Abstract
Cross-talk between the glucocorticoid receptor (GR) and other receptors is emerging as a mechanism for fine-tuning cellular responses. We have previously shown that gonadotropin-releasing hormone (GnRH) ligand-independently activates the GR and synergistically modulates glucocorticoid-induced transcription of an endogenous gene in LβT2 pituitary gonadotrope precursor cells. Here, we investigated GR and GnRH receptor (GnRHR) cross-talk that involves co-localization with lipid rafts in LβT2 cells. We report that the GnRHR and a small population of the GR co-localize with the lipid raft protein flotillin-1 (Flot-1) at the plasma membrane and that the GR is present in a complex with Flot-1, independent of the presence of ligands. We found that the SGK-1 gene is up-regulated by Dex and GnRH alone, whereas a combination of both ligands resulted in a synergistic increase in SGK-1 mRNA levels. Using siRNA-mediated knockdown and antagonist strategies, we show that the gene-specific synergistic transcriptional response requires the GR, GnRHR, and Flot-1 as well as the protein kinase C pathway. Interestingly, although several GR cofactors are differentially recruited to the SGK-1 promoter in the presence of Dex and GnRH, GR levels remain unchanged compared with Dex treatment alone, suggesting that lipid raft association of the GR has a role in enhancing its transcriptional output in the nucleus. Finally, we show that Dex plus GnRH synergistically inhibit cell proliferation in a manner dependent on SGK-1 and Flot-1. Collectively the results support a mechanism whereby GR and GnRHR cross-talk within Flot-1-containing lipid rafts modulates cell proliferation via PKC activation and SGK-1 up-regulation.
Highlights
The glucocorticoid receptor cross-talks with other receptors to integrate cell signaling
We report that the GnRH receptor (GnRHR) and a small population of the glucocorticoid receptor (GR) co-localize with the lipid raft protein flotillin-1 (Flot-1) at the plasma membrane and that the GR is present in a complex with Flot-1, independent of the presence of ligands
Having established the co-localization of the GnRHR and Flot-1 at the plasma membrane, we investigated whether the GR is associated with Flot-1 at the membrane and whether this potential association is affected by the presence of GR agonist
Summary
The glucocorticoid receptor cross-talks with other receptors to integrate cell signaling. That the cellular responses mediated by the classical GR are much more complex and involve multiple parallel mechanisms integrating simultaneous signals from several different hormones involving cross-talk with intracellular signaling proteins. These include rapid, non-genomic, cytoplasmic GC-dependent effects (4 –9). Caveolae-associated GR has been implicated in a role for Dex-mediated intracellular communication and cell proliferation in mouse neural progenitor cells [31] These findings suggest that the localization of the GR in lipid rafts or caveolae may regulate several GR-dependent responses in a cell- and/or promoter-specific manner. In the present study we identify Flot-1-associated GR in LT2 cells and investigate its role in mediating genomic transactivation, ligand-independent GR activation by GnRH, and synergy with the GnRHR signaling pathway
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