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Study on the intracellular Network of TBX3

TBX3细胞内网络的研究

基本信息

批准号：
7318509
负责人：
TAOSHENG HUANG
金额：
$ 22.74万
依托单位：
UNIVERSITY OF CALIFORNIA-IRVINE
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-08-01 至 2011-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7318509
关键词：
American Basic Research Breast Cancer Binding Binding Sites Biological Biological Assay Biological Markers Boxing Breast Cancer Cell CDKN2A gene Cancer cell line Cells Chromatin Chromatin Structure Clinical Development Embryo Fibroblasts Gene Expression Gene Family Gene Targeting Genes Genetic Genetic Transcription Genome Glutathione S-Transferase Goals Growth and Development function HDAC1 gene Histone Deacetylase Human Immunohistochemistry In Situ Hybridization In Vitro Individual Knock-out Knockout Mice Lead Ligation Luciferases Malignant Neoplasms Mammary gland Mediating Molecular Molecular Biology Mus Mutation Normal tissue morphology Oligonucleotides Outcome Pathogenesis Pathway interactions Patients Physiological Play Precipitation Promoter Regions Protein Overexpression Proteins Recruitment Activity Research Research Personnel Role Sensitivity and Specificity Series Serum Syndrome TP53 gene Techniques Testing Tissues Transcription Repressor/Corepressor Transfection Tumor Suppressor Genes Tumor Suppressor Proteins Woman base breast cancer diagnosis chromatin immunoprecipitation clinical application cofactor experience human HDAC1 protein human disease in vitro Assay in vivo inhibitor/antagonist malignant breast neoplasm novel novel therapeutics programs promoter research study therapeutic target tool transcription factor tumor growth

项目摘要

DESCRIPTION (provided by applicant): Although breast cancer is one of the most common malignancies in American women, the genetic mechanisms responsible for its pathogenesis and clinical course remain largely unclear. TBX3 is a T-box transcription factor that may play a role in breast cancer and has not been previously investigated. Mutations of TBX3 in humans and mice show the clinical features of hypoplastic or absence of the mammary gland. Our earlier studies demonstrate that TBX3 is overexpressed in breast cancer cell lines, and can immortalize and transform mouse embryo fibroblast cells. TBX3 can also inhibit the expression of the p14ARF gene, a tumor suppressor and an inhibitor of MDM2-mediated degradation of p53. Therefore, overexpression of TBX3 decreases the stability of p53. Most recently, TBX3 was found to be elevated in serum of the patients with breast cancer. Collectively, these findings suggest that TBX3 plays an important role in breast cancer. In this proposal, we show that TBX3 interacts with histone deacetylase (HDAC)1, 2 and 5. Our central hypothesis is that TBX3 overexpression is associated with breast cancer and that TBX3 recruits HDACs to the p14ARF promoter and inhibits p14ARF tumor suppressor expression causing breast cancer. Aim 1, we will examine whether TBX3 is overexpressed in primary breast cancer tissues. Expression levels of TBX3 will be correlated with clinical outcomes and other established biomarkers. We anticipate that TBX3 could be a novel biomarker for breast cancer. TBX3 is mechanistically implicated in tumor growth, presumably by inhibiting the expression of p14ARF tumor suppressor gene. Although TBX3 has been characterized as a transcriptional repressor, the mechanism by which TBX3 represses gene expression is still unknown. Our preliminary results show that TBX3 interacts with HDACs and that the interaction is physiologically important. For Aim 2, we hypothesize that TBX3 recruits HDACs to p14ARF promoter and represses p14ARF tumor suppressor gene expression. We will test the interaction between TBX3 and HDACs by a series of in vivo and in vitro assays, including immunohistochemical analysis, Glutathione-S-transferase (GST) pulldown assay and chromatin immunoprecipitation (CHIP). Physiological significance of the TBX3-HDAC interaction will be further investigated in breast cancer cells and with the p14ARF promoter. In Aim 3, we propose to identify the TBX3 direct targets via chromatin immunoprecipitation-guided ligation selection (CHIP-GLAS). The CHIP-GLAS assay combines chromatin precipitation and microarray, which could be used to identify 20,000 promoter DNA/transcription factor interactions in one experiment with high sensitivity and specificity. Using this approach, we found that TBX3 binds to more than 600 promoters. We will verify those target genes involved in breast cancer and mammary gland development using various in vivo and in vitro assays. An important feature of the proposed research is translational for the development of a novel biomarker for breast cancer and basic biological information to clinical application will be optimized. Elucidation of the TBX3- HDAC interaction will deepen our understanding of the functions of TBX3 and may also lead to the identification of a novel therapeutic target for breast cancer.

描述（由申请人提供）：虽然乳腺癌是美国女性最常见的恶性肿瘤之一，但其发病机制和临床过程的遗传机制仍不清楚。TBX 3是一种T-box转录因子，可能在乳腺癌中发挥作用，以前尚未研究过。人类和小鼠中TBX 3的突变显示出乳腺发育不全或缺失的临床特征。我们早期的研究表明TBX 3在乳腺癌细胞系中过表达，并且可以永生化和转化小鼠胚胎成纤维细胞。TBX 3还可以抑制p14 ARF基因的表达，p14 ARF基因是一种肿瘤抑制因子，也是MDM 2介导的p53降解的抑制剂。因此，TBX 3的过表达降低了p53的稳定性。最近，发现TBX 3在乳腺癌患者的血清中升高。总的来说，这些发现表明TBX 3在乳腺癌中起着重要作用。在这个提议中，我们表明TBX 3与组蛋白脱乙酰酶（HDAC）1，2和5相互作用。我们的中心假设是TBX 3过表达与乳腺癌相关，TBX 3将HDAC募集到p14 ARF中，启动子和抑制p14 ARF肿瘤抑制基因的表达，导致乳腺癌。目的1，我们将检查TBX 3是否在原发性乳腺癌组织中过表达。TBX 3的表达水平将与临床结果和其他已建立的生物标志物相关。我们预计TBX 3可能是乳腺癌的新生物标志物。TBX 3可能通过抑制p14 ARF肿瘤抑制基因的表达而在机制上参与肿瘤生长。虽然TBX 3已被鉴定为转录抑制因子，但TBX 3抑制基因表达的机制仍不清楚。我们的初步结果表明，TBX 3与HDAC相互作用，这种相互作用在生理上是重要的。对于目的2，我们假设TBX 3招募HDAC到p14 ARF启动子并抑制p14 ARF肿瘤抑制基因的表达。我们将测试TBX 3和HDAC之间的相互作用，通过一系列的在体内和体外试验，包括免疫组织化学分析，谷胱甘肽-S-转移酶（GST）下拉分析和染色质免疫沉淀（CHIP）。TBX 3-HDAC相互作用的生理学意义将在乳腺癌细胞中和p14 ARF启动子下进一步研究。在目标3中，我们提出通过染色质免疫沉淀引导的连接选择（CHIP-GLAS）来识别TBX 3直接靶标。CHIP-GLAS检测结合了染色质沉淀和微阵列，可用于在一个实验中以高灵敏度和特异性鉴定20，000个启动子DNA/转录因子相互作用。使用这种方法，我们发现TBX 3与600多个启动子结合。我们将使用各种体内和体外检测方法来验证这些参与乳腺癌和乳腺发育的靶基因。拟议研究的一个重要特征是转化为乳腺癌的新型生物标志物的开发，并将优化临床应用的基本生物信息。TBX 3- HDAC相互作用的阐明将加深我们对TBX 3功能的理解，也可能导致识别乳腺癌的新治疗靶点。