POSTTRANSCRIPTIONAL REGULATION OF ENOS

申请人和其他实验室先前的工作表明，内皮型一氧化氮合酶（eNOS）通过mRNA半衰期的变化在转录后受到调节。申请人已经证明，在内皮细胞生长期间，eNOS mRNA的半衰期大约是细胞静止时的3倍。这似乎是通过Enos 3'utr中的新序列与一个51 kDA蛋白之间的相互作用发生的，该蛋白被命名为Nos不稳定因子的信息（MONSDER）。这项研究的第一个目标是克隆和测序它们的蛋白质，命名为Nos不稳定因子信息（MONSDER）。第二个目标是使用四环素调控的逆转录病毒载体系统在内皮细胞中过表达MONSDER，并确定其对eNOS表达的影响。第三个目的是研究eNOS mRNA稳定性与核糖体组装之间的关系，并确定MONSDER过表达对这一过程的影响。总的来说，这些研究应该为内皮细胞一氧化氮生成的调节提供新的机制。作为该病房的一部分，申请人将在药理学部门与tj Murphy博士一起工作两年，他在mRNA稳定性领域具有专业知识。随后，他将在埃默里大学心脏病学部门获得教职和实验室空间，并将继续与dr。墨菲和大卫·哈里森博士，他以前的导师和心脏病学教员。申请人的最终目标是从事心血管基础研究工作。在本奖项期间所获得的研究经验将使Searles博士成为一名独立研究者，拥有实现这一目标所需的专业知识和工具。

Previous work, performed by the applicant, and by other laboratories, has shown that the endothelial nitric oxide synthase (eNOS) is regulated post- transcriptionally via changes in mRNA half-life. The applicant has shown that during endothelial cell growth, eNOS mRNA half-life is approximately 3 times longer than when cells are quiescent. This seems to occur via interactions between novel sequences in the Enos 3'utr AND A 51 kDA protein which has been named Message of Nos DestabilizER (MONSDER). The first aim of this research will be to clone and sequence their protein, named Message of Nos DestabilizER (MONSDER). The second aim will be to over-express MONSDER in endothelial cells using a tetracycline- regulated retroviral vector system, and determine the effect of this on eNOS expression. The third aim will be to examine the relationship between eNOS mRNA stability and ribosomal assembly, and determine the effect of MONSDER over-expression on this process. Overall, these studies should provide insight into a novel mechanism of regulation of endothelial nitric oxide production. As a portion of this ward, the applicant will spend two years working in the Department of Pharmacology with Dr. T.J. Murphy, who has expertise in the area of mRNA stability. Subsequently, he will be given a faculty position and laboratory space in the Cardiology Division at Emory, and will continue to have interactions with and guidance from Drs. Murphy and Dr. David Harrison, his previous mentor and a faculty member in Cardiology. The applicant's ultimate goal is to pursue a career in basic cardiovascular research. The experience gained performing this research during the period of this award should permit Dr. Searles to become an independent investigator with the expertise and tools necessary to achieve this goal.