Recognition of engineered tRNAs with an extended 3' end by Exportin-t (Xpo-t) and transport of tRNA-attached ribozymes to the cytoplasm in somatic cells.
Our recent analysis indicates that the cytoplasmic localization of tRNA-attached ribozymes (tRNA-Rz) is critical for its high-level intracellular activity, suggesting that mature mRNAs in the cytoplasm are more accessible to ribozymes than pre-mRNAs in the nucleus (Kato et al. J. Biol. Chem. 2001, 276, 15378-15385; Kuwabara et al. Nucleic Acids Res. 2001, 29, 2780-2788). Although studies in Xenopus oocytes led to the proposal that only correctly processed mature tRNAs are exported from nuclei in a RanGTP-dependent manner (Lund and Dahlberg Science 1998, 282, 2082-2085), our tRNA-Rz with an extended 3' end can also be exported to the cytoplasm in somatic cells. Xpo-t/RanGTP bound to tRNA-attached ribozymes in vitro and in somatic cells, with recognition basically resembling the recognition of mature tRNAs. In contrast, no binding to tRNA-attached ribozymes occurred in Xenopus oocytes. The injection of a nuclear extract of Xenopus oocytes together with tRNA-attached ribozymes inhibited the export of tRNA-attached ribozymes but not mature tRNAs in somatic cells, suggesting the existence of an inhibitor(s) of the Xpo-t-dependent export pathway. Moreover, the inhibitor(s) appears responsible for a proofreading mechanism that operates in oocytes.
Kuwabara
T
T
Department of Chemistry and Biotechnology, Graduate School of Engineering, The University of Tokyo, Hongo, Tokyo 113-8656, Japan.
Warashina
M
M
Sano
M
M
Tang
H
H
Wong-Staal
F
F
Munekata
E
E
Taira
K
K
D021581:Active Transport, Cell Nucleus; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D001693:Biological Transport, Active; D002352:Carrier Proteins / Q000378:metabolism* / Q000502:physiology; D002467:Cell Nucleus / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism; D005818:Genetic Engineering / Q000379:methods; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D029543:Nucleocytoplasmic Transport Proteins; D009865:Oocytes; D026721:RNA 3' End Processing / Q000235:genetics; D016337:RNA, Catalytic / Q000378:metabolism*; D012343:RNA, Transfer / Q000235:genetics* / Q000378:metabolism*; D014981:Xenopus
D002352:Carrier Proteins; D029543:Nucleocytoplasmic Transport Proteins; D016337:RNA, Catalytic; C503618:XPOT protein, human; D012343:RNA, Transfer