Regulation of elF4E activity during oxidant stress

氧化应激期间 eF4E 活性的调节

• 批准号: 6693070
• 负责人: JEFFREY S SHENBERGER
• 金额: $ 13.15万
• 依托单位: DARTMOUTH COLLEGE
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-01-01 至 2007-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6693070
• 关键词: DNA replication; apoptosis; biological signal transduction; calcium ion; calmodulin dependent protein kinase; cell growth regulation; cell line; cell sorting; cell transformation; flow cytometry; free radical oxygen; genetic regulation; genetic translation; hydrogen peroxide; oxidative stress; polymerase chain reaction; protein biosynthesis; site directed mutagenesis; transfection /expression vector; translation factor

DESCRIPTION (provided by applicant): This application encompasses a 5-year training program designed for the development of a clinician scientist. The principal investigator is an Assistant Professor in the Department of Pediatrics at the Dartmouth Medical School with an interest in the role of oxidants in the pathogenesis of bronchopulmonary dysplasia. This program will advance the candidate's knowledge of molecular biology and cell signaling and develop technical skills necessary for success as an independent researcher. Dr. Aaron Barchowsky, an Associate Professor in the Department of Pharmacology and Toxicology, will serve as the mentor. He is well recognized as an authority in oxidant-mediated cell signaling in the lung and vasculature. Dr. Barchowsky has trained numerous graduate level students and several post-doctoral fellows. Additional expertise in protein synthetic mechanisms will be garnered from Dr. Stephen G. Zimmer, Associate Professor of Immunology and Microbiology at the University of Kentucky. Dr. Zimmer is well known as a leader in the investigation of translational regulation and cell growth. Lastly, an advisory committee composed of highly regarded researchers in pharmacology and physiology will provide further guidance and career direction. The goal of this proposal is to establish the role of eIF4E activity in the regulation of cell growth and protein synthesis following oxidant exposure. Preliminary evidence indicates that oxidants slow pulmonary epithelial cell growth and alter the phosphorylation of key translational regulatory proteins. The specific aims entail: 1) determining that oxidants increase eIF4E activity; 2) establishing that increased eIF4E activity protects cells from oxidant-mediated growth arrest and cell death; and 3) determining that activation of eIF4E is mediated through CaZ+/CaMK. Experimental design includes establishing growth arrest in A549 cells with H202, analyzing eIF4E activity through analysis of phosphorylation state, expression, cleavage, and binding; altering eIF4E activity by cellular transformation and site-directed mutagenesis; and determining kinase and phosphatase activities integral to the changes in eIF4E activity. These studies will generate new information concerning the inter-dependence of translation and cell growth and form a novel basis for pharmacological intervention to minimize oxidative injury.

描述（由申请人提供）： 该应用程序包括为临床科学家的发展而设计的5年培训计划。 主要研究者是达特茅斯医学院儿科系的助理教授，对氧化剂在支气管肺发育不良发病机制中的作用感兴趣。该计划将推进候选人的分子生物学和细胞信号传导的知识，并发展作为独立研究人员成功所需的技术技能。 药理学和毒理学系副教授Aaron Barchowsky博士将担任导师。 他被公认为肺和血管系统中氧化剂介导的细胞信号传导的权威。 Barchowsky博士培养了许多研究生水平的学生和一些博士后研究员。在蛋白质合成机制的额外专业知识将获得博士斯蒂芬G。肯塔基州大学免疫学和微生物学副教授齐默说。Zimmer博士是众所周知的翻译调控和细胞生长研究的领导者。最后，一个由药理学和生理学领域备受推崇的研究人员组成的咨询委员会将提供进一步的指导和职业方向。 本提案的目的是确定eIF 4 E活性在氧化剂暴露后细胞生长和蛋白质合成调节中的作用。 初步证据表明，氧化剂减缓肺上皮细胞的生长和改变关键的翻译调节蛋白的磷酸化。 具体目标包括：1）确定氧化剂增加eIF 4 E活性; 2）确定增加的eIF 4 E活性保护细胞免于氧化剂介导的生长停滞和细胞死亡;和3）确定eIF 4 E的活化通过CaZ+/CaMK介导。 实验设计包括用H2 O2在A549细胞中建立生长停滞，通过分析磷酸化状态、表达、切割和结合来分析eIF 4 E活性;通过细胞转化和定点诱变来改变eIF 4 E活性;以及确定与eIF 4 E活性变化相关的激酶和磷酸酶活性。这些研究将产生关于翻译和细胞生长的相互依赖性的新信息，并为药物干预以最大限度地减少氧化损伤形成新的基础。