title: Subcellular fractionation of the ovine corpus luteum: association of progesterone with ovine luteal membranes?
abstract: Sheep corpus luteum homogenates were fractionated by centrifugation on continuous sucrose density gradients, with or without digitonin, and gradient fractions were assayed for progesterone, and for a range of intracellular organelle and plasma-membrane markers. Digitonin had little effect on the density distributions of mitochondrial, rough endoplasmic reticulum (RER) and Golgi-endoplasmic reticulum-lysosomal (GERL) membranes. However, digitonin did disrupt lysosomal membranes, leading to release of acid hydrolases, and induced a decrease in buoyant density of NADH-cytochrome c reductase, a putative smooth endoplasmic reticulum (SER) marker. Oxytocin-containing granules were clearly resolved from other organelles accumulating in a sharp peak (density, 1.20 g/cm3). Luteal cell-surface membrane marker activities equilibrated at similar buoyant densities in control gradients, and pretreatment with digitonin induced a marked increase in their buoyant densities. The majority of the progesterone of the sheep corpus luteum equilibrated at a buoyant density of 1.10 g/cm3 in control gradients, and was highly perturbed by digitonin. These fractions also accumulated [3H]progesterone. The buoyant density profile of progesterone in both control and digitonin-treated gradients most closely resembled that of sheep luteal lactogenic receptor, a putative plasma-membrane marker.
authors.affiliation: Department of Obstetrics and Gynaecology, Centre for Reproductive Biology, Edinburgh, U.K.
authors.forename: T A
authors.initials: TA
authors.lastname: Bramley
authors.forename: G S
authors.initials: GS
authors.lastname: Menzies
mesh_terms: D000818:Animals; D002458:Cell Fractionation; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D003338:Corpus Luteum / Q000166:cytology* / Q000378:metabolism / Q000648:ultrastructure; D004072:Digitonin / Q000494:pharmacology; D005260:Female; D007425:Intracellular Membranes / Q000187:drug effects / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D015388:Organelles / Q000648:ultrastructure; D011374:Progesterone / Q000378:metabolism*; D012756:Sheep
chemical_list: D008565:Membrane Proteins; D011374:Progesterone; D004072:Digitonin