Analysis of the Ribonucleases of Arabidopsis thaliana

基因的调控对于生长和发育的所有阶段都至关重要。 基因通过产生mRNA分子来发挥作用，mRNA分子将遗传信息携带到制造蛋白质的机器中，蛋白质是基因的产物。由于mRNA的量通常与产生的蛋白质的量相关，因此许多基因调控机制控制mRNA的合成或降解速率。本项目主要研究模式植物拟南芥中降解mRNA的两种核糖核酸酶。对一种类型，即5 '-3'核糖核酸外切酶的研究将主要集中在AtXRN 4的功能上。 由NSF资助的先前工作表明，AtXRN 4优先降解特定的mRNA分子，包括一个被称为miRNA的非常小的RNA靶向沉默的mRNA子集。 为了更好地理解这种特异性的基础，该项目将解决mRNA在细胞内的位置和mRNA内的序列的重要性。脱腺苷酸酶是第二种要研究的核糖核酸酶。 将检查拟南芥多聚（A）核糖核酸酶（AtPARN）和Ccr 4p去腺苷酸酶的拟南芥家族以确定它们优先降解哪些mRNA。这将通过使用缺乏一种或多种这些去腺苷酸酶的突变植物进行生物化学和分子生物学实验来实现，例如DNA微阵列分析，一种用于同时测定数千个基因的mRNA水平的技术。 这项工作将在植物领域之外产生广泛的影响，因为类似的问题尚未在其他多细胞生物中得到回答。特别是，涉及miRNA的基因调控机制对植物和动物都至关重要。 因此，理解AtXRN 4降解选定的miRNA靶标的原因对许多系统中的XRN同源物具有意义。类似地，PARN和多个Ccr 4p同源物不存在于酵母中，其中已经完成了许多关于mRNA降解的开创性工作，因此这些去腺苷酸酶的工作对其他真核生物也具有广泛的意义。 除了对基础知识的贡献外，该项目还将产生数据、突变体和其他工具，供公众使用。该项目的实施将有助于包括本科生和邻近机构的年轻科学家在内的各种年轻科学家的教育和职业发展。

The regulation of genes is of fundamental importance for all phases of growth and development. Genes function by producing mRNA molecules that carry genetic information to the machinery that makes proteins, which are the products of genes. Because the amount of mRNA is generally related to the amount of protein produced, many gene regulatory mechanisms control the synthesis or degradation rates of mRNAs. This project focuses on two types of ribonucleases (RNases) that degrade mRNA in the model plant Arabidopsis. Work on one type, the 5'-3' exoribonucleases, will concentrate primarily on the function of AtXRN4. Previous work funded by NSF showed that AtXRN4 preferentially degrades specific mRNA molecules, including a subset of the mRNAs targeted for silencing by very small RNAs called miRNAs. To better understand the basis of this specificity, this project will address the importance of mRNA location within the cell and of sequences within mRNAs. Deadenylases are the second type of RNase to be studied. Both Arabidopsis Poly(A) Ribonuclease (AtPARN) and the Arabidopsis family of Ccr4p deadenylases will be examined to determine which mRNAs they preferentially degrade. This will be accomplished by using mutant plants lacking one or more of these deadenylases for biochemical and molecular biological experiments such as DNA microarray analysis, a technology for determining mRNA levels for thousands of genes at once. This work will have broad impact beyond the plant field because similar questions have yet to be answered in other multicellular organisms. In particular, gene regulatory mechanisms involving miRNA are critical for both plants and animals. Therefore, understanding why AtXRN4 degrades selected miRNA targets has implications for XRN homologs in many systems. Similarly, PARN and multiple Ccr4p homologs are not present in yeast, in which much of the pioneering work on mRNA degradation has been done, so work on these deadenylases should also be of broad significance for other eukaryotes. Beyond its contributions to basic knowledge, this project will generate data, mutants and other tools that will be made publicly available. Execution of this project will contribute to the education and career development of diverse young scientists including undergraduates and those from a neighboring institution.