Biogenesis and nuclear export of ribosomal subunits in higher eukaryotes depend on the CRM1 export pathway.
The production of ribosomes constitutes a major biosynthetic task for cells. Eukaryotic small and large ribosomal subunits are assembled in the nucleolus and independently exported to the cytoplasm. Most nuclear export pathways require RanGTP-binding export receptors. We analyzed the role of CRM1, the export receptor for leucine-rich nuclear export signals (NES), in the biogenesis of ribosomal subunits in vertebrate cells. Inhibition of the CRM1 export pathway led to a defect in nuclear export of both 40S and 60S subunits in HeLa cells. Moreover, the export of newly made ribosomal subunits in Xenopus oocytes was efficiently and specifically competed by BSA-NES conjugates. The CRM1 dependence of 60S subunit export suggested a conserved function for NMD3, a factor proposed to be a 60S subunit export adaptor in yeast. Indeed, we observed that nuclear export of human NMD3 (hNMD3) is sensitive to leptomycin B (LMB), which inactivates CRM1. It had, however, not yet been demonstrated that Nmd3 can interact with CRM1. Using purified recombinant proteins we have shown here that hNMD3 binds to CRM1 directly, in a RanGTP-dependent manner, by way of a C-terminal NES sequence. Our results suggest that the functions of CRM1 and NMD3 in ribosomal subunit export are conserved from yeast to higher eukaryotes.
Thomas
Franziska
F
Swiss Federal Institute of Technology (ETH) Zürich, Institute of Biochemistry, HPM F11.1, Switzerland.
Kutay
Ulrike
U
D021581:Active Transport, Cell Nucleus / Q000187:drug effects / Q000502:physiology; D000818:Animals; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D005057:Eukaryotic Cells / Q000378:metabolism* / Q000648:ultrastructure; D019143:Evolution, Molecular; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D028884:Karyopherins / Q000378:metabolism*; D009865:Oocytes; D011485:Protein Binding / Q000502:physiology; D017434:Protein Structure, Tertiary / Q000502:physiology; D021122:Protein Subunits / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins / Q000494:pharmacology; D012270:Ribosomes / Q000187:drug effects / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D014714:Vertebrates / Q000378:metabolism; D014981:Xenopus; D020931:ran GTP-Binding Protein / Q000378:metabolism; D000097606:Exportin 1 Protein
D005231:Fatty Acids, Unsaturated; D028884:Karyopherins; D021122:Protein Subunits; D016601:RNA-Binding Proteins; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D020931:ran GTP-Binding Protein; D000097606:Exportin 1 Protein; C117894:NMD3 protein, S cerevisiae; C038753:leptomycin B