GENETIC & MOLECULAR STUDIES OF NEMATODE MRNA DEGRADATION

信使RNA的稳态水平由其相对分子水平确定 合成和降解的速度。越来越明显的是，mRNA 降解是基因表达及其调控的一个重要方面。 尽管许多调节mRNA稳定性的顺式作用序列 已经被定义，但对选择性地 将消息降级。大多数信使核糖核酸的成交量似乎与 翻译。这一点在无意义突变的情况下尤其明显。 MRNAs。含有无义突变(过早终止密码子)的mRNA是 在包括人类在内的所有有机体中都不稳定。我们将对此进行研究 线虫中的一种现象，称为无意义介导的mRNA衰变 秀丽隐杆线虫。影响六种突变中的任何一种的功能丧失突变 不同的基因(SMG-1到SMG-6)消除线虫系统 无稽之谈介导的信使核糖核酸衰变。六个SMG基因定义了 毫不含糊地作用于反式，以完成特定的信使核糖核酸的转换。我们会 研究SMG基因的遗传和分子特性，并回答 以下问题：SMG基因产品是什么？SMG在哪里 位于细胞内的蛋白质？他们把什么与什么联系在一起？还有，你好吗？ 以降解为目标的无稽之谈的突变mRNAs？我们的方法结合了 遗传、分子和生化方法。特定的SMG基因将 被克隆和测序。SMG基因对动物的生存能力至关重要 将使用一种新的基因筛查进行鉴定。多克隆抗SMG抗体 抗体将被用来定位SMG蛋白的原位和分级 细胞提取液。测试特定模型的定点突变 我们将构建无意义介导的衰变的专一性。我们的结果 应该确定参与无意义介导的衰变的蛋白质 概括了解系统如何选择要 被毁了。 我们的长期目标是了解信使核糖核酸的分子机制。 营业额。在线虫中，无意义介导的腐烂可能被证明类似于 许多情况下，结构性或受调控的mRNA周转。胡说八道 信使核糖核酸衰变是一种普遍现象，可能与人类遗传有关。 疾病。有受影响基因的无意义等位基因的患者通常 表达稳定低水平的突变体mRNA。我们相信这些都是 而其他的信使核糖核酸被人类的无义介导的信使核糖核酸系统降解。 腐烂。了解线虫SMG基因将有助于 了解这些疾病的病因。

The steady-state level of a messenger RNA is established by its relative rates of synthesis and degradation. It is increasingly apparent that mRNA degradation is an important aspect of gene expression and its regulation. Although numerous cis-acting sequences that regulate mRNA stability have been defined, little is known about trans-acting factors that selectively degrade a message. Most mRNA turnover appears to be intimately coupled to translation. This is particular clear in the case of nonsense mutant mRNAs. mRNAs that contain nonsense mutations (premature stop codons) are unstable in all organisms, including humans. We will study this phenomenon, termed nonsense-mediated mRNA decay, in the nematode Caenorhabditis elegans. Loss-of-function mutations affecting any of six different genes (smg-1 through smg-6) eliminate the C. elegans system of nonsense-mediated mRNA decay. The six smg genes define proteins that unambiguously act in trans to accomplish specific mRNA turnover. We will study genetic and molecular properties of the smg genes and answer the following questions: What are the smg gene products? Where are smg proteins located in the cell? With what do they associate? And, how are nonsense mutant mRNAs targeted for degradation? Our methods combine genetic, molecular, and biochemical approaches. Specific smg genes will be cloned and sequenced. smg genes essential for viability of the animal will be identified using a novel genetic screen. Polyclonal anti-Smg antibodies will be used to locate smg proteins in situ and in fractionated cell extracts. Site-directed mutations that test specific models for the specificity of nonsense-mediated decay will be constructed. Our results should identify the proteins involved in nonsense-mediated decay and provide an outline understanding of how the system selects mRNAs to be destroyed. Our long range goals are to understand the molecular mechanisms of mRNA turnover. Nonsense-mediated decay in C. elegans may prove to be similar to many cases of constitutive or regulated mRNA turnover. Nonsense-mediated mRNA decay is a universal phenomenon and may contribute to human genetic disease. Patients having nonsense alleles of an affected gene usually express low steady-state levels of the mutant mRNA. We believe that these and other mRNAs are degraded by the human system of nonsense-mediated mRNA decay. Understanding the smg genes of C. elegans will contribute to understanding the etiology of these diseases.