Functional Definition of MYC Acetylation

MYC 乙酰化的功能定义

• 批准号: 8717607
• 负责人: Matthew Thomas Hurd
• 金额: $ 3.54万
• 依托单位: UNIVERSITY OF CALIFORNIA RIVERSIDE
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-01 至 2015-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8717607
• 关键词: Acetylation; Affect; Animals; Antibodies; Antineoplastic Agents; Apoptosis; Apoptotic; Attenuated; Biochemical; Biological; Biological Assay; Biological Markers; Cancer cell line; Cell Death; Cell Proliferation; Cells; Cellular Stress; Data; Development; EP300 gene; Enzymes; Fibroblasts; Gene Expression; Gene Expression Regulation; Gene Targeting; Genes; Genetic Transcription; Genome; Growth Factor; Health; Hela Cells; Homeostasis; Human; Interleukin-2; Intervention; Lysine; MYC Family Protein; Malignant Neoplasms; Malignant neoplasm of cervix uteri; Mammalian Cell; Mediating; Methods; Molecular; Mus; Normal Cell; Outcome; Pharmacotherapy; Phosphorylation; Post-Translational Protein Processing; RNA Interference; Reporter Genes; Reporting; Research; Role; Serum; Signal Pathway; Signal Transduction; Site; Stimulus; System; Testing; Transcription Coactivator; c-myc Genes; cancer cell; cancer type; cell growth; cell type; chromatin immunoprecipitation; genome-wide; glycosylation; histone acetyltransferase; metaplastic cell transformation; multicatalytic endopeptidase complex; mutant; novel; overexpression; research study; transcription factor; tumor; tumorigenic

MYC is a transcription factor that targets a large fraction (~15%) of the genome and is tightly regulated in normal cells and essential for normal development and homeostasis in most animals. Deregulated MYC expression (e.g., overexpression) is tumorigenic in mice and associated with most types of cancer in humans. MYC deregulation may occur via alterations of the c-myc gene itself (e.g., translocation and amplification) or via aberrant activation of upstream signaling pathways that activate c-myc gene expression and/or increase MYC protein stability via post-translational modifications (PTMs). The PTMs of MYC are, however, poorly characterized and their roles in the multiple cellular and molecular activities of MYC are still poorly defined and controversial. Until recently phosphorylation and O-glycosylation of a few residues were the only PTMs reported, which affect MYC ubiqitination and degradation by the proteasome. Our lab recently uncovered novel PTMs of MYC and found that MYC is acetylated at several lysine (K) residues by transcription coactivators with histone acetyltransferase (HAT) activity (i.e., GCN5 and p300). The general objective of my research is to provide definitive evidence that native MYC in mammalian cells is regulated via acetylation and to define the cellular and molecular functions of MYC acetylation at specific residues. AIM 1: To establish and characterize acetylation of native MYC in mammalian cells using acetylated site-specific antibodies (MYC-K148Ac, MYC-K157Ac, MYC-K323Ac) Acetylation of endogenous MYC has yet to be demonstrated in any cell type. I will use new antibodies that recognize specifically human MYC acetylated residues K148, K157, and K323 to profile acetylation of MYC at these different sites in several non-transformed and transformed/cancer cell lines. AIM 2: To identify functions of MYC acetylated residues K148(149) and K157(158) I will test the role of MYC acetylated residues K148 and K157 (human coordinates homologous to mouse MYC K149 and K158) in MYC-induced cell proliferation, transformation and apoptosis. AIM 3: To establish and define the role(s) of MYC acetylation in target gene regulation Selected MYC target genes affected by the R158 (or R149) substitution identified in preliminary experiments (e.g. Nrf1) and in Aim 2 will be analyzed to tes the possible role of acetylation at K158 (or K149) in regulating MYC transcription functions. Various methods, including chromatin immunoprecipitation (ChIP) assays, RNA interference (RNAi), and reporter gene assays will be used.

MYC是一种转录因子，靶向大量基因组（约15％），在正常细胞中受到严格调节，对于大多数动物的正常发育和稳态至关重要。失控的MYC表达（例如，过表达）在小鼠中是肿瘤的，并且与人类的大多数癌症有关。 MYC放松管制可能通过改变C-MYC基因本身（例如易位和扩增）或通过异常激活上游信号传导途径的异常激活，从而激活C-MYC基因表达和/或通过翻译后修饰（PTMS）增加MYC蛋白质稳定性。但是，MYC的PTM的特征很差，并且它们在MYC的多个细胞和分子活性中的作用仍然很差，并且引起了争议。直到最近，少数残基的磷酸化和O-糖基化是唯一报告的PTM，它影响了蛋白酶体的Myc ubiQitation和降解。我们的实验室最近发现了MYC的新型PTM，发现MYC通过具有组蛋白乙酰基转移酶（HAT）活性的转录共激活剂在几种赖氨酸（K）残基上被乙酰化（即GCN5和P300）。我的研究的一般目标是提供明确的证据，表明哺乳动物细胞中的天然MYC通过乙酰化调节，并定义Myc乙酰化在特定残基处的细胞和分子功能。 目的1：使用乙酰化位点特异性抗体（MYC-K148AC，MYC-K157AC，MYC-K323AC）乙酰化乙酰化乙酰化乙酰化乙酰化乙酰化乙酰化乙酰化的内源性myc尚未在任何细胞类型中证明乙酰化的乙酰细胞中乙酰化。我将使用新的抗体，这些抗体在几个非转化和转化的/癌细胞系中，在这些不同位点识别人类Myc乙酰化残基K148，K157和K323来介绍MYC的乙酰化。 目标2：要确定MYC乙酰化残基K148（149）和K157（158）的功能，我将测试Myc乙酰化残基K148和K157的作用（与小鼠MYC MYC K149和K158同源的MyC诱导的细胞扩散和转化，转化，转化，转化和apoptosis和Apoptosis和Apoptosis。 目的3：建立并定义了在选定的MYC靶基因调节中的MYC乙酰化的作用，该目标基因受到初步实验（例如NRF1）的R158（或R149）替代的影响（或R149）的作用（例如NRF1），并且将在K158（或K158）（或K149）中分析乙酰化的可能作用。将使用各种方法，包括染色质免疫沉淀（CHIP）测定，RNA干扰（RNAI）和报告基因测定。