, mutations in the C-terminus of cyclin D1 that will disrupt cyclin D1 nuclear export have been reported in endometrial cancer. We hypothesize that cyclin D1b and mutant nuclear cyclin D1 isoforms are oncogenic variants of canonical cyclin D1 (D1a) whose overexpression contributes directly to neoplastic malignancy. To test this hypothesis, we propose to: 1) Determine the frequency of cyclin D1b overexpression in breast carcinoma; 2) Determine the capacity of cyclin D1 mutants that are constitutively nuclear to drive mammary carcinoma; and 3) Characterize cancer specific cyclin D1 mutants. To accomplish these goals, we will utilize resources here at the University of Pennsylvania to determine the frequency of the nuclear cyclin D1 variant, D1b, in primary human breast cancer and assess its prognostic value. In addition, we will use newly established mouse models to assess the oncogenicity of nuclear cyclin D1 isoforms in vivo. Finally, we propose to characterize newly identified cyclin D1 mutants for their ability to undergo active, CRM1-dependent nuclear export. While expression of wild-type cyclin D1 is not overtly oncogenic in vitro or in animal models, my laboratory has demonstrated that constitutively nuclear cyclin D1 isoforms are strongly oncogenic. Cancer Relevance. Findings from this work would support a model wherein constitutively nuclear cyclin D1 functions as an initiating oncogene and that mechanisms, which regulate its nuclear retention will be targeted during carcinogenesis. Consistent with this, in our preliminary data we provide evidence for a novel, constitutively nuclear cyclin D1 isoform, which is expressed exclusively in cancer. The work proposed herein will establish the relationship between cyclin D1 nuclear retention and the function of cyclin D1 as an oncogenic protein in mammary carcinoma.