الفهرس 
Antibodies to the pr1 receptorOnly 14 pages are availabe for public view
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Abstract
Prl receptors in five target tissues with distinct responses to Prl and different cellular locations of the receptor were characterized. Prl receptor from female rat liver was purified 168-2200-fold with an average recovery of 20-100% and a net yield of 0.03-0.23 mg protein from -1 gm of membranes. About 3-5 other proteins co-purified with Prl receptor. The molecular weight of the partially purified receptor was 40 KD under reducing conditions. Prl-binding activity in CHAPS extracts and the partially purified receptor was assessed by a PEI-filtration assay we developed. Over 90% of the 125~-~rl-receptcoorm plexes at 0.1% CHAPS were retained on the filters. This assay gave over a three-fold increase in the amount of receptor recovered compared to the traditional PEG assay. Polyclonal antibodies to the partially purified receptor were produced in guinea pigs. These antibodies inhibited Prlbinding in membranes from rat liver, DMBA-induced mammary tumor, lactating mammary gland and R3230AC mammary tumor indicating that these Prl receptors share antigenic determinants at or close to the hormone recognition domain. The antibodies also recognized antigenic epitopes (or associated proteins) on the receptor of these tissues adjacent or distant to the Prl-binding domain. The differential potency of the antibodies in recognizing Prl receptor indicated that there were some antigenic differences in the receptor among the tissues. The antibodies recognized two peptides with Mrls of 43 KD and 30 KD in DMBA-induced mammary tumor cells. The 43 KD is likely the Prl-binding peptide since the Mr of Prl receptor determined by affinity cross-linking and gel filtration in this tissue was 40-42 KD. The 30 KD peptide could be a degradative product of the 43 KD protein or another protein with high affinity for the receptor or an unrelated antigen. Using affinity cross-linking, Prl receptor in rat liver, and the surface and internal receptor in DMBA-induced mammary tumor had a molecular weight of 40 KD (reduced). In contrast, Prl receptor in lactating mammary gland and the R3230AC mammary tumor had a Mr of -38 KD (reduced and non-reduced). In Prl-dependent Nb, lymphoma cells, the Prl receptor in membranes had two major f oms; 73 KD and 66 KD, and 73 KD, 66 KD, 40 KD and 16 KD peptides in CHAPS extracts. Using Sephacryl- 300 gel chromatography, the Prl-binding peptide had a Mr of -42 KD in some tissues, while most of the Prl-receptor complexes were aggregated (180 KD to over 800 KD) . This aggregation is due to the low concentration of the detergent (0.1% CHAPS) in the column buffer. Prl receptor in the different tissues showed charge heterogeneity. In rat liver where most of Prl receptors are intracellular, the receptor is more negatively charged compared to the cell surface receptor in Nb, lymphoma cells. In cultured DMBA-inducedmammarytumor cells, Prl receptors exhibited two charges; the surface receptor is less negatively charged than the internal one. This indicates that when Prl receptor translocates from the cell interior to the cell surface upon energy depletion it becomes less negatively charged. This translocation did not lead to any detectable change in the Mr of the receptor. In lactating mammary gland, Prl receptor is more negatively charged than the surface receptor in Nb, cells and DMBA-induced mammary tumor cells. In R3230AC mammary tumor Prl receptor eluted in two isoforms. However, the location and etiology of the differences in these isoforms are not known. We conclude that: (1) PEI-filtration assay is a good alternative to the traditional PEG method to assess Prl-binding activity in CHAPS extracts; (2) Differences in the molecular weight and antigenicity of Prl receptors in these tissues suggest that there are structural differences in the receptors. These differences may relate to alternate mechanisms(s) for Prl action; (3) charge differences in Prl receptor in these tissues may be important in receptor trafficking and their predominant cellular location.