RNA decay and processing activities of the RNA exosome

RNA 外泌体的 RNA 降解和加工活性

• 批准号: 8653583
• 负责人: AMBRO VAN HOOF
• 金额: $ 28.54万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-05-01 至 2016-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8653583
• 关键词: Active Sites; Address; Animal Model; Antiviral Agents; Catalytic Domain; Catalytic RNA; Characteristics; Cleaved cell; Collaborations; Complement; Cytoplasm; Data; Enzymes; Exonuclease; Family; Gene Expression; Genes; Genetic; Goals; Hand; Homeostasis; Human; Individual; Initiator tRNA; Messenger RNA; Modeling; Mutation; Nuclear; Orthologous Gene; Pathway interactions; Play; Poly(A) Tail; Process; Proteins; Publishing; RNA; RNA Decay; RNA Degradation; RNA Helicase; RNA Processing; Research; Ribonucleases; Ribosomal RNA; Role; Site; Specificity; Structure; Surface; Terminator Codon; Testing; Transcript; Transfer RNA; Yeasts; base; cofactor; deep sequencing; endonuclease; helicase; in vitro activity; in vivo; mRNA Decay; mRNA Transcript Degradation; mutant; nuclease; rRNA Precursor; research study

DESCRIPTION (provided by applicant): Almost all RNAs are processed from a primary transcript to a mature RNA by a variety of enzymes, including RNases. During this process many mistakes are made and aberrant RNAs generated this way are rapidly degraded to maintain RNA homeostasis. The RNA exosome plays several critical roles in this. First, the nuclear exosome processes some RNAs from longer precursors. For example, it processes the 160 nt 5.8S rRNA from a 300 nt precursor. Second, the nuclear exosome completely degrades some RNAs that are byproducts of gene expression, including the 5' external transcribed spacer that is part of the rRNA precursor. Third, the nuclear exosome degrades aberrant RNAs that fail to complete proper processing. This includes the initiator tRNA that is not properly modified. Fourth, the exosome is also present in the cytoplasm, where it performs one of two general pathways of mRNA decay. Fifth, the cytoplasmic exosome appears especially important for degrading aberrant mRNAs, including those that lack a stop codon and those that lack a poly(A) tail. Probably related to the process of degrading aberrant mRNAs is that the exosome has an antiviral activity. The long term goal of this research is to understand the role of the RNA exosome in these processing and decay pathways. The core exosome consists of ten proteins, of which Rrp44p is the catalytic subunit. We and others have previously shown that Rrp44p contains a 3' to 5' exoribonucleolytic domain and an endonucleolytic domain. We propose research to better understand the contributions of the endonuclease activity to exosome function. While Rrp44p is the only catalytic subunit of the core exosome, the other nine core subunits are also essential for viability and for all exosome functions examined. We propose experiments aimed at better understanding the role of the other nine subunits. Although the purified exosome has RNase activity in vitro, its in vivo activity requires many cofactors. While some of these cofactors are required only for specific roles of the exosome, an RNA helicase of the Ski2-like family is required for all exosome functions. Ski2p itself is required for all cytoplasmic functions of the exosome, while the closely related Mtr4p is required for all nuclear functions of the exosome. Our proposed experiments seek to clarify how these helicases assist the exosome in its many function.

描述（由申请人提供）：几乎所有 RNA 都是通过多种酶（包括 RNA 酶）从初级转录物加工成成熟 RNA。在此过程中会出现许多错误，并且以这种方式产生的异常RNA会迅速降解以维持RNA稳态。 RNA 外泌体在这方面发挥着几个关键作用。首先，核外泌体处理来自较长前体的一些 RNA。例如，它从 300 nt 前体加工 160 nt 5.8S rRNA。其次，核外泌体完全降解一些基因表达副产品的 RNA，包括作为 rRNA 前体一部分的 5' 外部转录间隔区。第三，核外泌体降解无法完成正确加工的异常RNA。这包括未正确修饰的起始 tRNA。第四，外泌体也存在于细胞质中，它在细胞质中执行 mRNA 衰减的两种一般途径之一。第五，细胞质外泌体对于降解异常 mRNA 尤其重要，包括那些缺乏终止密码子和缺乏 Poly(A) 尾的 mRNA。外泌体具有抗病毒活性，这可能与降解异常 mRNA 的过程有关。这项研究的长期目标是了解 RNA 外泌体在这些加工和衰变途径中的作用。 核心外泌体由十种蛋白质组成，其中Rrp44p是催化亚基。我们和其他人之前已经证明 Rrp44p 包含 3' 至 5' 核糖核酸外切结构域和核酸内切结构域。我们提出研究以更好地了解核酸内切酶活性对外泌体功能的贡献。 虽然 Rrp44p 是核心外泌体的唯一催化亚基，但其他九个核心亚基对于活力和所有检查的外泌体功能也至关重要。我们提出了旨在更好地理解其他九个亚基的作用的实验。 虽然纯化的外泌体在体外具有 RNase 活性，但其体内活性需要许多辅助因子。虽然其中一些辅因子仅在外泌体的特定作用中需要，但所有外泌体功能都需要 Ski2 样家族的 RNA 解旋酶。 Ski2p本身是外泌体的所有细胞质功能所必需的，而密切相关的Mtr4p是外泌体的所有核功能所必需的。我们提出的实验旨在阐明这些解旋酶如何协助外泌体发挥多种功能。