GTP hydrolysis by Ran is required for nuclear envelope assembly.
Nuclear formation in Xenopus egg extracts requires cytosol and is inhibited by GTP gamma S, indicating a requirement for GTPase activity. Nuclear envelope (NE) vesicle fusion is extensively inhibited by GTP gamma S and two mutant forms of the Ran GTPase, Q69L and T24N. Depletion of either Ran or RCC1, the exchange factor for Ran, from the assembly reaction also inhibits this step of NE formation. Ran depletion can be complemented by the addition of Ran loaded with either GTP or GDP but not with GTP gamma S. RCC1 depletion is only complemented by RCC1 itself or by RanGTP. Thus, generation of RanGTP by RCC1 and GTP hydrolysis by Ran are both required for the extensive membrane fusion events that lead to NE formation.
European Molecular Biology Laboratory, Heidelberg, Germany.
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M
Hetzer
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Bilbao-Cortés
T C
TC
Walther
O J
OJ
Gruss
I W
IW
Mattaj
D019943:Amino Acid Substitution; D000818:Animals; D018797:Cell Cycle Proteins; D002843:Chromatin / Q000737:chemistry / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D005260:Female; D005455:Fluorescent Antibody Technique; D005456:Fluorescent Dyes; D020662:Guanine Nucleotide Exchange Factors; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism / Q000494:pharmacology; D006153:Guanosine Diphosphate / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism* / Q000494:pharmacology; D006868:Hydrolysis / Q000187:drug effects; D008297:Male; D008561:Membrane Fusion / Q000187:drug effects; D009154:Mutation; D009685:Nuclear Envelope / Q000187:drug effects / Q000378:metabolism*; D009687:Nuclear Proteins; D009865:Oocytes / Q000166:cytology / Q000378:metabolism; D012995:Solubility; D013077:Sperm Head / Q000378:metabolism; D029867:Xenopus Proteins; D014982:Xenopus laevis; D020931:ran GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*
D018797:Cell Cycle Proteins; D002843:Chromatin; D004268:DNA-Binding Proteins; D005456:Fluorescent Dyes; D020662:Guanine Nucleotide Exchange Factors; D009687:Nuclear Proteins; C499304:RCC1 protein, Xenopus; D029867:Xenopus Proteins; D006153:Guanosine Diphosphate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate; D020931:ran GTP-Binding Protein