Characterization of the chromatin acceptor sites for the avian oviduct progesterone receptor using monoclonal antibodies.
Monoclonal antibodies (MAb) against the chromatin acceptor sites for the avian oviduct progesterone receptor were prepared with highly purified hen oviduct acceptor proteins reconstituted to hen DNA. Addition of the MAbs to a cell-free assay blocked progesterone receptor from chick oviduct (PRov) binding to native-like acceptor sites on nucleoacidic protein (NAP) representing a partially deproteinized chromatin, which has been shown to be enriched in these binding sites. However, the antibodies do not block PRov binding to pure DNA, nor do they affect the receptor itself. Estrogen receptor binding to NAP was not inhibited, supporting a receptor specificity of the PRov acceptor sites as reported previously from direct competition studies. These data support earlier studies showing that (1) the reconstituted PRov acceptor sites resemble the native sites, (2) the acceptor sites are receptor specific, and (3) the PRov binding sites of NAP are different from those of pure DNA. While some animal-species specificity in the PRov binding inhibition was observed, no tissue specificity was seen. Direct binding of the antibodies to native acceptor sites was demonstrated in an enzyme-linked immunosorbent assay (ELISA) system. The antibodies showed little recognition of free acceptor protein or DNA alone, indicating specificity for the protein-DNA complex. A partial evolutionary conservation of the nuclear acceptor sites for PRov was shown by the fact that about 50% of the inhibition seen with hen NAP was obtained with NAPs from several other species, and this partial cross-reactivity of the MAbs with the same NAPs from other animal species was also seen in the ELISA.
Goldberger
A
A
Department of Cell Biology, Mayo Clinic, Rochester, Minnesota 55905.
Horton
M
M
Katzmann
J
J
Spelsberg
T C
TC
D000818:Animals; D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex / Q000032:analysis; D002645:Chickens; D002843:Chromatin / Q000378:metabolism*; D004247:DNA / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D007700:Kinetics; D009698:Nucleoproteins / Q000378:metabolism; D010057:Oviducts / Q000378:metabolism*; D011980:Receptors, Progesterone / Q000276:immunology / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D002843:Chromatin; D009698:Nucleoproteins; D011980:Receptors, Progesterone; D004247:DNA