Shu-chi Hsu's research while affiliated with University of Pittsburgh and other places

The restricted expression of some genes to distinct stages of the cell cycle is often brought about through alterations in the activity and/or abundance of specific transcription factors. Many cells have been shown to be unresponsive to glucocorticoid hormone action during the G2 phase of the mammalian cell cycle, suggesting that some activities of the glucocorticoid receptor (GR), a ligand-activated transcription factor, are subjected to cell cycle control. We show here that GR insensitivity in G2 is selective, affecting receptor-mediated transactivation from a simple glucocorticoid response element, but not repression from a composite glucocorticoid response element. Since glucocorticoid-dependent down-regulation of GR protein levels is also unaffected in G2, distinct activities of the receptor that participate in this homologous down-regulation must be operating as effectively in G2-synchronized cells as in asynchronous cells. Finally, the phosphorylation state of the GR is altered in G2-synchronized cells reflecting, in part, both site-specific phosphorylation and dephosphorylation events. These results suggest that, while GR may be a target for cell cycle regulated kinases and phosphatases, the resulting changes in receptor phosphorylation have an impact only on selected GR functions.

Glucocorticoid receptor (GR) nuclear translocation, transactivation and phosphorylation were examined during the cell cycle in mouse L cell fibroblasts. Glucocorticoid-dependent transactivation of the mouse mammary tumor virus promoter was observed in G0 and S phase synchronized L cells, but not in G2 synchronized cells. G2 effects were selective on the glucocorticoid hormone signal transduction pathway, since glucocorticoid but not heavy metal induction of the endogenous Metallothionein-1 gene was also impaired in G2 synchronized cells. GRs that translocate to the nucleus of G2 synchronized cells in response to dexamethasone treatment were not efficiently retained there and redistributed to the cytoplasmic compartment. In contrast, GRs bound by the glucocorticoid antagonist RU486 were efficiently retained within nuclei of G2 synchronized cells. Inefficient nuclear retention was observed for both dexamethasone- and RU486-bound GRs in L cells that actively progress through G2 following release from an S phase arrest. Finally, site-specific alterations in GR phosphorylation were observed in G2 synchronized cells suggesting that cell cycle regulation of specific protein kinases and phosphatases could influence nuclear retention, recycling and transactivation activity of the GR.