In vivo Imaging Analysis of SteroidNuclear Receptor Func

Hager, Gordon L.

Abstract

We studied the interaction of the progesterone receptor with the MMTV promoter in living cells, and found a pronounced ligand effect on mobility of the receptor. We compared the in vivo ligand effects with the ability of PR to carry out chromatin remodeling at the MMTV promoter in vitro. We found that ZK98229, a complete antagonist of PR, increases PR mobility in living cells, while the agonist R5020 decreases its rate of exchange on gene targets. in vitro, ZK98229 liganded receptor was unable to catalyze the SWI/SNF dependent displacement reaction, while R5020 occupied receptor was effective in the reaction. These findings suggest a direct role for chromatin remodeling in receptor movement in the nucleus.We developed an in vitro system to study nuclear receptor mobility. After permeablization of cells, we showed that receptor mobility was lost (for both PR and GR), but could be restored by the addition of highly purified molecular chaperones. In contrast, the equally mobile HP1a was refractory to addition of chaperones. These findings introduce a new paradigm for chaperone involvement in nuclear protein mobility.

Funding Agency

• Agency: National Institute of Health (NIH)
• Institute: Division of Basic Sciences - NCI (NCI)
• Type: Intramural Research (Z01)
• Project #: 1Z01BC010308-07
• Application #: 7291698
• Study Section: (LRBG)
• Support Year: 7
• Fiscal Year: 2005

Institution

• Name: Basic Sciences
• Country: United States

Publications

Stavreva, Diana A; Wiench, Malgorzata; John, Sam et al. (2009) Ultradian hormone stimulation induces glucocorticoid receptor-mediated pulses of gene transcription. Nat Cell Biol 11:1093-102

Meijsing, Sebastiaan H; Elbi, Cem; Luecke, Hans F et al. (2007) The ligand binding domain controls glucocorticoid receptor dynamics independent of ligand release. Mol Cell Biol 27:2442-51

Huh, Jung-Im; Qiu, Ting Hu; Chandramouli, Gadisetti V R et al. (2007) 2-methoxyestradiol induces mammary gland differentiation through amphiregulin-epithelial growth factor receptor-mediated signaling: molecular distinctions from the mammary gland of pregnant mice. Endocrinology 148:1266-77

Klokk, Tove I; Kurys, Piotr; Elbi, Cem et al. (2007) Ligand-specific dynamics of the androgen receptor at its response element in living cells. Mol Cell Biol 27:1823-43

Voss, Ty C; John, Sam; Hager, Gordon L (2006) Single-cell analysis of glucocorticoid receptor action reveals that stochastic post-chromatin association mechanisms regulate ligand-specific transcription. Mol Endocrinol 20:2641-55

Martinez, Elisabeth D; Rayasam, Geetha V; Dull, Angie B et al. (2005) An estrogen receptor chimera senses ligands by nuclear translocation. J Steroid Biochem Mol Biol 97:307-21

Rayasam, Geetha V; Elbi, Cem; Walker, Dawn A et al. (2005) Ligand-specific dynamics of the progesterone receptor in living cells and during chromatin remodeling in vitro. Mol Cell Biol 25:2406-18

Becker, Matthias; Becker, Antje; Miyara, Faical et al. (2005) Differential in vivo binding dynamics of somatic and oocyte-specific linker histones in oocytes and during ES cell nuclear transfer. Mol Biol Cell 16:3887-95

Dohoney, Kathleen M; Guillerm, Claire; Whiteford, Craig et al. (2004) Phosphorylation of p53 at serine 37 is important for transcriptional activity and regulation in response to DNA damage. Oncogene 23:49-57

Elbi, Cem; Walker, Dawn A; Lewis, Marcia et al. (2004) A novel in situ assay for the identification and characterization of soluble nuclear mobility factors. Sci STKE 2004:pl10