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Vol. 14, Issue 11, 4387-4396, November 2003
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Negatively Regulates Nuclear Membrane Fusion and Nuclear Pore Complex Assembly
* Section of Cell and Developmental Biology, Division of Biological Sciences 0347, University of California, San Diego, La Jolla, California 92093-0347;
Department of Materials and Interfaces, Weizmann Institute of Science, Rehovot 76100, Israel
Submitted May 2, 2003;
Revised July 11, 2003;
Accepted July 14, 2003
Monitoring Editor: Joseph Gall
Assembly of a eukaryotic nucleus involves three distinct events: membrane recruitment, fusion to form a double nuclear membrane, and nuclear pore complex (NPC) assembly. We report that importin 
negatively regulates two of these events, membrane fusion and NPC assembly. When excess importin is added to a full Xenopus nuclear reconstitution reaction, vesicles are recruited to chromatin but their fusion is blocked. The importin down-regulation of membrane fusion is Ran-GTP reversible. Indeed, excess RanGTP (RanQ69L) alone stimulates excessive membrane fusion, leading to intranuclear membrane tubules and cytoplasmic annulate lamellae-like structures. We propose that a precise balance of importin to Ran is required to create a correct double nuclear membrane and simultaneously to repress undesirable fusion events. Interestingly, truncated importin 45–462 allows membrane fusion but produces nuclei lacking any NPCs. This reveals distinct importin -regulation of NPC assembly. Excess full-length importin and 45–462 act similarly when added to prefused nuclear intermediates, i.e., both block NPC assembly. The importin NPC block, which maps downstream of GTP
S and BAPTA-sensitive steps in NPC assembly, is reversible by cytosol. Remarkably, it is not reversible by 25 µM RanGTP, a concentration that easily reverses fusion inhibition. This report, using a full reconstitution system and natural chromatin substrates, significantly expands the repertoire of importin . Its roles now encompass negative regulation of two of the major events of nuclear assembly: membrane fusion and NPC assembly.
Corresponding author. E-mail address: dforbes{at}ucsd.edu. This article has been cited by other articles:
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