© The Rockefeller University Press,
0021-9525/1998//309 $5.00
The Journal of Cell Biology, Volume 143, Number 2,
, 1998 309-318
Determination of the Functional Domain Organization of the Importin
Nuclear Import Factor
Andrea Herold*,
Ray Truant*,
,
Heather Wiegand
, and
Bryan R. Cullen*,
* Department of Genetics and
Howard Hughes Medical Institute, Duke University Medical Center, Durham, North Carolina 27710
Although importin
(Imp
) has been shown to act as the receptor for basic nuclear localization signals (NLSs) and to mediate their recruitment to the importin β nuclear import factor, little is known about the functional domains present in Imp
, with the exception that importin β binding is known to map close to the Imp
NH2 terminus. Here, we demonstrate that sequences essential for binding to the CAS nuclear export factor are located near the Imp
COOH terminus and include a critical acidic motif. Although point mutations introduced into this acidic motif inactivated both CAS binding and Imp
nuclear export, a putative leucine-rich nuclear export signal proved to be neither necessary nor sufficient for Imp
nuclear export. Analysis of sequences within Imp
that bind to the SV-40 T antigen NLS or to the similar LEF-1 NLS revealed that both NLSs interact with a subset of the eight degenerate armadillo (Arm) repeats that form the central part of Imp
. However, these two NLS-binding sites showed only minimal overlap, thus suggesting that the degeneracy of the Arm repeat region of Imp
may serve to facilitate binding to similar but nonidentical basic NLSs. Importantly, the SV-40 T NLS proved able to specifically inhibit the interaction of Imp
with CAS in vitro, thus explaining why the SV-40 T NLS is unable to also function as a nuclear export signal.
Key Words: importin
nuclear export nuclear import nuclear localization signal
Abbreviations used in this paper: aa, amino acid(s); Arm, armadillo motif; β-gal, β-galactosidase; CMV, cytomegalovirus; GST, glutathione-S-transferase; IBB, importin β binding domain; Imp
, importin
; Imp β, importin β; NES, nuclear export signal; NLS, nuclear localization signal; T NLS, SV-40 T antigen NLS.
This research was funded by the Howard Hughes Medical Institute. A. Herold was supported by funds from the German Academic Exchange Organization (DAAD).
Address all correspondence to B.R. Cullen, Department of Genetics, Room 426 CARL Building, Research Drive, Box 3025, Duke University Medical Center, Durham, NC 27710. Tel.: (919) 684-3369. Fax: (919) 681-8979. E-mail: culle002{at}mc.duke.edu

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