RNA Binding Protein CUGBP2 in Intestinal Epithelium

肠上皮细胞中的 RNA 结合蛋白 CUGBP2

基本信息

批准号：
7924796
负责人：
Shrikant Anant
金额：
$ 35.53万
依托单位：
UNIVERSITY OF KANSAS MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-09-01 至 2012-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7924796
关键词：
3&apos Untranslated Regions 5&apos Untranslated Regions Adenocarcinoma Adenoviruses Affect Alternative Splicing Amino Acids Apoptosis Apoptotic Area Behavior Binding Cell Death Cell Survival Cell physiology Cells Cessation of life Cis-Acting Sequence Code Colon Colon Carcinoma Colonic Neoplasms Colorectal Cancer Complementary DNA Diagnosis Disease Epithelial Cells Event Exclusion Exons Family Gene Expression Gene Targeting Genes Genetic Transcription Goals Human Intestines Introns Luciferases Malignant Neoplasms Mediating Messenger RNA Microtubule-Associated Proteins Mitotic Mus Mutagenesis Neoplasm Metastasis Normal Cell Nuclear Nuclear Export Nude Mice Open Reading Frames Pathway interactions Physiological Process Protein Isoforms Protein Overexpression Proteins Proto-Oncogenes RNA Binding RNA Splicing RNA-Binding Proteins Recurrence Regulation Research Role Signal Transduction Spliced Genes Therapeutic Therapeutic Intervention Trans-Activators Translations Tumor Promoters Tumor Suppressor Genes Tumor Suppressor Proteins United States Variant Vascularization Xenograft Model adenoma base cancer cell cell killing cis acting element cyclooxygenase 2 hnRNP A1 in vivo intestinal epithelium irradiation mRNA Stability member neoplastic cell novel overexpression promoter protein protein interaction public health relevance research study response tumor tumor growth tumor xenograft tumorigenesis

项目摘要

DESCRIPTION (provided by applicant): The broad goal of this application is to understand the mechanisms by which RNA binding protein CUGBP2 regulates gene expression at the posttranscriptional level of mRNA stability and translation in intestinal epithelial cells. Overexpression of the protein in cancer cells results in cell death. We have determined that CUGBP2 interacts with AU-rich sequences in the 3'untranslated region of cyclooxygenase-2 and Mcl-1 mRNAs and upon binding downregulates the translation of both mRNAs. We have also determined that CUGBP2 levels are decreased in cancer cells but its expression is elevated in cells undergoing mitotic catastrophe. In addition, CUGBP2 is regulated at the levels of transcription by three different promoters. Activation from promoters located upstream from the commonly used proximal promoter results in the inclusion of additional amino acids in the N-terminus of the protein. Our studies suggest that the consequence of this addition is differences in cellular localization of the protein, but more importantly in the activity of the protein. The variants with the added amino acids do not affect cell viability. In addition, there are differential effects on target gene splicing with the three variants. Based on these observations, we have proposed to determine three aims. In Aim 1, we will determine the mechanism(s) by which the three CUGBP2 promoters are regulated. In addition, CUGBP2 is regulated at the posttranscriptional level of alternative splicing resulting in the novel inclusion of one intron in the 5'untranslated region. We will identify the cis-acting sequences and trans-acting factors regulating this process. In Aim 2, we will determine the cellular functions of CUGBP2. We have identified cellular factors that bind to CUGBP2. We will perform systematic deletion and fine mutagenesis to identify the domains in CUGBP2 that are involved in RNA:protein and protein:protein interactions. In addition, we will determine the CUGBP2 domains that regulate CUGBP2 localization under basal conditions and when the cells are exposed to 3-irradiation. We will also determine the effect of these interactions on CUGBP2 mediated RNA splicing, mRNA stability and translation regulation. In Aim 3, we will determine whether the CUGBP2 gene is both a novel tumor suppressor and a protooncogene. We will determine the expression levels of the three isoforms in a panel of human colon tumors. Furthermore, we will determine whether the different CUGBP2 isoforms affect tumor behavior in xenograft models. Completion of these experiments should give us a better understanding of how the RNA binding protein CUGBP2 functions in normal epithelial cells, and whether changes in the CUGBP2 expression that is observed in tumor cells is responsible for tumor behavior. PUBLIC HEALTH RELEVANCE: Colorectal cancer is the leading in cancer related deaths in the United States. Understanding how the normal cell progresses to a cancer will aid in our developing novel therapies for both diseases. We have identified a protein, CUGBP2 whose expression is lost in cancer cells. Restoring the protein into the cancer cell kills the cells. We are in the process of identifying how the gene expressing CUGBP2 is silenced in cancer cells so that we can determine ways to reverse the process. This might stop or slow down the tumorigenesis.

