Size and steroid-binding characterization of membrane-associated glucocorticoid receptor in S-49 lymphoma cells.
The precise mechanism for glucocorticoid-mediated lymphocytolysis is not understood, although it is presumed to be receptor mediated. We have recently presented evidence that this response is mediated by a specialized form of the glucocorticoid receptor (GR) that resides in the plasma membrane (mGR). Confirmation of the previous receptor identification studies in a population of S-49 cells enriched for mGR is now made using another antibody specific for the rodent GR, BUGR-2. The membrane resident receptor could be labeled competitively with the affinity ligand dexamethasone 21-mesylate, and Scatchard analysis of whole cell binding revealed that receptor number, but not the affinity for hormone, varied between the mGR-enriched and -deficient cell populations. Steroid specificity displacement analyses showed an order of affinities as follows: triamcinolone acetonide greater than progesterone greater than dexamethasone greater than testosterone = estrogen. Studies of mGR by one- and two-dimensional gel electrophoresis, immunoblot, autoradiography, and density gradients revealed a species with an equivalent size to cytosolic receptor as well as multiple higher molecular weight species, confirming earlier studies. To offer a possible explanation for the nucleic acid origins of the mGR, RNA from the mGR-enriched cells was probed with rat GR cDNA; mGR-enriched cells contained higher levels of GR mRNA. Possible molecular etiologies of larger receptor species in membrane are discussed.
Gametchu
B
B
Department of Pediatrics, Medical College of Wisconsin, Milwaukee 53226.
Watson
C S
CS
Pasko
D
D
D000345:Affinity Labels; D001665:Binding Sites; D001667:Binding, Competitive; D015152:Blotting, Northern; D002462:Cell Membrane / Q000378:metabolism*; D002499:Centrifugation, Density Gradient; D003907:Dexamethasone / Q000031:analogs & derivatives / Q000378:metabolism; D015151:Immunoblotting; D008223:Lymphoma / Q000378:metabolism*; D008970:Molecular Weight; D012333:RNA, Messenger / Q000032:analysis; D011965:Receptors, Glucocorticoid / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D000345:Affinity Labels; D012333:RNA, Messenger; D011965:Receptors, Glucocorticoid; D003907:Dexamethasone; C031710:dexamethasone 21-methanesulfonate