Nucleocytoplasmic shuttling of JAZ, a new cargo protein for exportin-5.
Exportin-5 is a nuclear export receptor for certain classes of double-stranded RNA (dsRNA), including pre-micro-RNAs, viral hairpin RNAs, and some tRNAs. It can also export the RNA binding proteins ILF3 and elongation factor EF1A. However, the rules that determine which RNA binding proteins are exportin-5 cargoes remain unclear. JAZ possesses an unusual dsRNA binding domain consisting of multiple C2H2 zinc fingers. We found that JAZ binds to exportin-5 in a Ran-GTP- and dsRNA-dependent manner. Exportin-5 stimulates JAZ shuttling, and gene silencing of exportin-5 reduces shuttling. Recombinant exportin-5 also stimulates nuclear export of JAZ in permeabilized cells. JAZ also binds to ILF3, and surprisingly, this interaction is RNA independent, even though it requires the dsRNA binding domains of ILF3. Exportin-5, JAZ, and ILF3 can form a heteromeric complex with Ran-GTP and dsRNA, and JAZ increases ILF3 binding to exportin-5. JAZ does not contain a classical nuclear localization signal, and in digitonin-permeabilized cells, nuclear accumulation of JAZ does not require energy or cytosol. Nonetheless, low temperatures prevent JAZ import, suggesting that nuclear entry does not occur via simple diffusion. Together, these data suggest that JAZ is exported by exportin-5 but translocates back into nuclei by a facilitated diffusion mechanism.
Chen
Ting
T
Center for Cell Signaling, Department of Microbiology, Health Sciences Center, University of Virginia School of Medicine, Charlottesville, VA 22908-0577, USA.
Brownawell
Amy M
AM
Macara
Ian G
IG
D021581:Active Transport, Cell Nucleus; D000818:Animals; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D003001:Cloning, Molecular; D006224:Cricetinae; D003600:Cytosol / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D004058:Diffusion; D004305:Dose-Response Relationship, Drug; D005982:Glutathione Transferase / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D028884:Karyopherins / Q000737:chemistry* / Q000378:metabolism; D050778:NFATC Transcription Factors; D050779:Nuclear Factor 90 Proteins; D009687:Nuclear Proteins; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D012313:RNA / Q000378:metabolism; D012330:RNA, Double-Stranded / Q000737:chemistry; D012333:RNA, Messenger / Q000378:metabolism; D012343:RNA, Transfer / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry; D013696:Temperature; D013997:Time Factors; D014157:Transcription Factors / Q000737:chemistry; D014162:Transfection; D020798:Two-Hybrid System Techniques; D016335:Zinc Fingers; D020931:ran GTP-Binding Protein / Q000737:chemistry* / Q000378:metabolism
D004268:DNA-Binding Proteins; D028884:Karyopherins; D050778:NFATC Transcription Factors; D050779:Nuclear Factor 90 Proteins; D009687:Nuclear Proteins; D012330:RNA, Double-Stranded; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; D014157:Transcription Factors; C446511:XPO5 protein, human; C121935:ZNF346 protein, human; D012313:RNA; D012343:RNA, Transfer; D005982:Glutathione Transferase; D020931:ran GTP-Binding Protein