B. Gametchu et al., USE OF RECEPTOR ANTIBODIES TO DEMONSTRATE MEMBRANE GLUCOCORTICOID RECEPTOR IN CELLS FROM HUMAN LEUKEMIC PATIENTS, The FASEB journal, 7(13), 1993, pp. 1283-1292
Anti-peptide antibody to the human glucocorticoid receptor (GR) was pr
oduced and used to demonstrate that a subset of the GR population resi
des in the plasma membrane of human leukemic cells. Characterization o
f the antibody with intracellular GR (iGR) showed its ability to shift
[H-3]triamcinolone acetonide-labeled GR (4S protein) from two human l
eukemic cell lines to a higher density in sucrose gradients; Western a
nd autoradiographic analysis of affinity-labeled ([H-3]dexamethasone 2
1-mesylate) receptor revealed an immunoreactive and competitively labe
led band of 94 kDa. CCRF-CEM cell membrane GR (mGR) resolved as a >7S
protein on density gradients and immunoselected cell surface protein l
abeled by whole cell biotinylation or affinity-labeling with [H-3]dexa
methasone 21-mesylate was approximately 145 kDa, demonstrating that mG
R was larger in size than iGR, as has been shown previously for the mG
R of mouse lymphoma cells. Analysis of mGR in lymphocytes of leukemic
patients and the CCRF-CEM cell line indicated differences in levels of
expression as shown by FACS and immunocytochemical analyses. We are c
urrently using this system to study the correlation between the quanti
ty of membrane-resident GRs and the glucocorticoid-induced lytic respo
nse, a relationship previously shown in the murine (S-49 cell) system.