Mechanisms of Gastrointestional Growth & Transformation

胃肠生长机制

基本信息

批准号：
7895949
负责人：
JUANITA L. MERCHANT
金额：
$ 7.37万
依托单位：
UNIVERSITY OF MICHIGAN AT ANN ARBOR
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：
2009-08-15 至 2010-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7895949
关键词：
Acetylation Apoptosis Ataxia Telangiectasia Binding Binding Proteins Biological Assay Butyrates Cell Cycle Cell Cycle Arrest Cell Cycle Regulation Pathway Cell Fractionation Cells Chromatin Complex Confocal Microscopy DNA DNA Binding DNA Damage DNA-Binding Proteins EP300 gene Elements Event Exhibits Funding G1 Arrest GC Rich Sequence Gene Targeting Genes Genetic Goals Growth HDAC1 gene Histone Acetylation Histone Deacetylase Histone Deacetylase Inhibitor Histones Malignant neoplasm of gastrointestinal tract Mediating Molecular Mutate Mutation Neoplastic Cell Transformation Nuclear Protein Nuclear Proteins Phosphorylation Protein Binding Protein p53 Proteins Recruitment Activity Regulation Role Site Site-Directed Mutagenesis Spermatogenesis TP53 gene Testing Trichostatin A Tumor Suppressor Proteins Zinc Fingers ataxia telangiectasia mutated protein base cell growth chromatin remodeling gastrointestinal histone acetyltransferase prevent promoter protein protein interaction response transcription factor

项目摘要

The molecular basis for the gene specific effects of butyrate remains poorly defined. Butyrate's major known function involves inhibition of histone deacetylases (HDACs) resulting in increased acetylation. In addition to histone acetylation, it is now known that DNA binding proteins become acetylated. The proposed function of acetylated transcription factors varies and includes increased or decreased DNA binding as well as protein stability. In many instances, the genetic targets of butyrate are GC-rich sequences that bind Sp1 and Sp3. Recruitment of the histone acetyltransferase (HAT) p300 cooperates with Sp1 and Sp3 to mediate the effects of butyrate to the p21waf1 promoter. However, Sp1 does not complex with p300, but instead binds HDAC1. We have shown previously that ZBP-89 is another DNA binding protein that binds GC-rich sites and mediates butyrate induction of p21waf1. Understanding the mechanisms by which butyrate suppresses growth through ZBP-89 is the focus of this competing renewal. ZBP-89 is an 89 kDa Kruppel-type zinc finger protein composed of 794 residues. During the past funding period, we demonstrated that ZBP-89 interacts with the tumor suppressor protein p53 to induce G1 arrest. We have recently found that ZBP-89 interacts with the tumor suppressor protein ataxia telangiectasia, mutated (ATM) in a butyrate specific manner. ATM modulates factors involved in both G1 and G2 cell arrest after DNA damage. ATM mediates cell cycle arrest through phosphorylation of p53 at Ser15. ZBP-89 is required for phosphorylation of p53 at Ser15. Therefore the specific aims of this proposal are 1) To dissect the interaction of ZBP-89 with ATM in response to butyrate. 2) To dissect the regulation of p53 activation by ZBP-89 in response to butyrate. 3) To dissect the mechanisms of p300 HAT activation in the regulation of ZBP-89. 4) To determine whether ZBP-89 exhibits tumor suppressor function. Site-direct mutations in various domains of ZBP-89 will be used to dissect the interactions with these cell cycle regulators. ZBP-89 interactions with chromatin will be studied using confocal microscopy, cell fractionation and ChIP assays. In this way, we will further the understanding of how butyrate inhibition of HDACs ultimately suppresses cell growth and prevents neoplastic transformation.

丁酸酯基因特异性作用的分子基础仍然很差。丁酸酯的主要已知功能涉及抑制组蛋白脱乙酰基酶（HDAC），导致乙酰化增加。除了组蛋白乙酰化外，现在已经知道DNA结合蛋白成为乙酰化。乙酰化转录因子的拟议功能变化，包括增加或减少的DNA结合以及蛋白质稳定性。在许多情况下，丁酸酯的遗传靶标是结合SP1和SP3的富含GC的序列。组蛋白乙酰基转移酶（HAT）P300的募集与SP1和SP3合作，介导丁酸酯对P21WAF1启动子的影响。但是，SP1与p300不复杂，而是结合HDAC1。我们先前已经证明，ZBP-89是另一种DNA结合蛋白，它结合了富含GC的位点并介导P21WAF1的丁酸酯诱导。理解丁酸酯通过ZBP-89抑制生长的机制是这种竞争更新的重点。 ZBP-89是由794个残基组成的89 kDa kruppel型锌指蛋白。在过去的资金期间，我们证明了ZBP-89与肿瘤抑制蛋白p53相互作用以诱导G1停滞。我们最近发现，ZBP-89与肿瘤抑制蛋白共济失调的毛细血管扩张相互作用，以丁酸酯特异性方式突变（ATM）。 ATM调节DNA损伤后G1和G2细胞停滞涉及的因素。 ATM通过在Ser15处的p53磷酸化来介导细胞周期停滞。 ZBP-89是在Ser15处p53磷酸化所必需的。因此，该提案的具体目的是1）剖析ZBP-89与ATM的相互作用，以响应丁酸酯。 2）解剖ZBP-89对丁酸酯的调节激活。 3）在调节ZBP-89中剖析P300 HAT激活的机制。 4）确定ZBP-89是否表现出肿瘤抑制功能。 ZBP-89各个结构域中的位置突变将用于剖析与这些细胞周期调节剂的相互作用。 ZBP-89与染色质的相互作用将使用共聚焦显微镜，细胞分馏和CHIP分析进行研究。通过这种方式，我们将进一步理解丁酸HDAC抑制HDAC的最终抑制细胞生长并防止肿瘤转化。