The role of histone phosphorylation in arsenic-induced cell transformation and sk

组蛋白磷酸化在砷诱导的细胞转化和皮肤病中的作用

基本信息

批准号：
7561683
负责人：
Zigang Dong
金额：
$ 33.98万
依托单位：
UNIVERSITY OF MINNESOTA
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-02-01 至 2013-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7561683
关键词：
Address Adverse effects Agar Amino Acids Arsenic Arts Biological Assay Carcinogens Cells Chemicals Chemoprevention Chemopreventive Agent Computer Simulation Computers Crystallography Databases Development Environmental Carcinogens Genes Goals Histone H3 Histones Human Hybrids In Vitro Kaempferols Knock-out Knockout Mice Knowledge Malignant Neoplasms Molecular Molecular Models Molecular and Cellular Biology Mus Mutation Phosphorylation Phosphorylation Site Phosphotransferases Play Point Mutation Preclinical Drug Evaluation Process Protamine Kinase Protein Kinase Role Screening procedure Skin Cancer Skin Carcinogenesis Small Interfering RNA Structure Technology Testing Tumor Promotion Work base cancer chemoprevention cancer prevention carcinogenesis cell transformation design inhibitor/antagonist kaempferol mass spectrometer molecular modeling novel overexpression public health relevance stable cell line ultraviolet upstream kinase

项目摘要

DESCRIPTION (provided by applicant): Arsenic is a well-documented human carcinogen. Our goal is to address the central hypothesis that phosphorylation of histones and their upstream kinases play an important functional role in arsenic-induced cell transformation and carcinogenesis. Specific Aim 1 is to study the role of histone phosphorylation in arsenic-induced cell transformation; Specific Aim 2 is to investigate and identify the histone kinases that phosphorylate histone H3 and H2B at different amino acid residues; Specific Aim 3 is to study the crystal structure of histone kinase RSK2, perform in- silico screening and design RSK2 inhibitors for suppressing arsenic-induced histone phosphorylation and cell transformation; and Specific Aim 4 is to study the role of RSK2 in arsenic/ultraviolet A (UVA)-induced skin carcinogenesis and determine RSK2's potential as a target for chemoprevention of cancer. The strategy for Specific Aim 1 is to use point mutations at key phosphorylation sites of histone H3 and H2B siRNA gene knockdown and overexpressing stable cell lines to test the role of H3 and H2B in soft agar cell transformation assays. For Specific Aim 2, we will use in vitro kinase assays, specific mutations, LTQ Orbitrap hybrid mass spectrometer analysis and RSK2 knockout cells as well as inhibitors of RSK2. In Specific Aim 3, we will use x-ray crystallography to determine the structure of RSK2. Then we will use a super computer to screen a database of 2.5 million chemicals to find inhibitors for RSK2 to be tested in an in vitro kinase assay. In Specific Aim 4, we will test the effect of the RSK2 inhibitor kaempferol and RSK2 knockout mice in UVA/arsenic-induced mouse skin carcinogenesis. Such knowledge will facilitate the design of more effective and specific strategies with fewer side effects for chemoprevention of arsenic- induced cancer. PUBLIC HEALTH RELEVANCE: Environmental arsenic contamination is a major problem in many parts of the world and is a well-documented human carcinogen. By using state-of-the-art technology such as x-ray crystallography, super computer based molecular modeling and drug screen, cellular and molecular biology and gene knockout mice, we will study the novel mechanism involved in histone phosphorylation on arsenic-induced cell transformation to cancer and the carcinogenesis process. These studies will facilitate the development of more effective agents with fewer side effects for chemoprevention against environmental carcinogens such as arsenic- induced cancer.

描述（由申请人提供）：砷是一种有据可查的人类致癌物。我们的目标是解决以下中心假设：组蛋白及其上游激酶的磷酸化在砷诱导的细胞转化和致癌作用中起着重要的功能作用。具体目的1是研究组蛋白磷酸化在砷诱导的细胞转化中的作用。具体目的2是研究和鉴定在不同氨基酸残基上磷酸化组蛋白H3和H2B的组蛋白激酶。具体目的3是研究组蛋白激酶RSK2的晶体结构，进行内硅筛选和设计RSK2抑制剂，以抑制砷诱导的组蛋白磷酸化和细胞转化。具体目的4是研究RSK2在砷/紫外线A（UVA）诱导的皮肤致癌作用中的作用，并确定RSK2的潜力作为癌症化学预防的靶标。特定目标1的策略是在组蛋白H3和H2B siRNA基因敲低的关键磷酸化位点使用点突变，并过表达稳定的细胞系来测试H3和H2B在软琼脂细胞转化测定中的作用。对于特定目标2，我们将使用体外激酶测定，特异性突变，LTQ Orbitrap杂交质谱仪分析和RSK2基因敲除细胞以及RSK2的抑制剂。在特定的目标3中，我们将使用X射线晶体学来确定RSK2的结构。然后，我们将使用一台超级计算机筛选一个250万种化学品的数据库，以查找在体外激酶测定中测试的RSK2的抑制剂。在特定目标4中，我们将测试RSK2抑制剂Kaempferol和RSK2敲除小鼠在UVA/砷诱导的小鼠皮肤致癌作用中的作用。这些知识将促进更有效和具体的策略的设计，而对于砷诱导的癌症进行化学预防的副作用较少。公共卫生相关性：环境砷污染是世界许多地方的主要问题，是有据可查的人类致癌物。通过使用最先进的技术，例如X射线晶体学，基于超级计算机的分子建模和药物筛选，细胞和分子生物学以及基因敲除小鼠，我们将研究涉及砷诱导细胞转化为癌症的组蛋白磷酸化的新型机制，以及癌症。这些研究将促进对针对环境致癌物（如砷诱导的癌症）进行化学预防的副作用更少的更有效药剂的发展。