Regulation of spliceosomal ATPase activity

剪接体 ATP 酶活性的调节

• 批准号: 6848434
• 负责人: GRETCHEN M EDWALDS-GILBERT
• 金额: $ 17.44万
• 依托单位: CLAREMONT MC KENNA COLLEGE
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-01-01 至 2008-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6848434
• 关键词: RNA splicing; SDS polyacrylamide gel electrophoresis; adenosinetriphosphatase; enzyme activity; eukaryote; fungal genetics; gene expression; introns; messenger RNA; polymerase chain reaction; spliceosomes; yeast two hybrid system

DESCRIPTION (provided by applicant): An essential step in eukaryotic gene expression is pre-mRNA splicing, which involves the removal of introns and ligation of exons. Mutations that cause errors in splicing are found in at least 15% of human genetic diseases; additional diseases are caused by mutations in trans-acting splicing factors. Members of the DExD/H family of nucleic acid-dependent ATPases play essential roles in splicing; other members of this protein family participate in all aspects of cellular activities involving nucleic acids, including DNA replication, recombination, transcription, mRNA transport, translation, and RNA processing. Residues in DExD/H box proteins that are essential for function have been characterized; however, much less is known about how the proteins act in vivo.The goal of this AREA proposal is to identify and characterize co-factors that modulate the activity of one member of this protein family, Prp43. Prp43 is encoded by an essential gene in yeast and has homologues in other eukaryotes, including humans. Although highly conserved with other spliceosomal ATPases, Prp43 may participate in cellular pathways in addition to splicing. Its target(s) and how it achieves specificity within the spliceosome and other complexes in which it has been identified is not known. Clearly, to fully understand the activity and function of Prp43 and other DExD/H proteins, the cofactors that interact with them in vivo must be identified and characterized. We plan to use biochemical and genetic approaches to: 1) Identify the spliceosome interaction domain(s) of Prp43 2) Identify and characterize potential cofactors for Prp43 Understanding the target of Prp43 RNA-dependent ATPase activity within the spliceosome and other cellular complexes will assist in understanding how the human homologue of Prp43, as well as related DExD/H-box proteins, achieve target specificity in vivo.

描述（由申请人提供）：真核基因表达的一个重要步骤是前mRNA剪接，其涉及内含子的去除和外显子的连接。 至少 15% 的人类遗传疾病中发现了导致剪接错误的突变；其他疾病是由反式剪接因子的突变引起的。核酸依赖性 ATP 酶 DExD/H 家族的成员在剪接中发挥重要作用；该蛋白质家族的其他成员参与涉及核酸的细胞活动的各个方面，包括 DNA 复制、重组、转录、mRNA 运输、翻译和 RNA 加工。 DExD/H 盒蛋白中对功能至关重要的残基已得到表征；然而，人们对这些蛋白质如何在体内发挥作用知之甚少。该 AREA 提案的目标是识别和表征调节该蛋白质家族成员 Prp43 活性的辅助因子。 Prp43 由酵母中的一个必需基因编码，并且在包括人类在内的其他真核生物中具有同源物。尽管与其他剪接体 ATP 酶高度保守，Prp43 除了剪接之外还可能参与细胞途径。它的目标以及它如何在剪接体和其他复合物中实现特异性尚不清楚。显然，为了充分了解 Prp43 和其他 DExD/H 蛋白的活性和功能，必须鉴定和表征在体内与它们相互作用的辅因子。我们计划使用生化和遗传学方法来： 1) 识别 Prp43 的剪接体相互作用域 2) 识别和表征 Prp43 的潜在辅因子 了解剪接体和其他细胞复合物中 Prp43 RNA 依赖性 ATP 酶活性的靶标将有助于了解 Prp43 的人类同源物以及相关的 DExD/H-box 蛋白如何在体内实现靶标特异性。