Glucocorticoid repression of inflammatory gene expression shows differential responsiveness by transactivation- and transrepression-dependent mechanisms.
Binding of glucocorticoid to the glucocorticoid receptor (GR/NR3C1) may repress inflammatory gene transcription via direct, protein synthesis-independent processes (transrepression), or by activating transcription (transactivation) of multiple anti-inflammatory/repressive factors. Using human pulmonary A549 cells, we showed that 34 out of 39 IL-1β-inducible mRNAs were repressed to varying degrees by the synthetic glucocorticoid, dexamethasone. Whilst these repressive effects were GR-dependent, they did not correlate with either the magnitude of IL-1β-inducibility or the NF-κB-dependence of the inflammatory genes. This suggests that induction by IL-1β and repression by dexamethasone are independent events. Roles for transactivation were investigated using the protein synthesis inhibitor, cycloheximide. However, cycloheximide reduced the IL-1β-dependent expression of 13 mRNAs, which, along with the 5 not showing repression by dexamethasone, were not analysed further. Of the remaining 21 inflammatory mRNAs, cycloheximide significantly attenuated the dexamethasone-dependent repression of 11 mRNAs that also showed a marked time-dependence to their repression. Such effects are consistent with repression occurring via the de novo synthesis of a new product, or products, which subsequently cause repression (i.e., repression via a transactivation mechanism). Conversely, 10 mRNAs showed completely cycloheximide-independent, and time-independent, repression by dexamethasone. This is consistent with direct GR transrepression. Importantly, the inflammatory mRNAs showing attenuated repression by dexamethasone in the presence of cycloheximide also showed a significantly greater extent of repression and a higher potency to dexamethasone compared to those mRNAs showing cycloheximide-independent repression. This suggests that the repression of inflammatory mRNAs by GR transactivation-dependent mechanisms accounts for the greatest levels of repression and the most potent repression by dexamethasone. In conclusion, our data indicate roles for both transrepression and transactivation in the glucocorticoid-dependent repression of inflammatory gene expression. However, transactivation appears to account for the more potent and efficacious mechanism of repression by glucocorticoids on these IL-1β-induced genes.
Airways Inflammation Research Group, Snyder Institute for Chronic Diseases, Faculty of Medicine, University of Calgary, Calgary, Alberta, Canada.
Elizabeth M
EM
King
Joanna E
JE
Chivers
Christopher F
CF
Rider
Anne
A
Minnich
Mark A
MA
Giembycz
Robert
R
Newton
D015153:Blotting, Western; D045744:Cell Line, Tumor; D003513:Cycloheximide / Q000494:pharmacology; D016207:Cytokines / Q000235:genetics*; D003907:Dexamethasone / Q000494:pharmacology*; D015870:Gene Expression / Q000187:drug effects*; D020869:Gene Expression Profiling; D005938:Glucocorticoids / Q000494:pharmacology; D006801:Humans; D018836:Inflammation Mediators / Q000378:metabolism*; D053583:Interleukin-1beta / Q000494:pharmacology; D016014:Linear Models; D016328:NF-kappa B / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D034622:RNA Interference; D011965:Receptors, Glucocorticoid / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D015533:Transcriptional Activation / Q000187:drug effects*
D016207:Cytokines; D005938:Glucocorticoids; D018836:Inflammation Mediators; D053583:Interleukin-1beta; D016328:NF-kappa B; C497713:NR3C1 protein, human; D011500:Protein Synthesis Inhibitors; D011965:Receptors, Glucocorticoid; D003907:Dexamethasone; D003513:Cycloheximide