Characterization of progesterone-binding moieties in the little skate Raja erinacea.
In this study we report evidence of a [3H]progesterone-binding moiety in the liver and oviduct of the little skate Raja erinacea. It is characterized by high affinity, low capacity and DNA-cellulose-binding activity. Furthermore Western blot analysis revealed that monoclonal antibodies against the chicken progesterone receptor (PR) subunits A and B cross-reacted with a 110-kDa band in the liver and a 80-kDa band in the oviduct. When analyzed by DEAE-Sepharose ion-exchange column chromatography, [3H]progesterone-binding molecules resolved into two peaks, one nonadherent and one adherent to the column. The liver adherent peak eluted in a linear gradient at a NaCl concentration of about 0.07 M and resolved on Western blot as a single band of a 110 kDa. The oviduct adherent peak eluted at about 0.14 M NaCl and resolved on Western blot as a single band of 80 kDa. Competition studies showed that the progesterone-binding moiety in the cytosol was specific for progesterone. On the contrary, the nuclear component is not specific for progesterone; it also binds testosterone and estradiol 17 beta in the oviduct, and progesterone, testosterone, dihydrotestosterone, estradiol 17 beta, mibolerone, and R5020 in the liver. The [3H]progesterone-binding activity was monitored in both liver and oviduct of females in different reproductive stages. Females were separated into three groups; laying, nonlaying, and immature. [3H]Progesterone-binding activity levels were higher in the liver of immature than of nonlaying skates, and it was undetectable in laying skates. [3H]Progesterone binding was higher in the oviduct of laying and nonlaying skates than of immature skates. This PR-binding moiety has many characteristics of a true receptor: high affinity, low capacity, binds to DNA, and cross-reacts with antibodies against chicken PR. However, while the cytosolic form of this progesterone-binding component was quite specific for P, nuclear extracted material was nonspecific. If these progesterone-binding components are homologous with the PR A and PR B forms of other vertebrates, as we believe, it is clear that there are species differences that probably relate to phylogenetic level and physiology of the organism.
Paolucci
M
M
Department of Biology, Boston University, Massachusetts 02215, USA.
Callard
I P
IP
D000818:Animals; D001667:Binding, Competitive; D015153:Blotting, Western; D002352:Carrier Proteins / Q000032:analysis* / Q000378:metabolism; D002846:Chromatography, Affinity; D002848:Chromatography, DEAE-Cellulose; D003600:Cytosol / Q000737:chemistry / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005260:Female; D008099:Liver / Q000737:chemistry / Q000378:metabolism*; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism; D010057:Oviducts / Q000737:chemistry / Q000378:metabolism*; D010058:Oviposition / Q000502:physiology; D011374:Progesterone / Q000032:analysis / Q000378:metabolism*; D012860:Skates, Fish / Q000378:metabolism / Q000502:physiology*; D013261:Sterols / Q000145:classification / Q000378:metabolism; D014316:Tritium
D002352:Carrier Proteins; D009687:Nuclear Proteins; D013261:Sterols; D014316:Tritium; D011374:Progesterone