Immunological identification of a 50 kDa Mr FK506-binding immunophilin as a component of the non-DNA binding, hsp90 and hsp70 containing, heterooligomeric form of the chick oviduct progesterone receptor.
p59/HBI, a Heat shock protein (hsp90)-Binding-Immunophilin of approximately 59 kDa, was originally detected in the heterooligomeric, non-DNA binding form of numerous mammalian steroid hormone receptors. P59/HBI belongs to the FKBP family since it binds the immunosuppressants FK506 and Rapamycin. Using immobilized FK506, we purified [3H]Org 2058-progesterone receptor (PR)-hsp90-complexes from chick oviduct cytosol in a species migrating at 9S in density gradient. This led to suppose that a protein present in these complexes is a FK506 binding protein. Following incubation of this FK506-affinity purified 9S-PR with BF4, a specific anti-chick hsp90 monoclonal antibody, a shift of the [3H]Org 2058-PR complexes from 9S to 11S has been observed, indicating the presence of hsp90, hsp70 also is included in the 9S-PR complexes as demonstrated by Western blotting and density gradient experiments. Two lines of evidence support the notion that an immunophilin of 50 kDa also associates to heterooligomeric 9S-PR complexes. Firstly, a shift at 11S of the FK506 eluted 9S-PR occurs in sucrose gradient after incubation with 419, a new polyclonal antibody raised against a peptide corresponding to the Hinge I region [Callebaut I., Renoir J. M., Lebeau M. C., Massol N., Burny A., Baulieu E. E., Mornon J. P. (1992) Proc. Natl. Acad. Sci. USA 89, 6270-6274] of the rabbit p59/HBI (amino acids Phe135 to Gly149). Secondly, in Western blotting experiments, both 419 and another anti-FKBP antibody, raised against purified Streptomyces Crysomallus FKBP12 [Pahl A. and Keller U. (1992) J. Bacteriol., 3, 325-333], allow detection of the same 50 kDa protein in the affinity purified PR.(ABSTRACT TRUNCATED AT 250 WORDS)
Renoir
J M
JM
INSERM U33, Lab. Hormones, Hôpital de Bicêtre, France.
Pahl
A
A
Keller
U
U
Baulieu
E E
EE
D000818:Animals; D015153:Blotting, Western; D002352:Carrier Proteins / Q000302:isolation & purification* / Q000378:metabolism*; D002645:Chickens; D004268:DNA-Binding Proteins / Q000737:chemistry*; D005260:Female; D006360:Heat-Shock Proteins / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism*; D007166:Immunosuppressive Agents / Q000378:metabolism; D010057:Oviducts / Q000737:chemistry; D011090:Polyenes / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D011980:Receptors, Progesterone / Q000737:chemistry* / Q000378:metabolism; D020123:Sirolimus; D016559:Tacrolimus / Q000378:metabolism*; D022021:Tacrolimus Binding Proteins
D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D006360:Heat-Shock Proteins; D007166:Immunosuppressive Agents; D011090:Polyenes; D011980:Receptors, Progesterone; D022021:Tacrolimus Binding Proteins; D020123:Sirolimus; D016559:Tacrolimus