Translational control of EMT by the CELF1 RNA Binding Protein

 描述（由申请人提供）：根据一些估计，肿瘤转移是超过90%的癌症死亡率的原因。在细胞水平上，上皮间质转化（EMT） 是肿瘤转移的起始事件。我们一直在乳腺上皮模型中研究翻译调控对TGF-β诱导EMT的作用.我们的初步数据明确地表明，CELF 1 RNA结合蛋白，最有名的是其在1型肌强直性营养不良症中的作用，是必要的和足够的EMT在这个系统中。在上皮细胞中，CELF 1蛋白通过蛋白酶体主动降解。然而，在加入TGF-β 1和EMT后，这种蛋白质被稳定化，并且似乎增加了几种靶基因的翻译，其中一些已经被定义为EMT的正调节因子。据我们所知，这是第一个数据直接证明EMT中的基因产物的作用。我们的总体目标是进一步阐明TGF-β通过CELF 1蛋白诱导EMT的机制。我们预测，对该途径中起作用的细胞机制的更深入理解将揭示在EMT中起关键作用的酶活性，并且这些活性的最终治疗靶向可能是预防早期癌症转移的可行治疗靶点。我们希望通过测试我们的核心假设来实现我们的目标，即CELF 1蛋白的诱导稳定性通过不同下游效应蛋白的翻译激活来驱动EMT。为此，我们提出了三个具体目标。首先，我们将通过候选方法在实验模型中鉴定CELF 1促进EMT和肿瘤转移的下游效应物。使用经典的生物化学方法，我们接下来将建立TGF-β治疗导致CELF 1蛋白水平增加的机制。最后，我们将确定CELF 1的错误表达在多大程度上影响体内模型中的肿瘤定植和转移，并调查广泛的人类乳腺癌，以确定与正常组织相比，CELF 1是否在这些肿瘤中错误表达。这项工作与癌症直接相关，因为EMT是肿瘤转移的核心机制。在EMT背景下控制CELF 1稳定性和功能的酶活性的鉴定将揭示EMT的候选治疗靶点。 预防早期癌症的转移。CELF 1蛋白的下游效应物也可能被证明是潜在的治疗靶点，但肯定会提供对EMT潜在细胞机制的深入了解，EMT可能会转化为广泛的实体瘤。

 DESCRIPTION (provided by applicant): By some estimates, tumor metastasis is responsible for over 90% of cancer mortality. At the cellular level, epithelial to mesenchymal transition (EMT) is the initiating event in tumor metastasis. We have been investigating the contribution of translational control to TGF-ß-induced EMT in a breast epithelial model. Our preliminary data unambiguously demonstrate that the CELF1 RNA binding protein, most well-known for its role in Type 1 Myotonic Dystrophy, is both necessary and sufficient for EMT in this system. In epithelial cells, the CELF1 protein is actively degraded via the proteasome. However, upon TGF-ß addition and EMT, this protein is stabilized and appears to increase the translation of several target genes, some of which have already been defined as positive regulators of EMT. To our knowledge this is the first data directly demonstrating the role of this gene product in EMT. Our overall objective is to further elucidate the mechanism by which TGF-ß induces EMT via the CELF1 protein. We predict that a more developed understanding of the cellular mechanisms at play in this pathway will reveal enzymatic activities playing critical roles in EMT, and that eventual therapeutic targeting of these activities may be viable therapeutic targets for the prevention of metastasis in early stage cancers. We hope to achieve our objective by testing our core hypothesis that induced stability of CELF1 protein drives EMT via translational activation of distinct downstream effector proteins. We propose doing this via three Specific Aims. First, we will identify the downstream effectors by which CELF1 promotes EMT and tumor metastasis in experimental models via a candidate approach. Using classical biochemical approaches, we will next establish the mechanism by which TGF-ß treatment leads to an increase in CELF1 protein levels. Finally, we will determine to what extent misexpression of CELF1 impacts tumor colonization and metastasis in in vivo models, and survey a broad range of human breast cancers to determine whether CELF1 is misexpressed in these tumors as compared to normal tissue. The proposed work is directly relevant to cancer because EMT is a core mechanism underlying tumor metastasis. Identification of the enzymatic activities by which CELF1 stability and function are controlled in the context of EMT will reveal candidate therapeutic targets for the prevention of metastasis of early-stage cancers. Downstream effectors of the CELF1 protein may also prove to be potential therapeutic targets, but are certain to provide insight into the cellular mechanisms underlying EMT that may be translatable to a broad range of solid tumors.