Determining the Role of the Stem-loop Binding Protein in Histone mRNA Decay.

确定茎环结合蛋白在组蛋白 mRNA 衰变中的作用。

• 批准号: 7806727
• 负责人: Stacie Ann Meaux
• 金额: $ 4.76万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-01 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7806727
• 关键词: Abnormal Cell; Amino Acids; Back; Binding Proteins; Biochemical; Biochemistry; Biological; Cell Cycle; Cell Nucleus; Cells; Chromosomes; Codon Nucleotides; Complex; Coupled; Cytoplasm; DNA biosynthesis; Educational process of instructing; Elements; Eukaryota; Faculty; Gene Expression; Gene Expression Regulation; Genomic Instability; Goals; Histones; Human; In Vitro; Knowledge; Lead; Life; Link; Malignant Neoplasms; Messenger RNA; Molecular; Pathway interactions; Phase; Play; Positioning Attribute; Postdoctoral Fellow; Process; Proteins; Recruitment Activity; Regulation; Research; Role; Saccharomyces cerevisiae; Signal Transduction; Staging; Techniques; Testing; Training; Transcript; Translations; Work; Yeasts; cancer therapy; career; cell growth; experience; graduate student; human disease; in vivo; insight; mRNA Decay; mRNA Transcript Degradation; mutant; programs; protein function; research study; stem; tool

DESCRIPTION (provided by applicant): Histone expression in metazoans is a highly regulated process that is linked with DNA synthesis during the cell cycle. Histone mRNA levels remain at a low basal level throughout most of the life of a cell but increase about 30-fold at the beginning of S phase, when DNA synthesis begins. At the end of S-phase, histone mRNA levels decrease back to basal level due to rapid degradation of the transcripts in the cytoplasm; however, little evidence exists to explain how degradation of histone mRNAs is triggered. The stem-loop binding protein (SLBP) is required for processing, export and translation of histone mRNAs. SLBP is also hypothesized to play a critical role in histone mRNA degradation by recognizing, or causing, aberrant translation termination and/or recruiting decay proteins to degrade the transcript. However, the role of SLBP In histone mRNA decay has not been thoroughly evaluated. I hypothesize that SLBP plays a critical role In recognizing, or causing, aberrant translation termination and/or in initiating mRNA degradation of histone mRNAs. Understanding how histone mRNA gene expression is coupled with DNA synthesis will give a clearer picture of how the concentration of histone proteins is controlled in a cell. Also, these studies will give insight into how a cell regulates gene expression during different stages of its life (e.g. after DNA synthesis). Therefore, completion of the proposed experiments will provide a better understanding of how disruption of this type of gene expression can result in abnormal cell growth and could lead to better treatments for cancer. Specific Aim 1: To identify the region(s) and specific amino acid residue(s) of SLBP that Is required for histone mRNA degradation. I will utilize various biochemical and molecular techniques to specifically search for domains and/or amino acid residues of SLBP that are required for this function. I will also search for amino acid residues that could signal histone mRNA decay by being post-translationally modified. Specific Aim 2: To determine if SLBP functions to recruit decay proteins to a histone transcript and/or to recognize aberrant translation termination. I will Identify proteins that interact with SLBP once DNA ^ synthesis is terminated. I will then disrupt these interactions and evaluate the effect those mutant During my graduate work, I analyzed els elements that are important for translation and in determining the stability of mRNAs in the yeast Saccharomyces cerevisiae. These studies have taught me basic molecular biological techniques that are essential for studying gene expression In eukaryotes. As a postdoctoral fellow in Dr. Marzluff's lab, I plan to study the regulation of decay of histone mRNAs in human cells. The results from these studies will help me understand how expression of histone mRNAs is regulated during the cell cycle, and provide^insights into the signals that are generated in the nucleus which ultimately lead to histone mRNA degradation in the cytoplasm. Additionally, these studies will expand my knowledge to include techniques in more complex eukaryotes and in biochemistry. My ultimate career goal is to obtain a faculty position where I can develop.an independent research program. I plan to study post-transcriptional gene regulation in eukaryotes and its role in human disease. I believe that the experience that I have obtained as a graduate student, together with the training and experience I will obtain as a postdoctoral fellow, will provide me with the necessary tools to initiate my own independent program aimed at successfully understanding how eukaryotes regulate gene expression post-transcriptionally.

描述（由申请人提供）：后生动物中的组蛋白表达是一个高度调控的过程，与细胞周期中的DNA合成有关。组蛋白mRNA水平在细胞生命的大部分时间内保持在较低的基础水平，但在S期开始时，当DNA合成开始时，组蛋白mRNA水平增加约30倍。在s期结束时，由于转录本在细胞质中迅速降解，组蛋白mRNA水平下降到基础水平；然而，几乎没有证据可以解释组蛋白mrna的降解是如何被触发的。茎环结合蛋白（SLBP）是组蛋白mrna加工、输出和翻译所必需的。SLBP也被假设在组蛋白mRNA降解中发挥关键作用，通过识别或引起异常翻译终止和/或招募衰变蛋白来降解转录物。然而，SLBP在组蛋白mRNA衰变中的作用尚未得到全面评估。我假设SLBP在识别或引起异常翻译终止和/或启动组蛋白mRNA降解方面起着关键作用。了解组蛋白mRNA基因表达是如何与DNA合成相结合的，将使我们更清楚地了解细胞中组蛋白的浓度是如何控制的。此外，这些研究将深入了解细胞在其生命的不同阶段（例如DNA合成后）如何调节基因表达。因此，完成所提出的实验将更好地理解这种类型的基因表达的破坏如何导致异常细胞生长，并可能导致更好的癌症治疗。特异性目的1：确定组蛋白mRNA降解所需的SLBP区域和特定氨基酸残基。我将利用各种生物化学和分子技术专门搜索该功能所需的SLBP结构域和/或氨基酸残基。我还将寻找可能通过翻译后修饰来指示组蛋白mRNA衰变的氨基酸残基。特异性目的2：确定SLBP是否具有招募衰变蛋白到组蛋白转录物和/或识别异常翻译终止的功能。我将识别一旦DNA合成终止与SLBP相互作用的蛋白质。然后，我将破坏这些相互作用并评估这些突变的影响。在我的研究生工作中，我分析了对翻译和确定酵母菌中mrna稳定性重要的el元素。这些研究教会了我研究真核生物基因表达的基本分子生物学技术。作为Marzluff博士实验室的博士后，我计划研究人类细胞中组蛋白mrna衰变的调控。这些研究的结果将帮助我了解在细胞周期中组蛋白mRNA的表达是如何被调节的，并为细胞核中产生的最终导致组蛋白mRNA在细胞质中降解的信号提供见解。此外，这些研究将扩展我的知识，包括更复杂的真核生物和生物化学技术。我最终的职业目标是获得一个可以发展的教师职位。一个独立的研究项目。我计划研究真核生物转录后基因调控及其在人类疾病中的作用。我相信，我在研究生阶段获得的经验，以及我作为博士后将获得的训练和经验，将为我提供必要的工具，以启动我自己的独立项目，旨在成功地理解真核生物如何在转录后调节基因表达。