Changes in chromatin structure and methylation of the human interleukin‐1β gene during monopoiesis

Abstract

Summary Interleukin‐1β (IL‐1β) induces the expression of a variety of proteins responsible for acute inflammation and chronic inflammatory diseases. However, the molecular regulation of IL‐1β expression in myeloid differentiation has not been elucidated. In this study the chromatin structure of the IL‐1β promoter and the impact of methylation on IL‐1β expression in monocytic development were examined. The results revealed that the IL‐1β promoter was inaccessible in undifferentiated promyeloid HL‐60 cells but highly accessible in differentiated monocytic cells which additionally acquired the ability to produce IL‐1β. Accessibilities of differentiated cells were comparable to those of primary monocytes. Lipopolysaccharide (LPS) stimulation did not affect promoter accessibility in promyeloid and monocytic HL‐60 cells, demonstrating that the chromatin remodelling of the IL‐1β promoter depends on differentiation and not on the transcriptional status of the cell. Demethylation via 5‐aza‐2′‐deoxycytodine led to the induction of IL‐1β expression in undifferentiated and differentiated cells, which could be increased after LPS stimulation. Our data indicate that the IL‐1β promoter is reorganized into an open poised conformation during monopoiesis being a privilege of mature monocytes but not of the entire myeloid lineage. As a second mechanism, IL‐1β expression is regulated by methylation acting independently of the developmental stage of myeloid cells.

Early Inhibition of IL-1β Expression by IFN-γ Is Mediated by Impaired Binding of NF-κB to the IL-1β Promoter but Is Independent of Nitric Oxide

“…In resting monocytes and macrophages, the IL-1b promoter is packaged into a nucleosome-free, highly accessible chromatin structure that remains largely unchanged upon stimulation, whereas the IL-1b promoter is inaccessible in undifferentiated promyeloid cells, as well as in primary B and T cells (6,34,42). Moreover, several transcription factors, including IRF8, Pu.1, nonphosphorylated STAT1, and C/EBPb, were described to be constitutively bound to the IL-1b promoter and/or enhancer elements, enabling rapid gene transcription upon cellular activation (6,34,(43)(44)(45).…”

Zinc deficiency induces production of the proinflammatory cytokines IL-1β and TNFα in promyeloid cells via epigenetic and redox-dependent mechanisms

“…SDS-polyacrylamide gel electrophoresis using an equivalent of 4×10 5 cells and Western Blot analysis were performed as described previously [27]. Membranes were incubated with horseradish-peroxidase (HRP)-linked anti-rabbit IgG secondary antibody and HRP-coupled anti-biotin antibody for detection of biotinlabeled molecular weight standard for 1 h, followed by detection with LumiGlo reagent (Cell Signaling Technology) on a LAS-3000 (Fujifilm Lifescience, Düsseldorf, Germany).…”

“…As a zincsufficient control, chelexed medium was supplemented with 8 μM ZnSO 4 (HL-60 CHE + ). The differentiation of HL-60 cells into monocytic cells using 1α,25-dihydroxyvitamin D 3 (VD3; 100 nM) for 72 h was performed and monitored as described [25,27].…”

“…MNase accessibility assays were performed, and results were plotted as described previously [27]. Real-time PCR was performed in 25-μl reaction volumes in duplicates using Brilliant Sybr Green qPCR Master Mix (Applied Biosystems) containing 100 ng of DNA.…”

“…A total of 2×10 6 cells were lysed and sonicated in 100 μl lysis buffer [0.5 M Tris-HCl (pH 6.8), 26.6% glycerin, 10% sodium dodecyl sulfate (SDS), 1 mM Na 3 VO 4 and 1% β-mercaptoethanol] [27]. SDS-polyacrylamide gel electrophoresis using an equivalent of 4×10 5 cells and Western Blot analysis were performed as described previously [27].…”