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The Nuclear Pore Complex: Assembly Required

Objective

The nuclear pore complex (NPC) mediates all traffic between the nucleus and cytoplasm and serves as a central node of cellular communication. Despite the recent identification of the full complement of nuclear pore proteins, much of the molecular details of NPC structure and its mechanism of action, remain unknown. Moreover, the precise molecular pathway for NPC biogenesis is largely undefined. Here, I present a comprehensive approach for dissecting the mechanism of NPC assembly, using in vitro nuclear reconstitution in Xenopus egg extracts. My working hypothesis is that NPC assembly begins with the formation of fused double nuclear membranes. In this model, an initial fusion event between the inner and outer nuclear membranes triggers the assembly process, and soluble nucleoporins are then added in a sequential manner, to form the complete structure.

A prediction of this model is that pore-membrane protein(s) should play a vital role at the early stages of assembly. To test this prediction we will use morphological analysis and immunolocalization of the pore-membrane proteins by transmission electron microscopy. Aborted assembly intermediates will be isolated and used in combination with fractionated cytosol to identify subsequent steps in the process. Newly identified assembly intermediates will be characterize by fluorescence and electron microscopy. The overall goal is to reach a detailed step-by-step model of NPC assembly. This will promote our understanding of the biogenesis and function of one of the largest supramolecular assemblies in eukaryotic cells, and the gateway to the genome.

Call for proposal

FP6-2004-MOBILITY-12
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Funding Scheme

IRG - Marie Curie actions-International re-integration grants

Coordinator

TECHNION-ISRAEL INSTITUTE OF TECHNOLOGY
Address
Technion City
Haifa
Israel