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© The Rockefeller University Press, 0021-9525/1999//237 $5.00
The Journal of Cell Biology, Volume 145, Number 2, , 1999 237-254

Regular Articles

RAE1 Is a Shuttling mRNA Export Factor That Binds to a GLEBS-like NUP98 Motif at the Nuclear Pore Complex through Multiple Domains



Colin E.J. Pritchard*, Maarten Fornerod{ddagger}, Lawryn H. Kasper*, and Jan M.A. van Deursen*,§

* Department of Genetics, St. Jude Children's Research Hospital, Memphis, Tennessee 38105; {ddagger} The European Molecular Biology Laboratory, Heidelberg, Germany D69117; and § Department of Pediatrics & Adolescent Medicine, Mayo Clinic, Rochester, Minnesota 55905

Gle2p is implicated in nuclear export of poly(A)+ RNA and nuclear pore complex (NPC) structure and distribution in Saccharomyces cerevisiae. Gle2p is anchored at the nuclear envelope (NE) via a short Gle2p-binding motif within Nup116p called GLEBS. The molecular mechanism by which Gle2p and the Gle2p–Nup116p interaction function in mRNA export is unknown. Here we show that RAE1, the mammalian homologue of Gle2p, binds to a GLEBS-like NUP98 motif at the NPC through multiple domains that include WD-repeats and a COOH-terminal non–WD-repeat extension. This interaction is direct, as evidenced by in vitro binding studies and chemical cross-linking. Microinjection experiments performed in Xenopus laevis oocytes demonstrate that RAE1 shuttles between the nucleus and the cytoplasm and is exported from the nucleus in a temperature-dependent and RanGTP-independent manner. Docking of RAE1 to the NE is highly dependent on new mRNA synthesis. Overexpression of the GLEBS-like motif also inhibits NE binding of RAE1 and induces nuclear accumulation of poly(A)+ RNA. Both effects are abrogated either by the introduction of point mutations in the GLEBS-like motif or by overexpression of RAE1, indicating a direct role for RAE1 and the NUP98–RAE1 interaction in mRNA export. Together, our data suggest that RAE1 is a shuttling transport factor that directly contributes to nuclear export of mRNAs through its ability to anchor to a specific NUP98 motif at the NPC.

Key Words: nuclear pore complex • mRNA export • RNA polymerase II • NUP98 • RAE1

Abbreviations used in this paper: AMD, actinomycin D; DSS, disuccinimidyl suberate; GST, glutathione-S-transferase; HA, hemagglutinin; NE, nuclear envelope; NES, nuclear export signal; NLS, nuclear localization signal; NPC, nuclear pore complex.

We are very grateful to Iain Mattaj for his constructive criticisms and generous support throughout this study. We thank Haruhiko Siomi, Susanne Bailer, Richard Bram, Mike Matunis, Eva Neer, and Xiaosheng Wu for helpful discussions, and Paul Brindle and Mutsuhito Ohno for critical reading of the manuscript. Scott Kuersten, Brian Burke, Gerard Grosveld, Hermann Bujard, Derrick Person, Arthur Nienhuis, and Albert Reynolds kindly provided materials. This work would not have been possible without the confocal microscope of Jim Ihle.


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