1. Nxt1 Is Necessary for the Terminal Step of Crm1-Mediated Nuclear Export
- Author
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Lyne Lévesque, James M. Holaska, Carol Gwizdek, Ben E. Black, Catherine Dargemont, Bryce M. Paschal, Batool Ossareh-Nazari, University of Virginia [Charlottesville], and IJM (UMR_7592) - Institut Jacques Monod
- Subjects
green fluorescent protein ,Nucleocytoplasmic Transport Proteins ,Time Factors ,[SDV]Life Sciences [q-bio] ,NXT1 ,STV ,Receptors, Cytoplasmic and Nuclear ,Crm1 ,0302 clinical medicine ,RNA, Transfer ,glutathione-S -transferase ,Genes, Reporter ,RNA, Small Nuclear ,glucocorticoid receptor ,Nuclear protein ,Nuclear pore ,leptomycin B ,0303 health sciences ,PKI ,protein kinase inhibitor ,nuclear export signal ,nuclear envelope ,Cell biology ,GR ,medicine.anatomical_structure ,Gene Products, rev ,Original Article ,NPC ,NE ,Recombinant Fusion Proteins ,Active Transport, Cell Nucleus ,NLS ,nuclear transport ,Biology ,Karyopherins ,GFP ,Cell Line ,03 medical and health sciences ,Rev response element ,nuclear pore complex ,constitutive transport element ,medicine ,streptavidin ,Animals ,RNA, Messenger ,GST ,Nuclear export signal ,LMB ,030304 developmental biology ,Cell Nucleus ,Nucleoplasm ,Cell Biology ,nuclear localization signal ,CTE ,Cell nucleus ,Ran ,Cytoplasm ,Mutagenesis ,NES ,RRE ,Carrier Proteins ,030217 neurology & neurosurgery - Abstract
Soluble factors are required to mediate nuclear export of protein and RNA through the nuclear pore complex (NPC). These soluble factors include receptors that bind directly to the transport substrate and regulators that determine the assembly state of receptor–substrate complexes. We recently reported the identification of NXT1, an NTF2-related export factor that stimulates nuclear protein export in permeabilized cells and undergoes nucleocytoplasmic shuttling in vivo (Black, B.E., L. Lévesque, J.M. Holaska, T.C. Wood, and B.M. Paschal. 1999. Mol. Cell. Biol. 19:8616–8624). Here, we describe the molecular characterization of NXT1 in the context of the Crm1-dependent export pathway. We find that NXT1 binds directly to Crm1, and that the interaction is sensitive to the presence of Ran-GTP. Moreover, mutations in NXT1 that reduce binding to Crm1 inhibit the activity of NXT1 in nuclear export assays. We show that recombinant Crm1 and Ran are sufficient to reconstitute nuclear translocation of a Rev reporter protein from the nucleolus to an antibody accessible site on the cytoplasmic side of the NPC. Further progress on the export pathway, including the terminal step of Crm1 and Rev reporter protein release, requires NXT1. We propose that NXT1 engages with the export complex in the nucleoplasm, and that it facilitates delivery of the export complex to a site on the cytoplasmic side of NPC where the receptor and substrate are released into the cytoplasm.
- Published
- 2001