title: Evidence for the role of phospholipids in follicle-stimulating hormone binding to membrane-bound and soluble receptors from calf testis.
abstract: The role of phospholipids in the interaction of FSH with receptors in the calf testis was explored by studying the effects of phospholipase-C (PL-C) on the binding of radioiodinated human FSH ([125I]iodo-hFSH) to three previously characterized but different preparations of FSH receptor: a membrane fraction derived from calf testis homogenate, a buffer-soluble receptor present in the cytosol of the calf testis homogenate, and a detergent-soluble receptor prepared from the membrane fraction by extraction with Triton X-100. Prior incubation with PL-C markedly reduced specific [125I]iodo-hFSH binding to the membrane-bound and buffer-soluble receptors. This loss in binding was associated with hydrolysis of phospholipids, was prevented by the addition of o-phenanthroline, an inhibitor of PL-C, but not by the addition of a protease inhibitor, and could not be reproduced by the addition of the products of PL-C hydrolysis. Enzyme treatment did not dissociate [125I]iodo-hFSH already bound to the buffer-soluble receptor and dissociated only 20% of [125I]iodo-hFSH bound to membrane receptor. PL-C treatment did not reduce [125I]iodo-hFSH binding to detergent-solubilized receptor, nor did it hydrolyze constituent phospholipids. The apparent resistance of the detergent-solubilized receptor to PL-C treatment was studied by incubating membranes with or without PL-C before detergent solubilization. Triton X-100 itself (2%) inhibited phospholipid hydrolysis by PL-C, but this could be overcome by reducing the detergent concentration 10-fold by dilution. Despite a marked decrease in specific [125I]iodo-hFSH binding to PL-C-treated membranes compared to that of untreated controls, the total amount of [125I]iodo-hFSH binding to Triton X-100 extracts of each membrane preparations was not significantly different. Addition of Triton X-100 to the buffer-soluble receptor restored 40% of the hormone binding that had initially been lost concomitant with enzymic hydrolysis of membrane phospholipids. It appears that constituent phospholipids play an important role in the interaction of FSH with membrane-bound or solubilized receptor and that Triton X-100 is able to substitute, although imperfectly, for phospholipids in that regard, probably because of its related amphipathic properties.
mesh_terms: D000818:Animals; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D005640:Follicle Stimulating Hormone / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D008563:Membrane Lipids / Q000502:physiology*; D010618:Phenanthrolines / Q000494:pharmacology; D010743:Phospholipids / Q000502:physiology*; D011956:Receptors, Cell Surface / Q000187:drug effects / Q000378:metabolism*; D011962:Receptors, FSH; D013737:Testis / Q000378:metabolism*; D010738:Type C Phospholipases / Q000494:pharmacology
chemical_list: D008563:Membrane Lipids; D010618:Phenanthrolines; D010743:Phospholipids; D011956:Receptors, Cell Surface; D011962:Receptors, FSH; D005640:Follicle Stimulating Hormone; D010738:Type C Phospholipases; C025205:1,10-phenanthroline