The role of nucleolar enzymes in yeast ribosome biogenesis

核仁酶在酵母核糖体生物发生中的作用

• 批准号: RGPIN-2014-04053
• 负责人: Dragon, François
• 金额: $ 2.55万
• 依托单位: Université du Québec à Montréal
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2018
• 资助国家: 加拿大
• 起止时间: 2018-01-01 至 2019-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=649573
• 关键词: role; nucleolar; enzymes; yeast; ribosome

Ribosomes are the organelles that synthesize proteins in all living organisms. They are formed by the association of two subunits (a small and a large) that are constituted of large RNA molecules and multiple proteins. The key components of ribosomes are the ribosomal RNAs (rRNAs) as they are directly involved in the different steps of protein synthesis. The biogenesis of ribosomes is a major cellular process, and it is a prerequisite for cells to grow in size and proliferate. In eukaryotes, this process takes place in the nucleolus, a prominent compartment of the cell nucleus. The majority of nucleolar factors actually function at various steps of the intricate pathway that leads to the production of mature rRNAs.*My research program focuses on ribosome biogenesis in the yeast Saccharomyces cerevisiae, the best studied eukaryotic model system in the field. Work in my laboratory contributes to defining the molecular mechanisms involving nucleolar factors that participate in the production of mature rRNAs. In particular, we focus on the cleavage reactions that remove extra sequences from precursor rRNA molecules. My lab is using yeast as a model system to take advantage of its well-characterized genetic, biochemical and molecular biology tools. Our objective for the next five years is to define the role of three nucleolar enzymes: Dbp4, Kre33 and Nop38.**Dbp4 is an RNA helicase that acts as a molecular motor to rearrange RNA-RNA (or possibly RNA-protein) interactions during the pre-rRNA cleavage steps. Importantly, Dbp4 is associated with other nucleolar factors that are conserved among species: all are essential for growth, indicating that they perform crucial cellular functions. Human homologues of these proteins are involved in embryonic development and cancer (the human homologue of Dbp4, DDX10, has been identified as a "cancer gene").**Kre33 is a putative RNA acetyltransferase that is conserved from bacteria to humans. This high degree of conservation suggests it has a critical cellular function but almost nothing is known about the role of Kre33 in yeast. My lab identified specific features of Kre33 that distinguish it from its bacterial ancestors, and we suspect these are key elements for the function of Kre33 in yeast and humans. To determine how Kre33 functions at the molecular level we will combine genetic and biochemical approaches. These investigations are fundamental to understand why Kre33 is essential for cell survival.**Nop38 is a putative RNA methyltransferase that is highly conserved but not as much as Kre33. Nop38 is present in yeasts, plants, animals but it is not found in all bacteria, only those termed "extremophiles", which live in very hostile environments such as oceanic thermal vents; "normal" bacteria, like those that live in the guts of animals, do not have Nop38. We are very excited to work on this enzyme because nothing is known about its function, and everything remains to be discovered.**Our research on Dbp4, Kre33 and Nop38 is important to understand how these proteins function in the cell: elucidation of molecular mechanisms underlying ribosome biogenesis in eukaryotes could open new avenues to improve the growth of plants, to treat diseases or to target eukaryotic pathogens that affect humans and other animals.

核糖体是所有生物体中合成蛋白质的细胞器。它们是由两个亚基（一个小的和一个大的）组成的，这两个亚基由大RNA分子和多种蛋白质组成。核糖体的关键成分是核糖体RNA（rRNA），因为它们直接参与蛋白质合成的不同步骤。核糖体的生物发生是一个主要的细胞过程，是细胞生长和增殖的先决条件。在真核生物中，这一过程发生在核仁中，核仁是细胞核的一个突出部分。大多数核仁因子实际上在导致成熟rRNA产生的复杂途径的各个步骤中发挥作用。我的研究项目主要集中在酵母酿酒酵母中的核糖体生物合成，这是该领域研究最好的真核模型系统。在我的实验室工作有助于定义的分子机制，涉及核仁因子参与成熟rRNA的生产。特别是，我们专注于从前体rRNA分子中去除额外序列的切割反应。我的实验室正在使用酵母作为模型系统，以利用其良好的特征遗传，生物化学和分子生物学工具。我们未来五年的目标是确定三种核仁酶的作用：Dbp 4，Kre 33和Nop 38。Dbp 4是一种RNA解旋酶，在pre-rRNA切割步骤中作为分子马达重排RNA-RNA（或可能的RNA-蛋白质）相互作用。重要的是，Dbp 4与物种间保守的其他核仁因子相关：所有这些因子都是生长所必需的，表明它们执行关键的细胞功能。这些蛋白质的人类同源物参与胚胎发育和癌症（Dbp 4的人类同源物DDX 10已被鉴定为“癌症基因”）。Kre 33是一种假定的RNA乙酰转移酶，从细菌到人类都是保守的。这种高度的保守性表明它具有重要的细胞功能，但几乎没有人知道Kre 33在酵母中的作用。我的实验室确定了Kre 33与其细菌祖先的特定特征，我们怀疑这些是Kre 33在酵母和人类中功能的关键要素。为了确定Kre 33在分子水平上的功能，我们将联合收割机遗传和生物化学方法相结合。这些研究对于理解为什么Kre 33对细胞存活至关重要至关重要。Nop 38是一种推定的RNA甲基转移酶，其高度保守，但不如Kre 33保守。Nop 38存在于酵母、植物、动物中，但并非在所有细菌中发现，只有那些被称为“极端微生物”的细菌，它们生活在非常恶劣的环境中，如海洋热喷口;“正常”细菌，如生活在动物肠道中的细菌，不具有Nop 38。我们对这种酶的研究感到非常兴奋，因为对它的功能一无所知，一切都有待发现。我们对Dbp 4，Kre 33和Nop 38的研究对于了解这些蛋白质在细胞中的功能非常重要：阐明真核生物中核糖体生物发生的分子机制可以为改善植物生长，治疗疾病或靶向影响人类和其他动物的真核病原体开辟新途径。