RanGTP-regulated interactions of CRM1 with nucleoporins and a shuttling DEAD-box helicase.
CRM1 is an export receptor mediating rapid nuclear exit of proteins and RNAs to the cytoplasm. CRM1 export cargoes include proteins with a leucine-rich nuclear export signal (NES) that bind directly to CRM1 in a trimeric complex with RanGTP. Using a quantitative CRM1-NES cargo binding assay, significant differences in affinity for CRM1 among natural NESs are demonstrated, suggesting that the steady-state nucleocytoplasmic distribution of shuttling proteins could be determined by the relative strengths of their NESs. We also show that a trimeric CRM1-NES-RanGTP complex is disassembled by RanBP1 in the presence of RanGAP, even though RanBP1 itself contains a leucine-rich NES. Selection of CRM1-binding proteins from Xenopus egg extract leads to the identification of an NES-containing DEAD-box helicase, An3, that continuously shuttles between the nucleus and the cytoplasm. In addition, we identify the Xenopus homologue of the nucleoporin CAN/Nup214 as a RanGTP- and NES cargo-specific binding site for CRM1, suggesting that this nucleoporin plays a role in export complex disassembly and/or CRM1 recycling.
Askjaer
P
P
Department of Gene Expression, Deutsches Krebsforschungszentrum, Heidelberg, Germany.
Bachi
A
A
Wilm
M
M
Bischoff
F R
FR
Weeks
D L
DL
Ogniewski
V
V
Ohno
M
M
Niehrs
C
C
Kjems
J
J
Mattaj
I W
IW
Fornerod
M
M
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001693:Biological Transport, Active; D002352:Carrier Proteins / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism; D017931:DNA Primers / Q000235:genetics; D005260:Female; D019204:GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D028884:Karyopherins; D008954:Models, Biological; D008969:Molecular Sequence Data; D028861:Nuclear Pore Complex Proteins; D009687:Nuclear Proteins / Q000378:metabolism*; D009865:Oocytes / Q000378:metabolism; D011485:Protein Binding; D020365:RNA Helicases / Q000235:genetics / Q000378:metabolism*; D018160:Receptors, Cytoplasmic and Nuclear; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014981:Xenopus; D020931:ran GTP-Binding Protein; D000097606:Exportin 1 Protein
D002352:Carrier Proteins; D017931:DNA Primers; D019204:GTP-Binding Proteins; D028884:Karyopherins; D028861:Nuclear Pore Complex Proteins; D009687:Nuclear Proteins; D020365:RNA Helicases; D018160:Receptors, Cytoplasmic and Nuclear; D011994:Recombinant Proteins; D020931:ran GTP-Binding Protein; D000097606:Exportin 1 Protein; C085914:NUP214 protein, human; C091775:ran-binding protein 1