Transitions: Mechanistic insights into ribosome rescue and quality control in the Archaea

细胞根据mRNA分子的编码，使用一种叫做翻译的过程来合成蛋白质。翻译代码时的错误会造成许多有害的影响。核糖体是细胞的纳米机器，翻译mRNA代码并合成蛋白质。核糖体不断被使用，必须多次重复使用，这种重复使用有两个必要的过程。首先，在一个mRNA的翻译结束后，有一个被称为核糖体再循环的活跃过程，将核糖体从mRNA中分离出来，使其可以重复使用。其次，有时核糖体在转译过程中失速甚至发生碰撞，无法自行重启；在这种情况下，有一个活跃的过程将mRNA从被破坏的核糖体中分离出来，这样核糖体就可以被重复使用。这个研究项目将调查最不为人所知的生命领域，即古细菌，如何完成核糖体的拯救和回收。该项目还将为研究生和巴尔的摩市公立学校的高中生提供培训。为了进行科学调查，研究小组将使用嗜盐古菌（Haloferax volcanii），这是一种适合遗传和生化调查的嗜盐古菌。在目标1中，研究人员将确定识别停滞核糖体并促进古生菌核糖体分裂活性的蛋白质因子。他们将使用遗传和生化方法来发现目前在古细菌中未知的新因素。在目标2中，研究人员将确定古生菌中核糖体碰撞的景象。核糖体碰撞产生核糖体二聚体；二体保护片段的深度测序将提供碰撞基序的全基因组特征，并能够在体内测试核糖体拯救中感兴趣的新因素的作用。在目标3中，研究人员将确定由停滞核糖体产生的多肽的蛋白水解靶向机制。这些实验将提供古细菌核糖体相关质量控制（RQC）途径的初步知识。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

Cells use a process called translation to synthesize proteins according to the code in an mRNA molecule. Mistakes in translating the code can cause many deleterious effects. Ribosomes are the cellular nanomachines that translate the mRNA code and synthesize proteins. Ribosomes are in constant use and must be reused many times and there are two processes necessary for this reuse. First, after translation of one mRNA concludes, there is an active process called ribosome recycling that separates the ribosome from the mRNA so that it can be reused. Second, sometimes ribosomes stall or even collide during translation and cannot restart on their own; again in this case there is an active process that separates the mRNA from the disrupted ribosomes so that the ribosomes can be reused. This research project will investigate how the least-understood domain of life, known as the archaea, accomplish ribosome rescue and recycling. The project will also provide training for graduate students and include high school students from the Baltimore City public schools in the research.To do the scientific investigations, the research team will use Haloferax volcanii, a halophilic archaeon that is amenable to genetic and biochemical investigation. In aim 1, the researchers will identify protein factors that recognize stalled ribosomes and promote ribosome splitting activity in archaea. They will use genetic and biochemical approaches to discover novel factors that are currently unknown in the archaea. In aim 2, the researchers will determine the landscape of ribosome collisions in archaea. Ribosome collisions result in ribosome dimers called disomes; deep-sequencing of disome-protected fragments will provide a genome-wide characterization of collision motifs and the ability to test the role of novel factors of interest in ribosome rescue, in vivo. In aim 3, the researchers will identify the mechanisms for proteolytic targeting of polypeptides produced by stalled ribosomes. These experiments will provide first knowledge of Ribosome-associated Quality Control (RQC) pathways in archaea.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.