Genetic & Molecular Studies of Nematode mRNA Degradation

描述（由申请人提供）：我们研究的广泛目标是了解通过转录后过程调节真核生物基因表达的分子机制和生理后果。大量重要的生物学调控发生在转录后，但许多机制尚不清楚。我们将研究选择性mRNA降解，即特定mRNA从细胞中主动消除的过程。mrna的调控周转深刻地影响着基因表达，我们的主要目标是了解这种调控的机制和后果。我们关注的是含有过早翻译终止密码子的mrna的加速降解。含有过早终止密码子的信使rna比野生型的信使rna降解得更快，这种现象被称为“无义介导的mRNA衰变”（NMD）。NMD发生在所有被测试的真核生物中，其作用对人类遗传疾病有重大影响。在所有导致人类遗传疾病的已测序点突变中，大约有三分之一是无义突变或移码突变，其中大多数已知或预测会引起突变mRNA的NMD。我们将研究秀丽隐杆线虫NMD所需的7个基因（smg-1至smg-7），并研究它们的遗传和分子特性。我们的工作分为三大类：(1)研究NMD的分子机制；(2)研究NMD与RNA干扰的关系；(3)研究NMD在野生型动物中的生物学功能。我们的具体目标是：

DESCRIPTION (provided by applicant): The broad aims of our research are to understand the molecular mechanisms and physiological consequences of regulating eukaryotic gene expression by post-transcriptional processes. A large amount of important biological regulation occurs post-transcriptionally, but many of the mechanisms are not well understood. We will study selective mRNA degradation, the processes by which specific mRNAs are actively eliminated from cells. Regulated turnover of mRNAs profoundly influences gene expression, and our broad goals are to understand the mechanisms and consequences of that regulation. We focus on the accelerated degradation of mRNAs that contain premature translation termination codons. Messenger RNAs that contain premature stop codons are degraded more rapidly than their wild-type counterparts, a phenomenon termed "nonsense-mediated mRNA decay" (NMD). NMD occurs in all eukaryotes tested, and its action substantially impacts human genetic disease. Approximately one third of all sequenced point mutations that cause human genetic disease are nonsense or frameshift mutations, and most of these are known or predicted to elicit NMD of the mutant mRNA. We will study seven C. elegans genes required for NMD (smg-1 through smg-7) and will investigate their genetic and molecular properties. Our work falls into three broad categories: (1) investigating the molecular mechanisms of NMD; (2) investigating the relationship of NMD to RNA interference; and (3) investigating the biological function of NMD in wild-type animals. Our Specific Aims are: 1. We will investigate the structure of mRNPs containing premature translation termination codons. Such mRNPs differ from those not containing premature stop codons by selectively binding the SMG-2 protein. 2. We will study the roles of SMG-2 phosphorylation and dephosphorylation in NMD. 3. We will investigate a recently discovered relationship between NMD and RNAi. We will test whether NMD contributes to gene silencing via endogenous RNAi. 4. We will identify most or all of the endogenous mRNA targets of NMD. Our long-range goals are to understand both the molecular mechanisms of mRNA turnover and the biological roles of NMD. Our experiments contribute to these goals by identifying proteins that are required for NMD, by describing their biochemical activities in vivo, and by investigating the role of NMD during normal growth and development.