Labelled regulatory elements are pervasive features of the macrophage genome and are dynamically utilized by classical and alternative polarization signals

Abstract

The concept of tissue-specific gene expression posits that lineage-determining transcription factors (LDTFs) determine the open chromatin profile of a cell via collaborative binding, providing molecular beacons to signal-dependent transcription factors (SDTFs). However, the guiding principles of LDTF binding, chromatin accessibility and enhancer activity have not yet been systematically evaluated. We sought to study these features of the macrophage genome by the combination of experimental (ChIP-seq, ATAC-seq and GRO-seq) and computational approaches. We show that Random Forest and Support Vector Regression machine learning methods can accurately predict chromatin accessibility using the binding patterns of the LDTF PU.1 and four other key TFs of macrophages (IRF8, JUNB, CEBPA and RUNX1). Any of these TFs alone were not sufficient to predict open chromatin, indicating that TF binding is widespread at closed or weakly opened chromatin regions. Analysis of the PU.1 cistrome revealed that two-thirds of PU.1 binding occurs at low accessible chromatin. We termed these sites labelled regulatory elements (LREs), which may represent a dormant state of a future enhancer and contribute to macrophage cellular plasticity. Collectively, our work demonstrates the existence of LREs occupied by various key TFs, regulating specific gene expression programs triggered by divergent macrophage polarizing stimuli.

HDAC3 Mediates the Inflammatory Response and LPS Tolerance in Human Monocytes and Macrophages

“…We are unsure why this is the case. While searching for transcription factors bound by HDAC3, we observed in the data published by Czimmerer et al that HDAC3 bound upstream of the IκBα-encoding genes Nfkbiz and Nfkbia in mice, both of which were also upregulated in HDAC3i pre-treated macrophages ( Supplemental Figures 6A, B ) ( 51 , 52 ). As IκBα protein prevents NFκB nuclear translocation and gene expression in both in human and mice, we speculate that this mechanism explains how HDAC3 negatively regulates NFκB activity and that HDAC3i reduces expression of NFkB dependent pro-inflammatory genes as measured in this study.…”

“…To understand whether HDAC3 directly affects transcription of TFs, we analyzed ChIP-sequencing data on HDAC3 binding in mouse M-CSF differentiated bone marrow derived macrophages (BMDMs) obtained from GSE106701 ( 51 , 52 ). Combining the reported peaks from samples GSM2845618 and GSM2845619 with the differential expression analysis on TFs indicated that mouse orthologs of the differentially expressed TF genes HIVEP2 , ELK3 , IRF7 , NFKB1 , ARID5B , REL , KLF7 , ETV3 and MXD1 were bound by HDAC3, with one peak associated to the intergenic region of HIVEP2 belonging to the top 2.5% strongest peaks ( Figure 6C and Supplemental Table 4 ).…”

“…Taken together, many differentially expressed TFs appear subject to HDAC3 activity, and could therefore explain the functional effects of HDAC3i. However, further analyses of HDAC3 DNA interaction did not reveal direct promotor binding of these affected TF ( Supplemental Figure 5 ) ( 51 , 52 ), suggesting that HDAC3i likely affects TF gene expression indirectly.…”

“…The transcription factors were obtained from the supplementary files published alongside Lambert et al ( 50 ) and used to filter the differentially expressed genes for differentially expressed transcription factors. HDAC3 chromatin immunoprecipitation sequencing (ChIP-seq) peaks were obtained from untreated mice bone marrow derived macrophages (BMDMs) as stored in GSM2845618 and GSM2845619 from the dataset GSE106701 ( 51 , 52 ). The ChIP-seq peak scores were log 2 transformed and the mean was taken across samples.…”

Unorthodox Transcriptional Mechanisms of Lipid-Sensing Nuclear Receptors in Macrophages: Are We Opening a New Chapter?

“…The macrophage subset-specific enhancer repertoires serve as a binding platform for the signal-dependent transcription factors (SDTFs). Many microenvironmental signals including pathogenderived molecules, cytokines, and lipids can activate SDTFs turning on signal-specific gene expression programs (29)(30)(31)(32). Toll-like receptor (TLR) ligands such as LPS and poly(I:C) as well as tumor necrosis factors (TNFs) activate nuclear factor kappa-lightchain-enhancer of activated B cells (NFkB) and Activator protein 1 (AP-1) transcription factor complexes initiating a transcriptional program of the inflammation (33,34).…”