Class I lysine deacetylases promote glucocorticoid-induced transcriptional repression through functional interaction with LSD1.

Small molecule inhibitors of lysine deacetylases (KDACs) are approved for clinical use in treatment of several diseases. Nuclear receptors, such as the glucocorticoid receptor (GR) use lysine acetyltransferases (KATs or HATs) and KDACs to regulate transcription through acetylation and deacetylation of protein targets such as histones. Previously we have shown that KDAC1 activity facilitates GR-activated transcription at about half of all cellular target genes. In the current study we examine the role of Class I KDACs in glucocorticoid-mediated repression of gene expression. Inhibition of KDACs through two structurally distinct Class I-selective inhibitors prevented dexamethasone (Dex)-mediated transcriptional repression in a gene-selective fashion. In addition, KDAC activity is also necessary to maintain repression. Steroid receptor coactivator 2 (SRC2), which is known to play a vital role in GR-mediated repression of pro-inflammatory genes, was found to be dispensable for repression of glucocorticoid target genes sensitive to KDAC inhibition. At the promoters of these genes, KDAC inhibition did not result in altered nucleosome occupancy or histone H3 acetylation. Surprisingly, KDAC inhibition rapidly induced a significant decrease in H3K4Me2 at promoter nucleosomes with no corresponding change in H3K4Me3, suggesting the activation of the lysine demethylase, LSD1/KDM1A. Depletion of LSD1 expression via siRNA restored Dex-mediated repression in the presence of KDAC inhibitors, suggesting that LSD1 activation at these gene promoters is incompatible with transcriptional repression. Treatment with KDAC inhibitors does not alter cellular levels of LSD1 or its association with Dex-repressed gene promoters. Therefore, we conclude that Class I KDACs facilitate Dex-induced transcriptional repression by suppressing LSD1 complex activity at selected target gene promoters. Rather than facilitating repression of transcription, LSD1 opposes it in these gene contexts.

Expression of Cav1.3 calcium channel in the human and mouse colon: posttranscriptional inhibition by IFNγ.

It has been hypothesized that apically expressed L-type Ca2+ channel Cav1.3 (encoded by CACNA1D gene) contributes toward an alternative TRPV6-independent route of intestinal epithelial Ca2+ absorption, especially during digestion when high luminal concentration of Ca2+ and other nutrients limit TRPV6 contribution. We and others have implicated altered expression and activity of key mediators of intestinal and renal Ca2+ (re)absorption as contributors to negative systemic Ca2+ balance and bone loss in intestinal inflammation. Here, we investigated the effects of experimental colitis and related inflammatory mediators on colonic Cav1.3 expression. We confirmed Cav1.3 expression within the segments of the mouse and human gastrointestinal tract. Consistent with available microarray data (GEO database) from inflammatory bowel disease (IBD) patients, mouse colonic expression of Cav1.3 was significantly reduced in trinitrobenzene sulfonic acid (TNBS) colitis. In vitro, IFNγ most potently reduced Cav1.3 expression. We reproduced these findings in vivo with wild-type and Stat1-/- mice injected with IFNγ. The observed effect in Stat1-/- suggested a noncanonical transcriptional repression or a posttranscriptional mechanism. In support of the latter, we observed no effect on the cloned Cav1.3 gene promoter activity and accelerated Cav1.3 mRNA decay rate in IFNγ-treated HCT116 cells. While the relative contribution of Cav1.3 to intestinal Ca2+ absorption and its value as a therapeutic target remain to be established, we postulate that Cav1.3 downregulation in IBD may contribute to the negative systemic Ca2+ balance, to increased bone resorption, and to reduced bone mineral density in IBD patients.