RNA Binding Proteins in Cancer

该项目的广泛目标是了解RNA结合的机制 蛋白RBM3在转录后水平调控基因表达 癌细胞中的稳定性和转译。我们已经确定了该基因的过度表达 蛋白质导致正常细胞经历转化的表型，从而导致细胞 在免疫受损的小鼠体内形成肿瘤。另一方面，RBM3的敲除 表达导致有丝分裂灾难导致细胞死亡。RBM3与HUR和 HnRNP A1，并带有AU-RNA序列，以增强mRNA的稳定性和翻译 富含Au的转录本，如COX-2、VEGF和IL-8。此外，RBM3过表达 在Notch中增加哺乳动物雷帕霉素蛋白靶标的激活 依赖机制。我们还发现了受RBM3调控的microRNAs 过度表达，包括下调抑制其自身表达的基因。我们的 研究还表明，RBM3在翻译后水平上受到调控 磷酸化、泛素化和SUMO化。基于这些观察，我们 提出三个具体目标。在目标1中，我们将确定RNA 结合蛋白RBM3调节基因表达。在这里，我们将识别RNA 与RBM3相互作用的序列。此外，我们还将确定在 COX-2 3‘非编码区是RBM3介导的COX-2 mRNA稳定性所必需的 翻译。我们还将确定HUR和hnRNP-A1在这一过程中的作用。在目标2中， 我们将测定RBM3中经过磷酸化、泛素化和 通过诱变使之相加。我们还将确定突变体对 信使核糖核酸稳定性和翻译。在目标3中，我们建议通过以下方式确定机制 哪种RBM3可诱导肿瘤发生。我们将使用异种移植癌模型来 确定mTOR和Notch通路在RBM3介导的肿瘤发生中的作用。另外， MicroRNAs在肿瘤发生中的作用将被确定。完成这些工作 实验应该会让我们更好地理解RNA结合蛋白是如何 RBM3在正常上皮细胞中的功能，以及RBM3的表达是否发生变化 在肿瘤细胞中观察到的这一点与肿瘤的行为有关。

The broad goal of the project is to understand the mechanisms by which RNA binding protein RBM3 regulates gene expression at the posttranscriptional level of mRNA stability and translation in cancer cells. We have identified that overexpression of the protein causes a normal cell to undergo a transformed phenotype resulting in the cells forming tumors in immunocompromized mice. On the other hand, knockdown of RBM3 expression results in cell death due to mitotic catastrophe. RBM3 interacts with HuR and hnRNP A1, and with AU-RNA sequences to enhance mRNA stability and translation of AU-rich transcripts such as COX-2, VEGF and IL-8. In addition, RBM3 overexpression increases the activation of the mammalian target of rapamycin protein in a Notch dependent mechanism. We have also identified microRNAs regulated upon RBM3 overexpression, including downregulating one that inhibits its own expression. Our studies also suggest that RBM3 is regulated at the posttranslational levels of phosphorylation, ubiquitination and SUMOylation. Based on these observations, we propose three specific aims. In Aim 1, we will determine the mechanism by which RNA binding protein RBM3 regulates gene expression. Here, we will identify the RNA sequences that interact with RBM3. In addition, we will determine the sequenced in COX-2 3'UTR that are required for RBM3-mediated COX-2 mRNA stability and translation. We will also identify the role of HuR and hnRNP-A1 in the process. In Aim 2, we will determine the residues in RBM3 that undergo phosphorylation, ubiquitination and SUMOylation by mutagenesis. We will also determine the effect of the mutants on mRNA stability and translation. In Aim 3, we propose to determine the mechanisms by which RBM3 induces tumorigenesis. We will use an xenograft cancer model to determine the role of mTOR and Notch pathway in RBM3 mediated tumorigenesis. Also, the role of microRNAs in the tumorigenesis will be determined. Completion of these experiments should give us a better understanding of how the RNA binding protein RBM3 functions in normal epithelial cells, and whether changes in the RBM3 expression that is observed in tumor cells is responsible for tumor behavior.