Hormone-induced hyperphosphorylation of specific phosphorylated sites in the mouse glucocorticoid receptor.
The glucocorticoid receptor (GR) is phosphorylated in its basal state, and rapidly undergoes hormone-induced hyperphosphorylation after binding glucocorticoids. Previously, we have identified seven phosphorylated sites in the mouse GR. Most of the sites are located in the regions of the N-terminal domain that are necessary for maximum transcriptional activity and reduce nonspecific binding to DNA. Using WCL2 cells, which overexpress mouse GRs, we now quantitate hormone-induced hyperphosphorylation at each of these sites. Addition of triamcinolone acetonide to WCL2 cells results in significant hyperphosphorylation at the majority of the sites. The hyperphosphorylation ratio, i.e. the 32P incorporation into GRs from hormone-treated cells divided by 32P incorporation into GRs from untreated cells, was above 1.0 for all sites but serine 150 and threonine 159. Serine 220 displays marked hormone dependence, with a ratio of 3. For most sites the ratio was about 1.5. Hormone-induced hyperphosphorylation not only increases the charge at selected phosphorylated sites but also provides a substantial increase in the overall negative charge around the region of the N-terminal domain that is involved in transactivation.
Bodwell
J E
JE
Department of Physiology, Dartmouth Medical School, Lebanon, NH 03756-0001.
Hu
J M
JM
Orti
E
E
Munck
A
A
D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D007700:Kinetics; D051379:Mice; D010748:Phosphopeptides / Q000032:analysis; D010766:Phosphorylation / Q000187:drug effects; D011965:Receptors, Glucocorticoid / Q000378:metabolism*; D012694:Serine / Q000378:metabolism; D013912:Threonine / Q000378:metabolism; D014222:Triamcinolone Acetonide / Q000494:pharmacology*
D010748:Phosphopeptides; D011965:Receptors, Glucocorticoid; D013912:Threonine; D012694:Serine; D014222:Triamcinolone Acetonide