Pse1p mediates the nuclear import of the iron-responsive transcription factor Aft1p in Saccharomyces cerevisiae.
In Saccharomyces cerevisiae, the iron-responsive transcription factor Aft1p plays a critical role in maintaining iron homeostasis. The activity of Aft1p is induced in response to iron starvation and as a consequence the expression of the iron-regulon is increased. We have shown previously that Aft1p is localized to the cytoplasm under iron-replete conditions but that it is localized to the nucleus under iron-depleted conditions. In this study, we identified the transport receptor that mediates the import of Aft1p into the nucleus, located the nuclear localization signal (NLS) sequences of Aft1p, and examined whether the nuclear import of Aft1p is affected by iron status. In pse1-1 cells, which bear a temperature-sensitive mutation of PSE1, Aft1p was misdirected to the cytoplasm during iron starvation at the restrictive temperature. Aft1p could also directly bind to Pse1p and was dissociated from the complex by Ran-GTP in vitro. These results indicate that Aft1p is imported into the nucleus by Pse1p. Supporting this is that the induction of an Aft1p target gene, FTR1, in response to iron starvation was greatly reduced in pse1-1 cells. Furthermore, we demonstrated that the nuclear localization of a mutant Aft1 protein that contains an NLS derived from SV40 was regulated by iron status regardless of whether Pse1p could interact with Aft1p. This suggests that the interaction between Aft1p and Pse1p is not a critical step that controls the iron-regulated nucleo-cytoplasmic transport of Aft1p.
Ueta
Ryo
R
Department of Applied Molecular Biology, Graduate School of Biostudies, Kyoto University, Sakyo-ku, Kyoto 606-8502, Japan.
Fukunaka
Ayako
A
Yamaguchi-Iwai
Yuko
Y
D021581:Active Transport, Cell Nucleus; D000595:Amino Acid Sequence; D015152:Blotting, Northern; D002467:Cell Nucleus / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D019084:Fluorescent Antibody Technique, Indirect; D049452:Green Fluorescent Proteins; D006160:Guanosine Triphosphate / Q000378:metabolism; D007501:Iron / Q000378:metabolism*; D008164:Luminescent Proteins / Q000378:metabolism; D026901:Membrane Transport Proteins; D008957:Models, Genetic; D008969:Molecular Sequence Data; D019913:Nuclear Localization Signals; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D012313:RNA / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000502:physiology*; D017386:Sequence Homology, Amino Acid; D013696:Temperature; D014157:Transcription Factors / Q000378:metabolism / Q000502:physiology*; D020931:ran GTP-Binding Protein / Q000378:metabolism
C090297:AFT1 protein, S cerevisiae; D008164:Luminescent Proteins; D026901:Membrane Transport Proteins; D019913:Nuclear Localization Signals; C483822:PSE1 protein, S cerevisiae; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D049452:Green Fluorescent Proteins; D012313:RNA; D006160:Guanosine Triphosphate; D007501:Iron; D020931:ran GTP-Binding Protein