Regulation of cell growth by RNA binding proteins

描述（由申请人提供）：肝肿瘤的发生涉及许多基因表达的改变。阐明调节正常细胞和癌细胞中基因表达的分子机制是开发肿瘤治疗性治疗的重要步骤。从mRNA翻译蛋白质是基因表达的关键事件之一。我的实验室研究RNA CUG三联体重复结合蛋白CUGBP 1的生物学活性，以及这种蛋白在肝脏增殖和肝脏肿瘤中的作用。在肝脏中，CUGBP 1调节几种蛋白质的翻译，包括C/EBP家族的成员C/EBP？，这是肝脏增殖的关键调节因子。CUGBP 1与C/EBP <$mRNA的5'区结合，并增加显性负性分子C/EBP <$-LIP的翻译，其抑制全长C/EBP蛋白的生物活性并促进肝脏增殖。我们最近在肝脏中发现了CUGBP 1的新靶点。我们获得的证据表明，CUGBP 1结合到编码组蛋白去乙酰化酶1，HDAC 1的mRNA的5'区域，并增加肿瘤肝脏中HDAC 1蛋白的翻译。我们发现CUGBP 1的翻译活性主要受CUGBP 1磷酸化水平的控制。在肿瘤肝脏和部分肝切除术（PH）后增殖的肝脏中，CUGBP 1被过度磷酸化，这种磷酸化增加了CUGBP 1的翻译活性。我们已经发现cdk 4是一种磷酸化CUGBP 1并增加其翻译活性的激酶。因此，本申请的主要假设是肝脏中CDK 4介导的CUGBP 1活化导致C/EBP-LIP和HDAC 1翻译增加并导致肿瘤发展（见左图）。与这一假设相一致，细胞周期蛋白D1和细胞周期蛋白D3，这是cdk 4的激活剂，在人肝肿瘤的细胞质提取物和部分肝切除术后增殖的小鼠肝的细胞质中的水平增加。特异性目的1检查增殖的肝脏增加细胞质中cdk 4-D型细胞周期蛋白复合物的量的机制。具体目的2和3确定CUGBP 1-C/EBP-LIP和CUGBP 1-HDAC 1途径在肝增殖中的作用。我们将检验CUGBP 1介导的HDAC 1升高抑制肝脏特异性肿瘤抑制基因（如C/EBPa）启动子的假设。在本申请中，我们还将确定肝脏在某些条件下（如急性期反应）阻断C/EBP <$-LIP促生长活性的机制。了解C/EBP <$-LIP和HDAC 1在肿瘤中升高的机制将有助于开发预防肝脏恶性转化的方法。公共卫生相关性：本项目研究RNA结合蛋白CUGBP 1在肝肿瘤发生和部分肝切除术（PH）后肝脏增殖中的作用。本申请的目的是1）检查RNA结合蛋白在肝肿瘤发展中的作用; 2）阐明增殖的肝脏激活RNA结合蛋白的机制;和3）确定增殖的肝脏中RNA结合蛋白CUGBP 1的下游靶点。阐明肝细胞恶性转化的分子机制将为肿瘤的防治提供基础。

DESCRIPTION (provided by applicant): Development of liver tumors involves alterations in expression of many genes. The elucidation of molecular mechanisms that regulate gene expression in normal cells and in cancer cells is an important step for the development of therapeutic treatments of the tumor. Translation of proteins from mRNAs is one of the critical events in the expression of the genes. My laboratory investigates biological activities of RNA CUG triplet repeat binding protein, CUGBP1, and the role of this protein in liver proliferation and in liver tumors. In the liver, CUGBP1 regulates translation of several proteins including a member of C/EBP family, C/EBP¿, which is a key regulator of liver proliferation. CUGBP1 binds to the 5' region of C/EBP¿ mRNA and increases translation of a dominant negative molecule C/EBP¿-LIP, which inhibits biological activities of full-length C/EBP proteins and promotes liver proliferation. We have recently identified a new target of CUGBP1 in the liver. We obtain evidence that CUGBP1 binds to the 5' region of mRNA coding for a histone deacetylase 1, HDAC1, and increases translation of HDAC1 protein in tumor liver. We found that the translational activity of CUGBP1 is controlled mainly on the level of phosphorylation of CUGBP1. In tumor livers and in livers proliferating after partial hepatectomy (PH), CUGBP1 is hyper-phosphorylated and this phosphorylation increases translational activities of CUGBP1. We have found that cdk4 is a kinase which phosphorylates CUGBP1 and increases its translational activity. Therefore, the major hypothesis of this application is that the cdk4-mediated activation of CUGBP1 in the liver leads to the increased translation of C/EBP¿-LIP and HDAC1 and to development of tumor (see diagram on the left). Consistent with this hypothesis, levels of cyclin D1 and cyclin D3, which are activators of cdk4, are increased in cytoplasmic extracts of human liver tumors and in cytoplasm of mouse liver proliferating after partial hepatectomy. Specific Aim 1 examines mechanisms by which proliferating livers increase amounts of cdk4-D-type cyclins complexes in cytoplasm. Specific Aims 2 and 3 determine the role CUGBP1-C/EBP¿-LIP and CUGBP1-HDAC1 pathways in liver proliferation. We will test the hypothesis that CUGBP1-mediated elevation of HDAC1 inhibits promoters of liver specific tumor suppressor genes such as C/EBPa. In this application, we will also determine mechanisms by which liver blocks growth promotion activities of C/EBP¿-LIP under certain conditions such as Acute Phase Response. The understanding of the mechanisms of the elevation of C/EBP¿-LIP and HDAC1 in tumor will help to develop approaches to prevent malignant transformations in the liver. Public Health Relevance: This project investigates the role of RNA binding protein CUGBP1 in development of liver tumors and in the liver proliferation after partial hepatectomy (PH). The goals of this application are 1) to examine the role of RNA binding proteins in the development of liver tumors; 2) elucidate mechanisms by which proliferating livers activate RNA binding proteins; and 3) determine down-stream targets of RNA binding protein CUGBP1 in proliferating livers. The elucidation of molecular mechanisms of malignant transformations in liver will provide basement for the development of therapeutic approaches to prevent tumors.