Difference in nucleocytoplasmic shuttling sequences of rat and human constitutive active/androstane receptor.
Fluorescence recovery after photobleaching (FRAP) in spontaneous multinuclear cells shows that both rat and human constitutive active/androstane receptors (CARs) are shuttling proteins with both nuclear localization signals (NLSs) and nuclear export signals (NESs). We previously identified two NLSs in rat CAR: NLS1 in the hinge region (residues 100-108) and NLS2 in the ligand-binding domain (residues 111-320). In the present study, we compared the intracellular localization signals between rat and human CARs. There was a marked difference in their intracellular localization in COS-7 cells because, unlike rat CAR, human CAR does not contain NLS1 due to an amino acid change at position 106. A CRM1-dependent leucine-rich NES, which is sensitive to an inhibitory effect of leptomycin B, was found in the cytoplasmic retention region previously identified within the ligand-binding domain of rat CAR (residues 220-258). We found that human CAR instead has a NES in the ligand-binding domain between residues 170 and 220. Also, we detected CRM1-independent C-terminal NESs between residues 317-358 of rat and human CARs. Removal of NLS1 by N-terminal truncation and mutation of xenochemical response signal caused rat CAR to localize in the cytoplasm of COS-7 cells, which we suspect is due to the masking of NLS2.
Kanno
Yuichiro
Y
Faculty of Pharmaceutical Sciences, Toho University, 2-2-1 Miyama, Funabashi, Chiba 274-8510, Japan.
Suzuki
Motoyoshi
M
Miyazaki
Yukari
Y
Matsuzaki
Midori
M
Nakahama
Takayuki
T
Kurose
Kouichi
K
Sawada
Jun-ichi
J
Inouye
Yoshio
Y
D021581:Active Transport, Cell Nucleus / Q000187:drug effects / Q000235:genetics; D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D019556:COS Cells; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism*; D002522:Chlorocebus aethiops; D003593:Cytoplasm / Q000235:genetics / Q000378:metabolism*; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D006801:Humans; D019913:Nuclear Localization Signals / Q000235:genetics / Q000378:metabolism*; D038761:Photobleaching; D017434:Protein Structure, Tertiary / Q000235:genetics; D051381:Rats; D011944:Receptors, Androgen / Q000235:genetics / Q000378:metabolism*; D013045:Species Specificity; D028901:alpha Karyopherins / Q000235:genetics / Q000378:metabolism
D000903:Antibiotics, Antineoplastic; D005231:Fatty Acids, Unsaturated; D019913:Nuclear Localization Signals; D011944:Receptors, Androgen; D028901:alpha Karyopherins; C038753:leptomycin B