Novel monoclonal antibodies against human uterine progesterone receptor. Mapping of receptor immunogenic domains.
Progesterone receptor was purified in a single step from human uteri using immunoaffinity chromatography with monoclonal antibodies raised against the rabbit receptor. A total of 39 monoclonal antibodies were prepared against the human receptor and characterized. Immunoblot experiments using crude uterine cytosol or purified receptor showed that the antibodies belonged to three groups: they recognized either a single receptor species (apparent molecular mass 110 kDa), two species (110 and 79 kDa) or three species (110, 79 and 65 kDa). The species specificity of the antibodies was very variable; some recognized only the human receptor, others interacted with several mammalian receptors (rabbit, guinea pig and rat), while a single one also cross-reacted with the chick receptor. The epitopes recognized by 15 of the antibodies showing the strongest affinity for the human receptor were mapped using a method recently described [Lorenzo, Jolivet & Milgrom (1988) Eur. J. Biochem. 176, 53-60] which involves immunoprecipitation of C-terminally truncated proteins obtained by transcription and translation of cloned cDNA in vitro. The antibodies recognized five different regions of the receptor, all localized on the N-terminal half of the protein. None of the antibodies interacted with an epitope present in the DNA-binding or steroid-binding regions of the receptor. Comparison of the pattern of receptor species recognized by the antibodies and the localization of their epitopes showed that the 79 and 65 kDa receptor species were derived from the 110 kDa form by deletion of its N-terminal part. The N-terminus of the 79 kDa species was found to lie between amino acids 121 and 208, and that of the 65 kDa species between amino acids 208 and 296.
Vu Hai
M T
MT
Unité 135 INSERM, Hormones et Reproduction, Faculté de Médecine Paris-Sud, Le Kremlin-Bicêtre, France.
Jolivet
A
A
Ravet
V
V
Lorenzo
F
F
Perrot-Applanat
M
M
Citerne
M
M
Milgrom
E
E
D000911:Antibodies, Monoclonal / Q000276:immunology*; D015153:Blotting, Western; D002846:Chromatography, Affinity; D000939:Epitopes / Q000032:analysis; D005260:Female; D006801:Humans; D008970:Molecular Weight; D011980:Receptors, Progesterone / Q000276:immunology* / Q000302:isolation & purification; D014599:Uterus
D000911:Antibodies, Monoclonal; D000939:Epitopes; D011980:Receptors, Progesterone