ASSEMBLY, LOCALIZATION AND FUNCTION OF THE U3 SNRNP

描述（改编自申请人的摘要）：核糖体生物发生是 对真核细胞在细胞周期中的进展， 大小，并划分。这项应用的重点是核糖体RNA的加工 （rRNA），目标是识别和研究关键因素的功能 参与前体rRNA的切割，导致18 S rRNA。利用酵母S.酿酒酵母作为一个模型系统，拟议的工作将 重点关注U3小核仁核糖核蛋白（snoRNP）的功能， 它的亚基在这些切割步骤中。拟议的研究很重要 因为U3 snoRNP在真核生物中普遍存在， 在迄今为止研究的所有生物体中新生rRNA转录物的加工。在 酵母，三个U3 snoRNP依赖性切割步骤，导致释放 18 SrRNA通过两种不同U3 snoRNA：pre-rRNA碱基配对产生 交互.关于蛋白质成分的功能知之甚少， U3 snoRNP在这些RNA-RNA相互作用中或如果蛋白质-蛋白质和 蛋白质-RNA相互作用对于功能是必需的。许多预测的U3 snoRNP蛋白组分尚未鉴定。初步结果 研究人员的实验室发现了几种新的 U3 snoRNP组分：Mpp 10 p、Imp3 p和Imp 4p。他们的性格特征 提供了大量证据，指出了 snoRNP功能中的蛋白质-蛋白质、蛋白质-RNA和RNA-RNA相互作用。旨在 2、3和4将测试关于这些相互作用的特定假设， 研究prerRNA加工中每种相互作用的重要性。这将是 通过不同的遗传方法，包括突变分析， 蛋白质和RNA成分。在目标1中，研究人员将继续她的 有希望的初步努力，以纯化U3 snoRNP，并确定其余的 使用亲和捕获随后进行质量分析的未知蛋白质组分 光谱法在目标5中，他们建议利用一些 他们已经产生的MPP 10、JMP 3和IMP 4的条件等位基因 建立一系列的基因筛选这些屏幕的设计目的是 鉴定在前体rRNA加工的相同途径中起作用的新因子 或以某种方式调节它。总的来说，这项工作将 导致对这些机制的全面理解 U3 snoRNP介导的RNA加工事件。

DESCRIPTION (Adapted from the applicant's abstract): Ribosome biogenesis is essential for eukaryotic cells to progress through the cell cycle, grow in size, and divide. This application focuses on the processing of ribosomal RNA (rRNA), and the goal is to identify and study the function of critical factors involved in the cleavage of the pre-rRNAs that lead to the formation of the 18S rRNA. Using the yeast S. cerevisiae as a model system, the proposed work will focus on the function of the U3 small nucleolar ribonucleoprotein (snoRNP) and its subunits in these cleavage steps. The proposed research is important because the U3 snoRNP is ubiquitous in eukaryotes and is required for processing of the nascent rRNA transcript in all organisms studied so far. In yeast, the three U3 snoRNPdependent cleavage steps that lead to the release of the 1 8S rRNA occur via two different U3 snoRNA:pre-rRNA base-pairing interactions. Little is known about the function of the protein components of the U3 snoRNP in these RNA-RNA interactions or if protein-protein and protein-RNA interactions are essential for function. Many of the predicted U3 snoRNP protein components have not yet been identified. Preliminary results from the investigator's laboratory have led to the discovery of several novel U3 snoRNP components: MpplOp, Imp3p, and Imp4p. Their characterization has provided substantial evidence pointing to the critical nature of protein-protein, protein-RNA, and RNA-RNA interactions in snoRNP function. Aims 2, 3, and 4 will test specific hypotheses regarding these interactions and will examine the importance of each interaction in prerRNA processing. This will be accomplished by varied genetic approaches, including mutational analysis of both protein and RNA components. In Aim 1, the investigator will continue her promising initial efforts to purify the U3 snoRNP and to identify the remaining unknown protein components using affinity capture followed by mass spectrometry. In Aim 5, they propose to take advantage of a number of conditional alleles of MPP1O, JMP3, and IMP4 that they have already generated to set up a selected series of genetic screens. These screens are designed to identify new factors that function in the same pathway for pre-rRNA processing as the U3 snoRNP or regulate it in some manner. Taken together, this work will lead to a comprehensive understanding of the mechanisms underlying these essential U3 snoRNP-mediated RNA processing events.