Importin β Can Mediate the Nuclear Import of an Arginine-Rich Nuclear Localization Signal in the Absence of Importin α

Abstract

The import of proteins into the nucleus is dependent on cis-acting targeting sequences, nuclear localization signals (NLSs), and members of the nuclear transport receptor (importin-β-like) superfamily. The most extensively characterized import pathway, often termed the classical pathway, is utilized by many basic-type (lysine-rich) NLSs and requires an additional component, importin α, to serve as a bridge between the NLS and the import receptor importin β. More recently, it has become clear that a variety of proteins enter the nucleus via alternative import receptors and that their NLSs bind directly to those receptors. By using the digitonin-permeabilized cell system for protein import in vitro, we have defined the import pathway for the Rex protein of human T-cell leukemia virus type 1. Interestingly, the arginine-rich NLS of Rex uses importin β for import but does so by a mechanism that is importin α independent. Based on the ability of the Rex NLS to inhibit the import of the lysine-rich NLS of T antigen and of both NLSs to be inhibited by the domain of importin α that binds importin β (the IBB domain), we infer that the Rex NLS interacts with importin β directly. In addition, and in keeping with other receptor-mediated nuclear import pathways, Rex import is dependent on the integrity of the Ran GTPase cycle. Based on these results, we suggest that importin β can mediate the nuclear import of arginine-rich NLSs directly, or lysine-rich NLSs through the action of importin α.

ACKNOWLEDGMENTS

We thank Steve Adam, Bryan Cullen, Gideon Dreyfuss, Matt Michael, and Ray Truant for sharing reagents; Paul Eder, Ron Fouchier, Sara Nakielny, and Vicki Pollard for helpful discussions; and Laurie Zimmerman for excellent secretarial support.

This work was supported by the Howard Hughes Medical Institute, a U.S. Public Service grant AI41933 (to M.H.M.) from NIAID, and a Minority Predoctoral Fellowship (5-F31-HG-00065) from NIGMS (to D.P.).