Regulation of elF4E activity during oxidant stress

氧化应激期间 eF4E 活性的调节

• 批准号: 7001198
• 负责人: JEFFREY S SHENBERGER
• 金额: $ 5.37万
• 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 2006-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7001198
• 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）： 该应用程序包含一个为期 5 年的培训计划，旨在培养临床科学家。 主要研究者是达特茅斯医学院儿科系的助理教授，他对氧化剂在支气管肺发育不良发病机制中的作用感兴趣。该计划将提高候选人在分子生物学和细胞信号传导方面的知识，并培养作为独立研究人员取得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。 他被公认为肺和脉管系统氧化剂介导的细胞信号传导领域的权威。 Barchowsky 博士培养了众多研究生水平的学生和数名博士后研究员。肯塔基大学免疫学和微生物学副教授 Stephen G. Zimmer 博士将获得蛋白质合成机制方面的更多专业知识。 Zimmer 博士是众所周知的翻译调控和细胞生长研究领域的领导者。最后，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。 该提案的目标是确定 eIF4E 活性在氧化剂暴露后调节细胞生长和蛋白质合成中的作用。 初步证据表明，氧化剂会减缓肺上皮细胞的生长并改变关键翻译调节蛋白的磷酸化。 具体目标包括：1) 确定氧化剂增加 eIF4E 活性； 2) 确定增加的 eIF4E 活性可以保护细胞免受氧化剂介导的生长停滞和细胞死亡； 3)确定eIF4E的激活是通过CaZ+/CaMK介导的。 实验设计包括用 H202 在 A549 细胞中建立生长停滞，通过分析磷酸化状态、表达、裂解和结合来分析 eIF4E 活性；通过细胞转化和定点诱变改变 eIF4E 活性；确定与 eIF4E 活性变化相关的激酶和磷酸酶活性。这些研究将产生有关翻译和细胞生长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奠定新的基础。