Nuclear export of proteins in plants: AtXPO1 is the export receptor for leucine-rich nuclear export signals in Arabidopsis thaliana.
Transport across the nuclear envelope is mediated by transport receptors from the Importin beta family. We identified Exportin 1 from Arabidopsis (AtXPO1/AtCRM1) as the nuclear export receptor for proteins carrying leucine-rich nuclear export signals (NESs). AtXPO1 shares 42-50% identity with its functional homologues from humans and yeasts. We functionally characterised AtXPO1 by its interaction with NESs of animal and plant proteins, which is inhibited by the cytotoxin leptomycin B (LMB), and also by its interaction with the small GTPase Ran1 in the yeast two-hybrid system. Furthermore, we demonstrated the existence of a nuclear export pathway for proteins in plants. For the characterisation of nuclear export activities, we established an in vivo assay based on the localisation equilibrium of a GFP reporter protein fused to both a nuclear localisation signal (NLS) and an NES motif. Using this in vivo assay we demonstrated that the NES of the heterologous protein Rev is also functional in plants and that its export is inhibited by LMB. In addition, we identified a leucine-rich NES in the Arabidopsis protein AtRanBP1a. The NES, which is located at the carboxy terminus of the protein, is disrupted by mutating three long chain hydrophobic amino acid residues to alanine (L176A, L179A, V181A). In BY-2 protoplasts the NES of AtRanBP1a is functionally indistinguishable from the Rev NES. Our results demonstrate that the machinery for the nuclear export of proteins is functionally conserved in plants.
Haasen
D
D
University of Freiburg, Institute for Biology II, Cell Biology, Schänzlestr. 1, D-79104 Freiburg, Germany.
Köhler
C
C
Neuhaus
G
G
Merkle
T
T
D000595:Amino Acid Sequence; D000818:Animals; D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism*; D001693:Biological Transport, Active; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics / Q000302:isolation & purification; D018744:DNA, Plant / Q000235:genetics / Q000302:isolation & purification; D015870:Gene Expression; D017343:Genes, Plant; D006801:Humans; D028884:Karyopherins; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D030821:Plants, Genetically Modified; D010947:Plants, Toxic; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D018749:RNA, Plant / Q000235:genetics / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014018:Tissue Distribution; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism; D020798:Two-Hybrid System Techniques; D000097606:Exportin 1 Protein
D002352:Carrier Proteins; D017931:DNA Primers; D018076:DNA, Complementary; D018744:DNA, Plant; D028884:Karyopherins; D009687:Nuclear Proteins; D010940:Plant Proteins; D012333:RNA, Messenger; D018749:RNA, Plant; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; D000097606:Exportin 1 Protein