Mechanisms of progesterone receptor export from nuclei: role of nuclear localization signal, nuclear export signal, and ran guanosine triphosphate.
Steroid hormone receptors are, in most cases, mainly nuclear proteins that undergo a continuous nucleocytoplasmic shuttling. The mechanism of the nuclear export of these proteins remains largely unknown. To approach this problem experimentally in vivo, we have prepared cell lines permanently coexpressing the wild-type nuclear progesterone receptor (PR) and a cytoplasmic receptor mutant deleted of its nuclear localization signal (NLS) [(deltaNLS)PR]. Each receptor species was deleted from the epitope recognized by a specific monoclonal antibody, thus allowing separated observation of the two receptor forms in the same cells. Administration of hormone provoked formation of heterodimers during nucleocytoplasmic shuttling and import of (deltaNLS)PR into the nucleus. Washing out of the hormone allowed us to follow the export of (deltaNLS)PR into the cytoplasm. Microinjection of BSA coupled to a NLS inhibited the export of (deltaNLS)PR. On the contrary, microinjection of BSA coupled to a nuclear export signal (NES) was without effect. Moreover, leptomycin B, which inhibits NES-mediated export, was also without effect. tsBN2 cells contain a thermosensitive RCC1 protein (Ran GTP exchange protein). At the nonpermissive temperature, the nuclear export of (deltaNLS)PR could be observed, whereas the export of NES-BSA was suppressed. Microinjection of GTPgammaS confirmed that the export of (deltaNLS)PR was not dependent on GTP hydrolysis. These experiments show that the nuclear export of PR is not NES mediated but probably involves the NLS. It does not involve Ran GTP, and it is not dependent on the hydrolysis of GTP. The nucleocytoplasmic shuttling of steroid hormone receptors thus appears to utilize mechanisms different from those previously described for some viral, regulatory, and heterogeneous ribonuclear proteins.
Tyagi
R K
RK
Hormones et Reproduction, INSERM U135, Faculté de Médecine Paris-Sud, Le Kremlin-Bicêtre, France.
Amazit
L
L
Lescop
P
P
Milgrom
E
E
Guiochon-Mantel
A
A
D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D019556:COS Cells; D018797:Cell Cycle Proteins; D002467:Cell Nucleus / Q000378:metabolism*; D002478:Cells, Cultured; D006224:Cricetinae; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D020662:Guanine Nucleotide Exchange Factors; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D006160:Guanosine Triphosphate / Q000502:physiology; D006801:Humans; D007668:Kidney; D007739:L Cells; D008647:Mesocricetus; D051379:Mice; D008845:Microinjections; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism*; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000378:metabolism; D011374:Progesterone / Q000494:pharmacology; D021382:Protein Sorting Signals / Q000378:metabolism*; D011980:Receptors, Progesterone / Q000378:metabolism*; D017384:Sequence Deletion; D012710:Serum Albumin, Bovine / Q000378:metabolism; D020931:ran GTP-Binding Protein
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D005231:Fatty Acids, Unsaturated; D020662:Guanine Nucleotide Exchange Factors; D009687:Nuclear Proteins; D010446:Peptide Fragments; D021382:Protein Sorting Signals; C062521:RCC1 protein, human; C499302:Rcc1 protein, mouse; D011980:Receptors, Progesterone; D012710:Serum Albumin, Bovine; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D011374:Progesterone; D006160:Guanosine Triphosphate; D020931:ran GTP-Binding Protein; C038753:leptomycin B