) The Nuclear Pore Complex (NPC) is a large cylindrical assembly embedded in the nuclear envelope, central for nuclear function at two related levels. First, as a regulator of transport, the NPC controls signalling access to the DNA and the passage of genetic information from DNA. Second, the NPC is an important regulator of genes by binding chromatin and its regulators to control expression states, a phenomenon that is poorly understood at the molecular level. These pivotal roles in all eukaryotic cells involve dozens of interacting pathways influencing virtually all aspects of cellular function. As a consequence, disruption of the NPC leads to many human disorders. Despite this, and though the nuclear transport machinery is a valid and powerful drug target, the NPC and the nuclear transport machinery have not been a significant part of therapeutic strategies. Arguably, there are two fundamental reasons why this is the case: (i) we do not know enough about the structure of the NPC to predict its behavior; (ii) the nuclear transport machinery impacts a bewildering array of cellular functions - thus even with a deep understanding of its structure, we still require complementary functional information to be able to predict the outcome of the targeted disruption of key elements of the transport pathway. We propose two Specific Aims that inform each other in a synergistic fashion. First, we will perform structural mapping of disease-associated Nup complexes, focusing on components of the cytoplasmic export platform and inner rings that have been linked to oncogenic and developmental defects. We will use enhanced versions of the methods we have already successfully deployed to generate high resolution maps of these two regions and their attachment sites. On completion of this study, we will have mapped most of the NPC at high precision, allowing the two regions to be seen in the context of the whole NPC assembly. Second, and in parallel, we will map the functions of disease- associated Nup complexes. We will dissect the functionalities associated with the target Nup complexes, and determine the defects associated with their alteration - testing the hypothesis that these Nups are linked to diseases because their disruption alters critical gene expression patterns in a manner distinct from other nucleoporins. Realizing these aims will generate NPC structure-function maps in unprecedented detail and which are essential to understanding how different parts of the NPC act together to determine its functionality. This project will shed light on the nature of numerous disorders associated with human NPC dysfunction;
aim ed ultimately to open the nuclear transport machinery to rational and predictive drug design.

Public Health Relevance

Nuclear pore complexes (NPCs) selectively transport material into and out of the cell?s nucleus and directly control gene expression. We still do not know much about how NPCs are constructed or how they work, therefore this project intends to understand how they mediate and control key elements of nuclear transport and regulate their associated DNA, and how defects in these processes lead to many developmental, viral or oncogenic diseases. This knowledge will ultimately open the possibility of therapeutically controlling nuclear pore complexes to correct defects that result in these diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM112108-05
Application #
9740117
Study Section
Nuclear and Cytoplasmic Structure/Function and Dynamics Study Section (NCSD)
Program Officer
Preusch, Peter
Project Start
2015-03-01
Project End
2023-04-30
Budget Start
2019-05-01
Budget End
2020-04-30
Support Year
5
Fiscal Year
2019
Total Cost
Indirect Cost
Name
Rockefeller University
Department
Biology
Type
Graduate Schools
DUNS #
071037113
City
New York
State
NY
Country
United States
Zip Code
10065
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Yoshizawa, Takuya; Ali, Rustam; Jiou, Jenny et al. (2018) Nuclear Import Receptor Inhibits Phase Separation of FUS through Binding to Multiple Sites. Cell 173:693-705.e22
Hayama, Ryo; Sparks, Samuel; Hecht, Lee M et al. (2018) Thermodynamic characterization of the multivalent interactions underlying rapid and selective translocation through the nuclear pore complex. J Biol Chem 293:4555-4563
Sparks, Samuel; Temel, Deniz B; Rout, Michael P et al. (2018) Deciphering the ""Fuzzy"" Interaction of FG Nucleoporins and Transport Factors Using Small-Angle Neutron Scattering. Structure 26:477-484.e4
Herricks, Thurston; Mast, Fred D; Li, Song et al. (2017) ODELAY: A Large-scale Method for Multi-parameter Quantification of Yeast Growth. J Vis Exp :
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Upla, Paula; Kim, Seung Joong; Sampathkumar, Parthasarathy et al. (2017) Molecular Architecture of the Major Membrane Ring Component of the Nuclear Pore Complex. Structure 25:434-445
Hayama, Ryo; Rout, Michael P; Fernandez-Martinez, Javier (2017) The nuclear pore complex core scaffold and permeability barrier: variations of a common theme. Curr Opin Cell Biol 46:110-118
Rout, Michael P; Field, Mark C (2017) The Evolution of Organellar Coat Complexes and Organization of the Eukaryotic Cell. Annu Rev Biochem 86:637-657
Rout, Michael P; Obado, Samson O; Schenkman, Sergio et al. (2017) Specialising the parasite nucleus: Pores, lamins, chromatin, and diversity. PLoS Pathog 13:e1006170

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