The monopartite nuclear localization signal of autoimmune regulator mediates its nuclear import and interaction with multiple importin alpha molecules.
Autoimmune regulator (AIRE) is a transcriptional regulator involved in establishing immunological self-tolerance. Mutations in the AIRE gene lead to the development of the autosomal, recessively inherited, organ-specific autoimmune disease, autoimmune polyendocrinopathy candidiasis ectodermal dystrophy (APECED). The AIRE protein is mainly localized in the cell nucleus where it is associated with nuclear bodies. The N-terminal part of AIRE has been previously shown to mediate nuclear localization of the protein. However, the functional nuclear localization signal (NLS) and nuclear import mechanisms of AIRE have not been identified. We show that, although the amino-acid sequence of AIRE contains a potential bipartite NLS consisting of amino acids 110-114 and 131-133, only the latter part constitutes a functional NLS. Furthermore, we show by in vitro binding assays that AIRE interacts with multiple members of the nuclear transport receptor importin alpha family, mainly alpha1, alpha3, and alpha5, and that these interactions depend on the intactness of the Arg-Lys-rich NLS of AIRE. In addition, we found that AIRE binds to the 'minor' NLS-binding site of importin alpha3 and alpha5 proteins consisting of the C-terminal armadillo repeats 7-9. Our findings strongly suggest that the nuclear import of AIRE is mediated by the classical importin alpha/beta pathway through binding to several importin alpha family members.
Ilmarinen
Tanja
T
Department of Molecular Medicine, National Public Health Institute, Helsinki, Finland. tanja.ilmarinen@ktl.fi
Melén
Krister
K
Kangas
Hannele
H
Julkunen
Ilkka
I
Ulmanen
Ismo
I
Eskelin
Petra
P
D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002467:Cell Nucleus / Q000378:metabolism*; D002522:Chlorocebus aethiops; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019913:Nuclear Localization Signals; D010957:Plasmids; D011485:Protein Binding; D021381:Protein Transport; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism*; D020798:Two-Hybrid System Techniques; D028901:alpha Karyopherins / Q000378:metabolism*; D000096927:AIRE Protein
D019913:Nuclear Localization Signals; D014157:Transcription Factors; D028901:alpha Karyopherins; D000096927:AIRE Protein