Impact of glucocorticoid receptor density on ligand- independent dimerization, cooperative ligand-binding and basal priming of transactivation : a cell culture model
Date
2013-05
Authors
Robertson, Steven
Rohwer, Johann M.
Hapgood, Janet P
Louw, Ann
Journal Title
Journal ISSN
Volume Title
Publisher
Public Library of Science -- PLoS
Abstract
Glucocorticoid receptor (GR) levels vary between tissues and individuals and are altered by physiological and
pharmacological effectors. However, the effects and implications of differences in GR concentration have not been fully
elucidated. Using three statistically different GR concentrations in transiently transfected COS-1 cells, we demonstrate, using
co-immunoprecipitation (CoIP) and fluorescent resonance energy transfer (FRET), that high levels of wild type GR (wtGR),
but not of dimerization deficient GR (GRdim), display ligand-independent dimerization. Whole-cell saturation ligand-binding
experiments furthermore establish that positive cooperative ligand-binding, with a concomitant increased ligand-binding
affinity, is facilitated by ligand-independent dimerization at high concentrations of wtGR, but not GRdim. The down-stream
consequences of ligand-independent dimerization at high concentrations of wtGR, but not GRdim, are shown to include
basal priming of the system as witnessed by ligand-independent transactivation of both a GRE-containing promoterreporter
and the endogenous glucocorticoid (GC)-responsive gene, GILZ, as well as ligand-independent loading of GR onto
the GILZ promoter. Pursuant to the basal priming of the system, addition of ligand results in a significantly greater
modulation of transactivation potency than would be expected solely from the increase in ligand-binding affinity. Thus
ligand-independent dimerization of the GR at high concentrations primes the system, through ligand-independent DNA
loading and transactivation, which together with positive cooperative ligand-binding increases the potency of GR agonists
and shifts the bio-character of partial GR agonists. Clearly GR-levels are a major factor in determining the sensitivity to GCs
and a critical factor regulating transcriptional programs.
Description
The original publication is available at www.plosone.org
Publication of this article was funded by the Stellenbosch University Open Access Fund.
Publication of this article was funded by the Stellenbosch University Open Access Fund.
Keywords
Glucocorticoids -- Receptors, Inflammatory diseases, Ligand binding (Biochemistry), DNA
Citation
Robertson, S. Rohwer, J. M. Hapgood, J. P. & Louw, A. 2013. Impact of glucocorticoid receptor density on ligand-independent dimerization, cooperative ligand-binding and basal priming of transactivation: a cell culture model. PLoS ONE, 8(5): e64831. doi:10.1371/journal.pone.0064831.