Eukaryotic Ribosome Assembly

PI：Arlen W约翰逊 总结 核糖体负责所有蛋白质的快速准确生产 存在于地球上所有生命形式的细胞中。这些分子机器执行 忠实的翻译依赖于它们的复杂结构，这种结构允许动态交互 配体。真核细胞中成熟的核糖体由两部分组成， 大的60 S亚基进行多肽合成，小的40 S亚基 解码mRNA。真核生物核糖体的组装涉及200多个附属物， 组装因素，其功能，在许多情况下，仍然未知。考虑 核糖体结构和功能的复杂性及其在解码我们的遗传密码中的关键作用 信息，确保其正确组装似乎是一项必要但艰巨的任务 细胞的。最近，相当大的兴趣集中在质量机制上， 控制核糖体生物合成途径。本提案侧重于两个不同的主题 （1）完成60 S的肽基转移酶中心， 亚基及其功能完整性评估机制;（2）过渡 从90年代早期的前核糖体前体到40年代前体。 本建议旨在了解质量控制机制， 评估肽基转移酶中心的功能和结构完整性， 在核糖体蛋白Rpl 10插入后，这一建议建立在我们最近的 前核糖体的原子结构的测定和两个因子Tif 6的释放 NMD 3。Nmd 3和Tif 6的释放依赖于两种GTP酶Efl 1和Efl 2。 Lsg 1，并构成60 S成熟过程中的主要质量控制检查点。在 在人类中，质量控制步骤的缺陷导致T细胞急性淋巴细胞白血病 和Shwachman-Diamond综合征 核糖体小亚基的组装涉及逐步共转录 组装90 S颗粒，一个大的蛋白质-RNA复合物，支架在U3-snoRNA上。 然而，U3的存在与U3的最终折叠结构相互不相容。 小亚基RNA，一旦RNA转录完成， 90 S粒子已经完全组装好了从90年代到40年代前的过渡很差 明白我们认为，U3被RNA解旋酶Dhr 1置换是一个重要的机制。 驱动90 S转变为40 S前粒子的主要事件。我们将 确定如何调节Dhr 1的活性以确保U3的及时释放。

PI: Arlen W Johnson Summary Ribosomes are responsible for the rapid and accurate production of all proteins in cells in all forms of life on earth. The ability of these molecular machines to carry out faithful translation depends on their complex structure that allows dynamic interaction with ligands. The mature ribosome in eukaryotic cells is composed of two parts, the large 60S subunit that carries out polypeptide synthesis and the small 40S subunit that decodes mRNA. The assembly of a eukaryotic ribosome involves over 200 accessory assembly factors, whose function, in many cases, is still unknown. Considering the complexity of ribosome structure and function and its critical role in decoding our genetic information, ensuring their correct assembly would seem a necessary but daunting task for cells. Lately, considerable interest has been focused on mechanisms of quality control in the ribosome biogenesis pathway. This proposal focuses on two distinct topics within ribosome assembly; (1) completion of the peptidyl transferase center of the 60S subunit and the mechanisms for assessing its functional integrity and (2) the transition from the early 90S pre-ribosomal precursor to the pre-40S precursor. This proposal is directed at understanding the quality control mechanisms that assess the functional and structural integrity of the peptidyl transferase center of the subunit, upon insertion of ribosomal protein Rpl10. This proposal builds on our recent determination of the atomic structure of a preribosome and release of two factors, Tif6 and Nmd3. The release of Nmd3 and Tif6 is dependent on the two GTPases Efl1 and Lsg1 and constitutes the primary quality control check point in during 60S maturation. In humans, defects in the quality control step lead to T-cell acute lymphoblastic leukemia and Shwachman-Diamond syndrome. Assembly of the small ribosomal subunit involves stepwise cotranscriptional assembly of the 90S particle, a large protein-RNA complex, scaffolded on U3-snoRNA. However, the presence of U3 is mutually incompatible with the final folded structure of small subunit RNA and must be removed once transcription of the RNA is complete and the 90S particle has fully assembled. The transition from the 90S to pre-40S is poorly understood. We propose that the displacement of U3 by the RNA helicase Dhr1 is a primary event that drives the transition of the 90S into the pre-40S particle. We will determine how the activity of Dhr1 is regulated to ensure the timely release of U3.