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Supplement: Atomic Structure of the Nuclear Pore Complex

补充：核孔复合体的原子结构

基本信息

批准号：
10705495
负责人：
Andre Hoelz
金额：
$ 9.29万
依托单位：
CALIFORNIA INSTITUTE OF TECHNOLOGY
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-09-05 至 2024-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10705495
关键词：
Accounting Address Anatomy Architecture Binding Biochemical Biogenesis Biological C-terminal Caliber Cell Cycle Cell Nucleus Cell physiology Cells Characteristics Complex Cryo-electron tomography Crystallization Cytoplasmic Filaments Disease Equipment Eukaryotic Cell Event Evolution Functional disorder Funding Gene Expression Regulation Genetic Transcription Grant Health Human Inherited Integral Membrane Protein Knowledge Life Link Maps Membrane Methodology Mitosis Molecular N-terminal Nature Nuclear Nuclear Envelope Nuclear Import Nuclear Pore Complex Nuclear Pore Complex Proteins Nuclear Structure Nucleic Acids Organelles Outcome Physiological Post-Translational Protein Processing Proteins Recombinants Research Role Structure Time Validation Work Yeasts flexibility genetic information human disease in vivo insight leukemia mRNA Export macromolecular assembly macromolecule molecular mass nucleocytoplasmic transport protein folding reconstitution reconstruction scaffold success

项目摘要

This equipment supplement does not modify the scope of the funded research but replaces vital equipment to realize the original aims in a timely manner. As such, the original abstract is included. Abstract The enclosure of genetic information in the nucleus is one of the great hallmarks of evolution, but creates the necessity for dedicated portals through which folded proteins and protein/nucleic acid complexes can cross the nuclear envelope (NE). The nuclear pore complex (NPC), a cylindrical supramolecular structure embedded in circular pores permeating the NE, is the sole gateway for passage through the NE and can accomplish the selective bidirectional transport of macromolecules of up to ~40 nm in diameter at a rate of several hundred events per second. Beyond its primary role in nucleocytoplasmic transport, the NPC also contributes to additional modes of gene regulation for example through direct interaction with the transcription and mRNA export machineries. The NPC thus represents an essential organelle for all eukaryotic life and, accordingly, NPC dysfunction has been associated with various forms of human disease. Architecturally, the NPC consists of a central symmetric core to which asymmetric components called cytoplasmic filaments and nuclear basket are attached. The NPC is built from ~34 different proteins termed nucleoporins that are each present in multiple copies such that the entire assembly reaches the extraordinary mass of ~110 MDa in humans. Nucleoporins are organized into distinct subcomplexes which constitute physiological building blocks of the intact NPC in vivo. To determine the atomic architecture of the NPC, my group has been pursuing a divide-and-conquer approach, in which we have mapped nucleoporin interactions, reconstituted recombinant nucleoporin complexes and determined their crystal structures to be fit into cryo-electron tomographic reconstructions of the intact NPC. In this way, we achieved a near-atomic composite structure of the ~60MDa human NPC symmetric core in the previous grant period. Building on this progress, we now propose to expand our structural characterization to still unresolved parts of the NPC and to use our already gained knowledge to address fundamental NPC-associated cell biological questions. Specifically, we plan to elucidate the molecular interactions in the NPC’s inner ring that are essential for the formation of its central transport channel, and between the symmetric core and transmembrane NPC components that are essential for NPC anchoring in the NE pores. The outcome of the proposed research is expected to greatly increase our understanding of the molecular mechanisms by which the NPC regulates nucleocytoplasmic transport and associated cellular processes, while simultaneously creating a mechanistic basis for currently untreatable “nup diseases.” Furthermore, the methodologies developed herein will serve as a paradigm for the characterization of other essential cellular mega-assemblies as large, flexible and complex as the NPC whose functional mechanisms have remained elusive due to lack of structural insight.

这项设备补充并不改变资助研究的范围，而是取代了至关重要的及时实现原定目标的设备。因此，原始摘要是包括在内。摘要遗传信息被包围在细胞核中是进化的最大标志之一，但需要专门的门户，折叠的蛋白质和蛋白质/核酸复合体可以穿过核膜(NE)。核孔复合体(NPC)，一种圆柱形嵌入在贯穿NE的圆形孔中的超分子结构是通过NE，并可完成大分子的选择性双向传输直径可达~40纳米，速度为每秒数百个事件。除了它的主要角色之外核质运输，鼻咽癌还有助于额外的基因调控模式例如，通过与转录和信使核糖核酸输出机制直接相互作用。全国人大就是这样代表着所有真核生物生命所必需的细胞器，因此，鼻咽癌功能障碍与各种形式的人类疾病有关。在建筑上，全国人大由一个中央被称为细胞质细丝和核篮的不对称成分的对称核心附在这里。NPC是由大约34种不同的称为核孔素的蛋白质组成的，每种蛋白质都存在于多个拷贝，使整个组装达到人类体内~110个丙二醛的非凡质量。核孔蛋白被组织成不同的亚复合体，这些亚复合体构成了活体完整的鼻咽癌。为了确定全国人大的原子结构，我的小组一直在研究一种分而治之的方法，其中我们已经绘制了核孔素相互作用图，重新构建重组核孔蛋白络合物及其低温电子结构的确定完整鼻咽癌的断层重建。通过这种方式，我们实现了一种近原子的复合材料上一次授权期内~60MDa人NPC对称核的结构。以此为基础为了取得进展，我们现在建议将我们的结构定性扩大到仍未解决的全国人大部分并利用我们已经获得的知识来研究与鼻咽癌相关的基本细胞生物学问题。具体地说，我们计划阐明NPC内环中的分子相互作用对于其中央传输通道的形成以及对称核心和跨膜鼻咽癌成分，对于鼻咽癌在东北孔洞中的锚定是必不可少的。结果是这项拟议的研究有望大大增加我们对分子的了解鼻咽癌调节核质转运及相关细胞的机制过程，同时为目前无法治疗的“nup疾病”创造一个机制基础。此外，这里开发的方法论将作为表征其他像NPC一样大、灵活和复杂的基本细胞巨型组件，其功能是由于缺乏结构性洞察力，机制仍然难以捉摸。

项目成果

期刊论文数量（14）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Günter Blobel (1936-2018).

冈特·布洛贝尔（1936-2018）。

DOI：
10.1038/s41556-018-0081-8
发表时间：
2018
期刊：
Nature cell biology
影响因子：
21.3
作者：
Hoelz,André
通讯作者：
Hoelz,André

Evidence for an evolutionary relationship between the large adaptor nucleoporin Nup192 and karyopherins.

大接头核孔蛋白 Nup192 和核传递蛋白之间进化关系的证据。

DOI：
10.1073/pnas.1311081111
发表时间：
2014
期刊：
Proceedings of the National Academy of Sciences of the United States of America
影响因子：
11.1
作者：
Stuwe,Tobias;Lin,DanielH;Collins,LeslieN;Hurt,Ed;Hoelz,André
通讯作者：
Hoelz,André

Forced entry into the nucleus.