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The Ran GTPase cycle is required for yeast nuclear pore complex assembly
Kathryn J Ryan1, J Michael McCaffery, Susan R Wente
1Department of Cell and Developmental Biology, Vanderbilt University Medical Center, Nashville, TN 37232, USA.
The Journal of Cell Biology
|March 26, 2003
Summary
The Ran GTPase cycle is essential for nuclear pore complex (NPC) assembly in yeast. Genetic studies reveal that defects in Ran and its regulators disrupt NPC formation, leading to membrane abnormalities and vesicle accumulation.
Area of Science:
- Cell Biology
- Molecular Biology
- Genetics
Background:
- Nuclear pore complexes (NPCs) are crucial for nucleocytoplasmic transport.
- The Ran GTPase cycle regulates various nuclear processes, but its role in NPC assembly was unknown.
Purpose of the Study:
- To investigate the involvement of the Ran GTPase cycle in the de novo assembly of NPCs.
- To identify genetic factors essential for NPC biogenesis in Saccharomyces cerevisiae.
Main Methods:
- Genetic screening in yeast to identify mutants defective in NPC assembly.
- Localization studies using GFP-tagged nucleoporins (nups) and Pom152.
- Electron microscopy and biochemical fractionation to analyze mutant phenotypes and vesicle contents.
Main Results:
- Mutations in Ran and its regulators (GTPase-activating protein, guanine nucleotide exchange factor, GDP import factor) caused temperature-dependent mislocalization of nups and Pom152.
- Defects led to reduced nuclear envelope-associated GFP fluorescence and increased cytoplasmic signal.
- Electron microscopy revealed membrane perturbations and the accumulation of nup-containing vesicles in arrested mutants.
- NPC stability was unaffected, and defects depended on de novo protein synthesis.
Conclusions:
- The Ran GTPase cycle is required for efficient nuclear pore complex assembly.
- A model is proposed where Ran-mediated vesicular fusion is a critical step in integrating NPCs into the nuclear envelope.