Abstract
Heat shock and other forms of stress increase glucocorticoid receptor (GR) activity in cells, suggesting cross-talk between the heat shock and GR signal pathways. An unresolved question concerning this cross-talk is whether heat shock factor (HSF1) activity is required for this response. We addressed this issue by modulating HSF1 activity with compounds acting by distinct mechanisms: sodium vanadate (SV), an inhibitor of protein phosphatases; and wortmannin, an inhibitor of DNA-dependent protein kinase. Using HSF1- and GR-responsive CAT reporters, we demonstrate that SV inhibits both HSF1 activity and the stress potentiation of GR, while having no effect on the hormone-free GR or HSF1. Paradoxically, SV increased hormone-induced GR activity in the absence of stress. In contrast, wortmannin increased HSF1 activity in stressed cells and had no effect on HSF1 in the absence of stress. Using the pMMTV-CAT reporter containing the negative regulatory element 1 site for DNA-dependent protein kinase, wortmannin was found to increase the GR response. However, in cells expressing a minimal promoter lacking negative regulatory element 1 sites, wortmannin had no effect on the GR in the absence of stress but increased the stress potentiation of GR. Our results show that the mechanism by which GR activity is increased in stressed cells requires intrinsic HSF1 activity.
Highlights
The glucocorticoid receptor (GR)1 is a member of the nuclear receptor family of proteins that act as hormone-activated transcription factors [1, 2]
To accurately measure the effects of sodium vanadate (SV) on this HSF1 function, we have stably transfected L929 cells with the p2500-chloramphenicol acetyltransferase (CAT) construct (LHSECAT cells), in which high level CAT gene expression is controlled by the human HSP70 promoter [36]
In recent work from our laboratory [40], we obtained our first evidence of a correlation between HSF1 activity and the heat shock potentiation of GR by demonstrating a similar pattern of HSF1 and GR activity in stressed cells and by showing that inhibition of HSF1 activity by the flavonoid compound quercetin results in a similar inhibition of the stress effect on the GR
Summary
The glucocorticoid receptor (GR) is a member of the nuclear receptor family of proteins that act as hormone-activated transcription factors [1, 2]. The potentiation of hormone-induced GR function by heat shock was observed at saturating concentrations of dexamethasone (Dex) agonist (1 M) and was observed in cells stably transfected with complex (pMMTV-CAT) or minimal (pGRE2E1BCAT) promoters controlling expression of CAT [13] These observations suggest that the GR-mediated response typically seen in unstressed cells is not maximal, even in the presence of hormone concentrations that yield saturation results for reporter gene expression. We report here that wortmannin treatment of intact cells results in a marked enhancement of HSF1-mediated CAT expression, but only under stress conditions, suggesting that DNA-PK does act as a negative regulator of HSF1. In cells stably transfected with a GR-responsive CAT reporter, wortmannin treatment caused a corresponding increase in the stress potentiation of GR, under conditions where wortmannin had no effect on GR signaling in the absence of stress
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
