Minihelix-containing RNAs mediate exportin-5-dependent nuclear export of the double-stranded RNA-binding protein ILF3.
The karyopherin-related nuclear transport factor exportin-5 preferentially recognizes and transports RNAs containing minihelix motif, a structural cis-acting export element that comprises a double-stranded stem (>14 nucleotides) with a base-paired 5' end and a 3-8-nucleotide protruding 3' end. This structural motif is present in various small cellular and viral polymerase III transcripts such as the adenovirus VA1 RNA (VA1). Here we show that the double-stranded RNA-binding protein, ILF3 (interleukin enhancer binding factor 3) preferentially binds minihelix motif. Gel retardation assays and glutathione S-transferase pull-down experiments revealed that ILF3, exportin-5, RanGTP, and VA1 RNA assembled in a quaternary complex in which the RNA moiety bridges the interaction between ILF3 and exportin-5. Formation of this complex is facilitated by the ability of both exportin-5 and ILF3 to mutually increase their apparent affinity for VA1 RNA. Using microinjection in the nucleus of HeLa cells and transfection experiments, we show here that formation of the cooperative RanGTP-dependent RNA/ILF3/exportin-5 complex promotes the co-transport of VA1 and ILF3 from the nucleus to the cytoplasm. Exportin-5 thus appears as the first example of a nuclear export receptor that mediates RNA export but also promotes transport of proteinaceous cargo through appropriate and specific RNA adaptors.
Institut Jacques Monod, Unité Mixte de Recherche 7592, CNRS, Universités Paris VI et VII, 2 Place Jussieu, Tour 43, Paris 75251 Cedex 05, France.
Carole
C
Gwizdek
Batool
B
Ossareh-Nazari
Amy M
AM
Brownawell
Stefan
S
Evers
Ian G
IG
Macara
Catherine
C
Dargemont
D021581:Active Transport, Cell Nucleus; D000256:Adenoviridae / Q000235:genetics; D020816:Amino Acid Motifs; D000818:Animals; D019921:Biotinylation; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D006224:Cricetinae; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D005982:Glutathione Transferase / Q000378:metabolism; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D028884:Karyopherins / Q000378:metabolism; D050778:NFATC Transcription Factors; D050779:Nuclear Factor 90 Proteins; D009687:Nuclear Proteins; D011233:Precipitin Tests; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012313:RNA / Q000737:chemistry / Q000378:metabolism; D012330:RNA, Double-Stranded; D012367:RNA, Viral / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D014162:Transfection; D020931:ran GTP-Binding Protein / Q000378:metabolism
D004268:DNA-Binding Proteins; D028884:Karyopherins; D050778:NFATC Transcription Factors; D050779:Nuclear Factor 90 Proteins; D009687:Nuclear Proteins; D012330:RNA, Double-Stranded; D012367:RNA, Viral; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; C446511:XPO5 protein, human; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D012313:RNA; D005982:Glutathione Transferase; D020931:ran GTP-Binding Protein