Abstract
BackgroundIt has recently been found that both nuclear epithelial-expressed histone deacetylases Hdac1 and Hdac2 are important to insure intestinal homeostasis and control the mucosal inflammatory response in vivo. In addition, HDAC inhibitors modulate epithelial cell inflammatory responses in cancer cells. However, little is known of the specific role of different HDAC, notably Hdac1, in the regulation of inflammatory gene expression in intestinal epithelial cells (IEC).MethodsWe investigated the role of Hdac1 in non-transformed IEC-6 rat cells infected with lentiviral vectors expressing specific Hdac1 shRNAs, to suppress Hdac1 expression. Proliferation was assessed by cell counting. Deacetylase activity was measured with a colorimetric HDAC assay. Cells were treated with IL-1β and/or the JQ1 bromodomain acetyl-binding inhibitor. Nuclear protein levels of Hdac1, Hdac2, phosphorylated or unphosphorylated NF-κB p65 or C/EBPβ, and NF-κB p50 and actin were determined by Western blot. Chemokine and acute phase protein expression was assessed by semi-quantitative RT-PCR analysis. Secreted cytokine and chemokine levels were assessed with a protein array. Chromatin immunoprecipitation experiments were done to assess RNA polymerase II recruitment.ResultsReduced Hdac1 protein levels led to Hdac2 protein increases and decreased cell proliferation. Hdac1 depletion prolonged nuclear IL-1β-induced phosphorylation of NF-κB p65 protein on Ser536 as opposed to total p65, and of C/EBPβ on Ser105. In addition, semi-quantitative RT-PCR analysis revealed three patterns of expression caused by Hdac1 depletion, namely increased basal and IL-1β-stimulated levels (Hp, Kng1), increased IL-1β-stimulated levels (Cxcl2) and decreased basal levels with normal IL-1β induction levels (Ccl2, Ccl5, Cxcl1, C3). Secreted cytokine and chemokine measurements confirmed that Hdac1 played roles both as an IL-1β signalling repressor and activator. Hdac1 depletion did not alter the JQ1 dependent inhibition of basal and IL-1β-induced inflammatory gene expression. Hdac1 depletion led to decreased basal levels of RNA polymerase II enrichment on the Ccl2 promoter, as opposed to the Gapdh promoter, correlating with decreased Ccl2 basal mRNA expression.ConclusionsHdac1 is a major nuclear HDAC controlling IL-1β-dependent inflammatory response in IEC, notably by regulating gene-specific transcriptional responses. Hdac1 may be important in restricting basal and inflammatory-induced gene levels to defined ranges of expression.Electronic supplementary materialThe online version of this article (doi:10.1186/s12950-014-0043-2) contains supplementary material, which is available to authorized users.
Highlights
It has recently been found that both nuclear epithelial-expressed histone deacetylases Hdac1 and Hdac2 are important to insure intestinal homeostasis and control the mucosal inflammatory response in vivo
Hdac1 depletion reduces intestinal epithelial cells (IEC)-6 cell proliferation To determine the role of the histone deacetylase Hdac1 in IEC, we infected IEC-6 cells with a retroviral construct expressing a selected short hairpin RNA (shRNA) against Hdac1
While Hdac2 mRNA levels were not affected, Hdac2 protein levels were increased (Figure 1B). This post-transcriptional increase in Hdac2 protein levels after Hdac1 depletion was observed in other cell types [11]
Summary
It has recently been found that both nuclear epithelial-expressed histone deacetylases Hdac and Hdac are important to insure intestinal homeostasis and control the mucosal inflammatory response in vivo. Little is known of the specific role of different HDAC, notably Hdac, in the regulation of inflammatory gene expression in intestinal epithelial cells (IEC). Histone acetylation plays a dual role, either by reducing histone-DNA interactions, thereby creating an open chromatin configuration, or by acting as an anchor recognized by bromodomaincontaining chromatin modifiers [8]. Some of these recruited proteins, such as Brd family members, play important roles in the regulation of transcriptional elongation as well as inflammation. HDAC-dependent regulation of protein acetylation levels leads to cell- and gene-specific transcriptional repression or activation [8]
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