Biochemistry of Small Nucleolar RNAs in Yeast

核仁含有许多小的核仁rna -蛋白复合物（snoRNPs），它们修饰rRNA中的核苷酸（大多数snoRNPs都这样做）或参与prerna的加工（切割）。修饰的snoRNPs形成2'- o -甲基化核苷酸和假尿嘧啶，主要在核糖体的功能重要区域。这些变化的影响实际上是未知的。关于处理snornp的主要未解决的问题是它们的实际作用。该提案有两个主要目的——旨在回答这些重要问题。两者都建立在前一笔拨款的基础上。首先，我们发现核糖体反应中心区域的核苷酸修饰的系统损耗会导致翻译缺陷。其次，我们亲和分离了18S rRNA加工和甲基化所需的保守snoRNP (U14)；

DESCRIPTION (provided by applicant) The nucleolus contains scores of small nucleolar RNA-protein complexes (snoRNPs), which modify nucleotides in rRNA (most snoRNPs do this) or participate in processing (cleavage) of prerRNA. The modifying snoRNPs form 2'-O-methylated nucleotides and pseudouridine, mostly in functionally important regions of the ribosome. The effects of these alterations are virtually unknown. The main unanswered question with the processing snoRNPs is their actual role. The proposal has two main aims -designed to answer these important questions. Both build on results from the previous grant. First, we showed that systematic depletion of nucleotide modifications from the reaction center region of the ribosome causes defects in translation. Second, we have affinity-isolated a conserved snoRNP (U14) required for processing and methylation of 18S rRNA, and identified several novel proteins, including factors already tied to ribosome synthesis. The major aims of our application are: 1) To determine the influence of nucleotide modifications in the mRNA decoding center and two other domains that interact dynamically with tRNA in the decoding region, and; 2) To determine the roles of the U14 snoRNP proteins in rRNA processing and modification. Effects of nucleotide modification will be investigated by depleting selected modifications from the target regions and examining ribosome synthesis and function. Studies of the U14 snoRNP will focus initially on defining the roles of the new snoRNP components in ribosome synthesis, by genetic depletion, mutation and affinity isolation strategies. Other goals include describing the assembly, structure and activity of the snoRNP complex. Results from the studies will yield valuable new insights into ribosome synthesis and function, and could shed light on the basis of established links between snoRNPs, aging and cancer.