Regulation of Retrotransposition in S. cerevisiae

酿酒酵母逆转录转座的调控

• 批准号: 8197651
• 负责人: M Joan CURCIO
• 金额: $ 31.43万
• 依托单位: WADSWORTH CENTER
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-08-01 至 2013-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8197651
• 关键词: Acquired Immunodeficiency Syndrome; Affect; Biogenesis; Biological Assay; Biological Models; Capsid Proteins; Cells; Collection; Complement; Complementary DNA; Complex; Coupled; DNA; Defect; Degradation Pathway; Dependency; Development; Elements; Exonuclease; Family; Fungal Genome; Genetic; Genetic Transcription; Genetic Translation; Genome; Genomics; Goals; Grant; HIV-1; Holoenzymes; Homologous Gene; Human; Integration Host Factors; Long Terminal Repeats; Mediating; Messenger RNA; Molecular; Mutagens; Nature; Nonsense-Mediated Decay; Organism; Orthologous Gene; Parasites; Pathway interactions; Peptide Initiation Factors; Pharmaceutical Preparations; Play; Process; Proteins; RNA; RNA-Directed DNA Polymerase; Regulation; Retroelements; Retrotransposition; Retrotransposon; Retroviridae; Reverse Transcription; Ribonucleoproteins; Ribosomal Proteins; Ribosomes; Role; Saccharomyces cerevisiae; Saccharomycetales; Sucrose; Translational Regulation; Translational Repression; Translations; Virus; Virus-like particle; Yeasts; antiretroviral therapy; human disease; mRNA Decay; mRNA Transcript Degradation; mRNA decapping; mutant; paralogous gene; particle; pathogen; poly A specific exoribonuclease; public health relevance; research study; small molecule; success

DESCRIPTION (provided by applicant): Retroviruses and long terminal repeat (LTR) retrotransposons comprise a family of mobile elements that encode reverse transcriptase and copy their RNA genome into a cDNA that is integrated into the host genome. These prolific genomic parasites constitute a significant fraction of virtually all eukaryotic genomes. Their success as parasites relies on an array of host functions, since retroviruses and retrotransposons have small genomes and complex modes of replication. Host factors that are required for retroelement replication are potential targets for small molecules to treat AIDS, yet few have been characterized thus far. The family of active Ty1 LTR-retrotransposons in the yeast, S. cerevisiae, provides a unique model system to explore the eukaryotic host-retrotransposon relationship. Using a synthetic genetic array screen and secondary molecular screens, my lab has identified 48 retrotransposition host factors (RHFs) that are required for accumulation of Ty1 cDNA. Many of these RHFs are global regulators of translation and mRNA localization and turnover, including ribosomal protein paralogs, ribosome biogenesis factors, a paralog of translation initiation factor eIF4G, nonsense-mediated decay proteins Upf1-Upf3, the activator of decapping, Dhh1, the decapping holoenzyme, Dcp1/Dcp2 and the 5' to 3' exoribonuclease, Xrn1. The RNA genome of Ty1, like that of retroviruses, functions as a template for both translation of virus- like particle (VLP) proteins and for reverse transcription within VLPs. We have recently found that eIF4G1 and two ribosomal protein paralogs are required for accumulation of TyA, the major capsid protein, while Xrn1 influences Ty1 RNA packaging into VLPs. Moreover, we have found that Ty1 RNA is translationally repressed in sucrose-dense ribonucleoprotein particles and that TyA is associated with cytoplasmic mRNA processing bodies (P bodies), where translational regulators and mRNA decay factors are concentrated. We propose to examine the mechanism of translational regulation of Ty1 RNA and determine whether this regulation plays a role in partitioning Ty1 RNA between translation and packaging in VLPs. In addition, we will examine the role of mRNA decay proteins in the formation of VLPs that are functional for reverse transcription. The specific aims are: (1) to identify the steps in retrotransposition that are blocked in rhf mutants with reduced levels of Ty1 cDNA; (2) to examine the role of ribosomal protein paralogs, ribosome biogenesis factors and eIF4G1 in the translational regulation and localization of Ty1 RNA and protein; and, (3) to characterize Ty1 RNPs and determine whether mRNA decay factors influence Ty1 RNA or protein localization, RNA packaging in VLPs and/or reverse transcription. This project will elucidate specific mechanisms by which eukaryotic host factors promote the replication of retrotransposons and retroviruses. PUBLIC HEALTH RELEVANCE: Retroviruses are a type of infectious virus that are the known or suspected causative agents in a variety of human diseases, including AIDS. This proposal uses a simple organism, budding yeast, to illuminate the role of cellular proteins that are necessary for replication of retroviruses in human cells. Cellular proteins that are required for retroviral replication are potential targets for the development of new drugs to treat AIDS.

描述（由申请人提供）：逆转录病毒和长末端重复（LTR）逆转录转座子包括一个可移动元件家族，其编码逆转录酶并将其RNA基因组复制成整合到宿主基因组中的cDNA。这些多产的基因组寄生虫构成了几乎所有真核生物基因组的重要部分。它们作为寄生虫的成功依赖于宿主的一系列功能，因为逆转录病毒和反转录转座子具有小基因组和复杂的复制模式。逆转录因子复制所需的宿主因子是小分子治疗艾滋病的潜在靶点，但迄今为止很少有表征。酵母中活跃的Ty1 ltr -反转录转座子家族为探索真核宿主-反转录转座子关系提供了一个独特的模型系统。使用合成基因阵列筛选和二级分子筛选，我的实验室已经确定了48个逆转录转座宿主因子（rhf），这些因子是Ty1 cDNA积累所必需的。这些rhf中有许多是翻译、mRNA定位和转换的全局调控因子，包括核糖体蛋白类似物、核糖体生物发生因子、翻译起始因子eIF4G类似物、无义介导的衰变蛋白Upf1-Upf3、脱帽激活因子Dhh1、脱帽全酶Dcp1/Dcp2和5‘至3’外核糖核酸酶Xrn1。与逆转录病毒一样，Ty1的RNA基因组作为病毒样颗粒（VLP）蛋白翻译和VLP内逆转录的模板。我们最近发现eIF4G1和两个核糖体蛋白类似物是主要衣壳蛋白TyA积累所必需的，而Xrn1影响Ty1 RNA包装成VLPs。此外，我们发现Ty1 RNA在蔗糖密集核糖核蛋白颗粒中被翻译抑制，并且TyA与细胞质mRNA加工体（P体）有关，其中翻译调节因子和mRNA衰变因子集中。我们建议研究Ty1 RNA的翻译调控机制，并确定这种调控是否在VLPs中Ty1 RNA的翻译和包装之间的划分中起作用。此外，我们将研究mRNA衰变蛋白在形成具有反转录功能的VLPs中的作用。具体目的是：(1)确定Ty1 cDNA水平降低的rhf突变体中被阻断的反转录转位步骤；(2)研究核糖体蛋白类似物、核糖体生物发生因子和eIF4G1在Ty1 RNA和蛋白的翻译调控和定位中的作用；(3)表征Ty1 RNPs，确定mRNA衰变因子是否影响Ty1 RNA或蛋白质定位、RNA在VLPs中的包装和/或逆转录。本项目将阐明真核宿主因子促进逆转录转座子和逆转录病毒复制的具体机制。