A Molecular Genetic Analysis of Transcription Termination in Yeast

该项目的长期目标是了解真核生物转录终止的机制。有充分的证据表明，前mrna的切割与转录终止有关。然而，终止反应的许多方面仍然不明确，特别是新生转录物释放的机制和参与3'端形成的新因子的鉴定。因此，本研究的目的是研究RNA加工和转录终止协调形成功能性mRNA 3'端的机制。采用酵母模型系统，利用分子遗传学和生物化学相结合的方法。GRS1是一种假定的甘酰tRNA合成酶，在筛选参与3'端形成的基因时被鉴定出来。携带grsi1-1突变的菌株在野生型水平的体外切割和聚腺苷化前mrna，但表现出增加的聚腺苷化信号的读通转录。因此，假设是Grslp促进有效的转录终止。为了验证这一点，实验正在进行：a.检验Grs1p在终止反应中的作用；b.开展Grs1p的结构/功能研究，以确定负责3'端形成的相同结构域是否对tRNA酰化很重要。基因调控和表达的机制是复杂的。由于基因转录在人类生物学和疾病机制中起着至关重要的作用，这一领域正受到密切的研究。转录反应的一个组成部分还没有被很好地理解，它与从DNA模板合成RNA的酶如何结束反应有关。这项研究描述了一个新的发现，即终止反应的一个组成部分也参与了其他细胞功能。这很重要，因为它提供了细胞中两个基本过程之间的联系，RNA的合成和蛋白质的合成。因此，这项研究的意义在于深入了解细胞如何表达个体基因，以及这一过程如何与其他细胞功能相关

Hyman The long term objective of the project is to understand the mechanism of transcription teruiination in eukaryotes. It is well documented that cleavage of pre-mRNA is coupled to transcription termination. However, many aspects of the termination reaction remain ill-defined, in particular, the mechanism of nascent transcript release and the identification of novel factors that participate in 3' end formation. Thus, the objective of this research is to examine mechanism(s) by which RNA processing and transcription termination coordinate formation of a functional mRNA 3' end. The yeast model system was employed to take advantage of combined molecular genetic and biochemical approaches. GRS1, a putative glycyl tRNA synthetase, was identified in a screen for genes involved in 3' end formation. Strains carrying a grsi1-1 mutation cleave and polyadenylate pre-mRNAs in vitro at wild type levels but exhibit increased readthrough transcription of a polyadenylation signal. Thus, the hypothesis is that Grslp promotes efficient transcription termination. To test this, experiments are underway to: a. examine the role of Grs1p in the termination reaction; b. carry out structure/function studies of Grs1p to determine if the same domains responsible for 3' end formation are important for tRNA acylation. The mechanism by which genes are regulated and expressed is complex. This area is under intense investigation as gene transcription is critical for human biology and disease mechanisms. A component of the transcription reaction that is not well understood deals with how the enzyme that synthesizes the RNA from the DNA template ends the reaction. This research describes the novel finding that a component of the termination reaction is also involved in other cellular functions. This is important as it provides a connection between two fundamental processes in cells, the synthesis of RNA and the synthesis of proteins. Thus, the significance of the research is to provide insight into how cells express individual genes and how this process is related to other cellular functions