描述（由申请人提供）：本申请的主要目标是了解RNA结合蛋白CUGBP2在肠上皮细胞中mRNA稳定性和翻译的转录后水平调节基因表达的机制。癌细胞中蛋白质的过度表达会导致细胞死亡。我们已经确定，CUGBP2 与 cyclooxygenase-2 和 Mcl-1 mRNA 3'非翻译区中富含 AU 的序列相互作用，并在结合后下调这两种 mRNA 的翻译。我们还确定，癌细胞中的 CUGBP2 水平降低，但在经历有丝分裂灾难的细胞中其表达升高。此外，CUGBP2 在转录水平上受到三个不同启动子的调节。位于常用近端启动子上游的启动子的激活导致在蛋白质的 N 末端包含额外的氨基酸。我们的研究表明，这种添加的结果是蛋白质细胞定位的差异，但更重要的是蛋白质活性的差异。添加氨基酸的变体不会影响细胞活力。此外，三种变体对靶基因剪接的影响也不同。基于这些观察，我们建议确定三个目标。在目标 1 中，我们将确定三个 CUGBP2 启动子的调控机制。此外，CUGBP2 在选择性剪接的转录后水平受到调节，导致 5' 非翻译区新包含一个内含子。我们将确定调节该过程的顺式作用序列和反式作用因子。在目标 2 中，我们将确定 CUGBP2 的细胞功能。我们已经鉴定出与 CUGBP2 结合的细胞因子。我们将进行系统删除和精细诱变，以鉴定 CUGBP2 中参与 RNA:蛋白质和蛋白质:蛋白质相互作用的结构域。此外，我们将确定在基础条件下以及当细胞暴露于 3 辐射时调节 CUGBP2 定位的 CUGBP2 结构域。我们还将确定这些相互作用对 CUGBP2 介导的 RNA 剪接、mRNA 稳定性和翻译调控的影响。在目标 3 中，我们将确定 CUGBP2 基因是否既是一种新型肿瘤抑制基因，又是一种原癌基因。我们将确定一组人类结肠肿瘤中三种亚型的表达水平。此外，我们将确定不同的 CUGBP2 亚型是否影响异种移植模型中的肿瘤行为。这些实验的完成应该让我们更好地了解 RNA 结合蛋白 CUGBP2 在正常上皮细胞中如何发挥作用，以及在肿瘤细胞中观察到的 CUGBP2 表达变化是否与肿瘤行为有关。公共卫生相关性：结直肠癌是美国癌症相关死亡的主要原因。了解正常细胞如何发展为癌症将有助于我们开发针对这两种疾病的新疗法。我们已经鉴定出一种蛋白质 CUGBP2，其表达在癌细胞中丢失。将蛋白质恢复到癌细胞中会杀死细胞。我们正在确定表达 CUGBP2 的基因如何在癌细胞中被沉默，以便我们能够确定逆转这一过程的方法。这可能会阻止或减缓肿瘤发生。

项目成果

期刊论文数量（19）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Crocetin: an agent derived from saffron for prevention and therapy for cancer.

DOI：
10.2174/138920112798868566
发表时间：
2012-01
期刊：
Current pharmaceutical biotechnology
影响因子：
2.8
作者：
Gutheil WG;Reed G;Ray A;Anant S;Dhar A
通讯作者：
Dhar A

Reduced Expression of RNA Binding Protein CELF2, a Putative Tumor Suppressor Gene in Colon Cancer.

DOI：
10.7178/ig.1.1.7
发表时间：
2012
期刊：
Immuno-gastroenterology
影响因子：
0
作者：
Ramalingam S;Ramamoorthy P;Subramaniam D;Anant S
通讯作者：
Anant S

Honokiol in combination with radiation targets notch signaling to inhibit colon cancer stem cells.

DOI：
10.1158/1535-7163.mct-11-0999
发表时间：
2012-04
期刊：
Molecular cancer therapeutics
影响因子：
5.7
作者：
Ponnurangam S;Mammen JM;Ramalingam S;He Z;Zhang Y;Umar S;Subramaniam D;Anant S
通讯作者：
Anant S

Urine and serum analysis of consumed curcuminoids using an IkappaB-luciferase surrogate marker assay.

使用 IkappaB-荧光素酶替代标记物测定对消耗的类姜黄素进行尿液和血清分析。

DOI：
发表时间：
2010
期刊：
In vivo (Athens, Greece)
影响因子：
0
作者：
Ponnurangam,Sivapriya;Mondalek,FadeeG;Govind,Janita;Subramaniam,Dharmalingam;Houchen,CourtneyW;Anant,Shrikant;Pantazis,Panayotis;Ramanujam,RamaP
通讯作者：
Ramanujam,RamaP

Cancer stem cells: a novel paradigm for cancer prevention and treatment.