title: Homodimerization of glucocorticoid receptor from single cells investigated using fluorescence correlation spectroscopy and microwells.
abstract: Glucocorticoid receptor α (GR) binds to the promoter regions of target genes as a homodimer and activates its transcriptional process. Though the homodimerization is thought to be the initial and essential process, the dissociation constant for homodimerization of GR remains controversial. To quantify homodimerization of (enhanced green fluorescence protein) EGFP-(glucocorticoid receptor) GR, the particle brightness in lysates from single cell was estimated for the fraction of homodimeric EGFP-GR using fluorescence correlation spectroscopy and microwells. Fitting the data with a bimolecular reaction model, the dissociation constant was determined. Moreover slow-diffusion complex was observed. These results suggest that EGFP-GR forms not only a monomer-dimer equivalent state but also a large-molecular-weight complex.
authors.affiliation: Laboratory of Molecular Cell Dynamics, Faculty of Advanced Life Science, Hokkaido Univesity, Sapporo 001-0021, Japan.
authors.forename: Sho
authors.initials: S
authors.lastname: Oasa
authors.affiliation: Bio-Analytical Research Group, Biomedical Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), Ibaraki 305-8566, Japan.
authors.forename: Akira
authors.initials: A
authors.lastname: Sasaki
authors.affiliation: Laboratory of Molecular Cell Dynamics, Faculty of Advanced Life Science, Hokkaido Univesity, Sapporo 001-0021, Japan.
authors.forename: Johtaro
authors.initials: J
authors.lastname: Yamamoto
authors.affiliation: Laboratory of Molecular Cell Dynamics, Faculty of Advanced Life Science, Hokkaido Univesity, Sapporo 001-0021, Japan.
authors.forename: Shintaro
authors.initials: S
authors.lastname: Mikuni
authors.affiliation: Laboratory of Molecular Cell Dynamics, Faculty of Advanced Life Science, Hokkaido Univesity, Sapporo 001-0021, Japan. Electronic address: kinjo@sci.hokudai.ac.jp.
authors.forename: Masataka
authors.initials: M
authors.lastname: Kinjo
mesh_terms: D000465:Algorithms; D015153:Blotting, Western; D004129:Dimethylpolysiloxanes / Q000737:chemistry; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D018613:Microscopy, Confocal; D055503:Protein Multimerization; D011965:Receptors, Glucocorticoid / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D059010:Single-Cell Analysis / Q000295:instrumentation / Q000379:methods*; D013050:Spectrometry, Fluorescence / Q000379:methods*
chemical_list: D004129:Dimethylpolysiloxanes; D011965:Receptors, Glucocorticoid; D049452:Green Fluorescent Proteins; C013830:baysilon
keywords: Dissociation constant; Fluorescence correlation spectroscopy; Glucocorticoid receptor; Homodimerization; Microwell; Single cell