Nuclear pore complexes and nucleocytoplasmic transport - Methods / edited by Val�erie Doye.

Description based upon print version of record.

Front Cover; Nuclear Pore Complexes and Nucleocytoplasmic Transport -- Methods; Copyright; Contents; Contributors; Preface; Chapter 1: Fifty Years of Nuclear Pores and Nucleocytoplasmic Transport Studies: Multiple Tools Revealing Complex Rules; Introduction; 1.1. The NPCs: A Modular Macromolecular Assembly; 1.1.1. Toward a refined view of the NPC structure; 1.1.2. Nucleoporins: The building blocks of NPCs; 1.1.3. Integrating the Nups into the 3D architecture of the NPC; 1.1.3.1. Nup localization; 1.1.3.2. Interactions among Nups; 1.1.3.3. Nup stoichiometry

1.1.3.4. Toward a detailed map of NPCs1.1.4. Nups are composed of a limited set of structural domains; 1.2. Nucleocytoplasmic Trafficking: The Rules of the Road; 1.2.1. Investigating nucleocytoplasmic transport; 1.2.1.1. Historical overview; 1.2.1.2. Expanding the toolbox; 1.2.2. The signals for nucleocytoplasmic exchanges; 1.2.3. A family of protein transport receptors: The karyopherins; 1.2.4. The Ran GTPase: A key to transport directionality; 1.2.5. The case of INM targeting; 1.2.6. Distinct pathways contribute to RNA export

1.2.7. Translocation across the NPCs: A dual function for FG-Nups as barrier and gate1.2.8. Noncanonical transport pathways through the NE; 1.3. The Nuclear Transport Machinery: A Dynamic and Versatile Device; 1.3.1. NPC biogenesis throughout the cell cycle; 1.3.1.1. NPC disassembly; 1.3.1.2. Post-mitotic NPC assembly; 1.3.1.3. De novo NPC assembly; 1.3.2. Multiple functions of the nuclear transport machinery during the cell cycle; 1.3.3. NPCs, nuclear organization, and gene expression; 1.3.4. NPCs and genetic stability; Concluding Remarks; Acknowledgments; References

Chapter 2: Imaging Metazoan Nuclear Pore Complexes by Field Emission Scanning Electron MicroscopyIntroduction; Prehistoric landmarks (before EM): From cells to the nuclear envelope; The EM era: From the nuclear envelope to nuclear pore complexes; High resolution SEM: Direct surface imaging of NPCs; 2.1. Rationale; 2.1.1. Purpose; 2.1.2. An overview of sample preparation; 2.1.2.1. Fixation; 2.1.2.2. Dehydration and critical point drying; 2.1.2.3. Sputter coating; 2.2. Materials; 2.2.1. Equipment; 2.2.2. Materials; 2.2.3. Reagents; 2.2.4. Buffers; 2.3. Anchored Nuclei

2.3.1. Coating silicon chips2.3.2. Chromatin decondensation and attachment; 2.3.3. In vitro assembly reaction; 2.4. Mammalian Cell Nuclei; 2.5. Immunogold Labeling; 2.6. Sample Preparation for FESEM; 2.6.1. Fixation, dehydration, and CPD; 2.6.2. Sputter coating and FESEM analysis; Acknowledgments; References; Chapter 3: Imaging Yeast NPCs: From Classical Electron Microscopy to Immuno-SEM; Introduction; 3.1. Conventional TEM; 3.1.1. Materials; 3.1.1.1. Equipment; 3.1.1.2. Spheroplast preparation; 3.1.1.3. Fixation, dehydration, contrasting, and embedding; 3.1.1.4. Sectioning and poststaining

3.1.2. Procedure

Volume 122 of Methods in Cell Biology describes modern tools and techniques used to study nuclear pore complexes and nucleocytoplasmic transport in diverse eukaryotic model systems (including mammalian cells, Xenopus, C. elegans, yeast). The volume enables investigators to analyze nuclear pore complex structure, assembly, and dynamics; to evaluate protein and RNA trafficking through the nuclear envelope; and to design in vivo or in vitro assays appropriate to their research needs. Beyond the study of nuclear pores and transport as such, these protocols will also.