Immunoprecipitation of the lutropin receptor. Loss of receptor molecules during down-regulation

Abstract

Specific anti-(lutropin receptor) antibodies were produced by immunizing rabbits with lutropin receptor purified from pseudopregnant rat ovary. The anti-receptor serum at 1:100 dilution together with anti-(rabbit gamma-globulin) serum immunoprecipitated 70% of 3H-labelled, purified lutropin receptor and 42% of 125I-chorio-gonadotropin-receptor complex. The antiserum inhibited hormone binding to rat ovarian particles. Pseudopregnant rat ovarian particles were labelled with periodate/NaB3H4 and solubilized with Triton X-100. The Triton X-100 extract was subjected to immunoprecipitation using the anti-receptor serum. When the immunoprecipitate was dissolved and analysed by polyacrylamide-gel electrophoresis in sodium dodecyl sulphate under reducing conditions followed by fluorography, a receptor polypeptide with an apparent Mr 95000 was detected. A receptor down-regulating dose of choriogonadotropin was injected into pseudopregnant rats and their ovaries were removed and homogenized 4 days later, and analysed for immunoprecipitable receptors as above. No receptor molecules were found. Accordingly, the lutropin receptor molecules actually disappear rather than merely become masked from hormone during homologous down-regulation.