Subcellular fractionation of the human corpus luteum: distribution of GnRH agonist binding sites.
We have investigated the subcellular localization of GnRH agonist (GnRHA) binding sites in the human corpus luteum. Corpora lutea (CL) were obtained from 13 women undergoing laparotomy during the luteal phase of the menstrual cycle. CL homogenates were subjected to subcellular fractionation by either differential rate centrifugation or by sucrose density gradient fractionation with or without digitonin perturbation. Fractions were then assayed for a number of marker activities characteristic of the major intracellular organelles and cell-surface membranes, and for specific binding of [125I]GnRHA. Specific binding of [125I]GnRH agonist was greatest in regions of the gradient (density, 1.12-1.15 g/cm3) enriched in cell-surface and endoplasmic reticulum membrane markers. Little or no GnRHA binding was associated with fractions enriched in nuclei, mitochondria or lysosomes. Digitonin treatment demonstrated that the majority of GnRHA binding sites were associated with luteal cell plasma-membranes: however, some GnRHA binding was also associated with endoplasmic reticulum. These results indicate that: (i) human luteal GnRHA binding sites were not associated with lysosomal proteases, (ii) a significant fraction (59%) of GnRHA binding sites was localised on the luteal cell-surface membrane. Specific GnRHA binding to both cell-surface and endoplasmic reticulum membranes may indicate an extensive turnover of membrane binding sites.
D001665:Binding Sites; D002064:Buserelin / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D003338:Corpus Luteum / Q000378:metabolism*; D004072:Digitonin; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005260:Female; D006801:Humans; D009940:Organoids / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism
D004072:Digitonin; D002064:Buserelin