Nuclear progesterone receptor is mainly heat shock protein 90-free in vivo.
Heat shock protein 90 (hsp90) is associated with many steroid receptors in tissue homogenates. It is widely accepted that hsp90 regulates the binding of the receptor to the corresponding gene regulatory element. However there is no unequivocal evidence that steroid receptor-hsp90 complexes are present in the intact cells. We demonstrate here the absence of progesterone receptor (PR)-hsp90 complexes in intact target cell nuclei, using immunohistochemical and biochemical methods to determine the location and composition of the nonliganded (aporeceptor) and liganded (holoreceptor) PR complexes. In the chicken oviduct cells, both apo- and holoreceptors were nuclear, while hsp90 was exclusively cytoplasmic. When expressed transiently in HeLa cells, hsp90 was detected in the cytoplasm and PR was detected in the nucleus. Their location or staining intensity was not affected when they were coexpressed in the same cells. To confirm that the sensitivity of the immunohistochemical detection of hsp90 and PR did not differ significantly, a chimeric hsp90-PR was transiently expressed in HeLa cells. Both hsp90 and PR antigens of the chimera were detected in nuclei with the same intensity. In homogenates of the same tissue samples that were used for immunohistochemistry, the PR was complexed with hsp90. Hsp90-PR complexes were formed in vitro when immature bursa of Fabricius, known to contain high levels of hsp90, was homogenized in the presence of hsp90-free aporeceptor, while holoreceptor did not associate with hsp90. Our data show that nuclear PR is not complexed with hsp90 in vivo and suggest that the 8S-PR may be an in vitro artifact generated during tissue processing.
Department of Biomedical Sciences, University of Tempere, Finland.
P
P
Tuohimaa
A
A
Pekki
M
M
Bläuer
T
T
Joensuu
P
P
Vilja
T
T
Ylikomi
D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism*; D002645:Chickens; D003001:Cloning, Molecular; D003600:Cytosol / Q000378:metabolism; D004247:DNA; D004958:Estradiol / Q000494:pharmacology*; D005260:Female; D006367:HeLa Cells; D006360:Heat-Shock Proteins / Q000032:analysis*; D006801:Humans; D015151:Immunoblotting; D007150:Immunohistochemistry; D010057:Oviducts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D011374:Progesterone / Q000378:metabolism*; D011980:Receptors, Progesterone / Q000032:analysis / Q000096:biosynthesis / Q000378:metabolism*; D014162:Transfection
D006360:Heat-Shock Proteins; D011980:Receptors, Progesterone; D011374:Progesterone; D004958:Estradiol; D004247:DNA