Importin α Nuclear Localization Signal Binding Sites for STAT1, STAT2, and Influenza A Virus Nucleoprotein

Krister Melén,Riku Fagerlund,Jacqueline Franke,Matthias Köhler,Leena Kinnunen,Ilkka Julkunen

doi:10.1074/jbc.m303571200

Abstract

Proteins actively transported into the nucleus via the classical nuclear import pathway contain nuclear localization signals (NLSs), which are recognized by the family of importin alpha molecules. Importin alpha contains 10 armadillo (arm) repeats, of which the N-terminal arm repeats 2-4 have been considered as the "major" NLS binding site. Interferon-activated, dimerized signal transducers and activators of transcription (STAT1 and STAT2) directly bind to importin alpha5 via a dimeric nonclassical NLS. Here we show by site-directed mutagenesis that the very C-terminal arm repeats 8 and 9 of importin alpha5 form a unique binding site for STAT1 homodimers and STAT1-STAT2 heterodimers. Influenza A virus nucleoprotein also contains a nonclassical NLS that is recognized by the C-terminal NLS binding site of importin alpha5, comprising arm repeats 7-9. Binding of influenza A virus nucleoprotein to importin alpha3 also occurs via the C-terminal arm repeats. Simian virus 40 large T antigen instead binds to the major N-terminal arm repeats of importin alpha3, indicating that one importin alpha molecule is able to use either its N- or C-terminal arm repeats for binding various NLS containing proteins.

Highlights

Proteins actively transported into the nucleus via the classical nuclear import pathway contain nuclear localization signals (NLSs), which are recognized by the family of importin ␣ molecules
We show by site-directed mutagenesis that the very C-terminal arm repeats 8 and 9 of importin ␣5 form a unique binding site for STAT1 homodimers and STAT1-STAT2 heterodimers
We show by site-directed mutagenesis that importin ␣5 recognizes the nonclassical STAT1, STAT2, and influenza A virus NP NLSs by its arm repeat domain

Summary

Introduction

Proteins actively transported into the nucleus via the classical nuclear import pathway contain nuclear localization signals (NLSs), which are recognized by the family of importin ␣ molecules. Importin ␣ contains 10 armadillo (arm) repeats, of which the N-terminal arm repeats 2– 4 have been considered as the “major” NLS binding site. Influenza A virus nucleoprotein contains a nonclassical NLS that is recognized by the C-terminal NLS binding site of importin ␣5, comprising arm repeats 7–9. Simian virus 40 large T antigen instead binds to the major N-terminal arm repeats of importin ␣3, indicating that one importin ␣ molecule is able to use either its N- or C-terminal arm repeats for binding various NLS containing proteins. Proteins transported into the nucleus contain nuclear location signals (NLSs) that are recognized by importin ␣/importin ␤ heterodimers. A typical bipartite NLS contains two clusters of basic amino acids separated by a linker of 10 –11 amino acids [21]

Methods

Results

Conclusion