Project description:According to current models, transcription factors (TFs) activated by extracellular stimuli operate in the context of a pre-established enhancer repertoire induced and maintained by lineage-specific TFs. Here, we uncovered the existence of latent enhancers, defined as regions of the genome that in terminally differentiated cells are poorly accessible and lack the histone marks characteristic of enhancers, but readily acquire these features in response to extracellular cues. Stimulation of resting macrophages caused simultaneous binding of stimulus-activated TFs and lineage-determining TFs to these regions, enabling deposition of enhancer-specific features. Once unveiled, these enhancers did not return to a latent state even when stimulation ceased; instead, they persisted and mediated a faster and stronger response upon restimulation. We suggest that stimulus-specific expansion of the available cis-regulatory repertoire provides an epigenomic memory of the exposure to environmental agents. Formaldehyde-Assisted Isolation Of Regulatory Elements (FAIRE) followed by multiparallel sequencing was performed in untreated murine bone marrow-derived macrophages.
Project description:According to current models, transcription factors (TFs) activated by extracellular stimuli operate in the context of a pre-established enhancer repertoire induced and maintained by lineage-specific TFs. Here, we uncovered the existence of latent enhancers, defined as regions of the genome that in terminally differentiated cells are poorly accessible and lack the histone marks characteristic of enhancers, but readily acquire these features in response to extracellular cues. Stimulation of resting macrophages caused simultaneous binding of stimulus-activated TFs and lineage-determining TFs to these regions, enabling deposition of enhancer-specific features. Once unveiled, these enhancers did not return to a latent state even when stimulation ceased; instead, they persisted and mediated a faster and stronger response upon restimulation. We suggest that stimulus-specific expansion of the available cis-regulatory repertoire provides an epigenomic memory of the exposure to environmental agents. Poly(A) fraction of the total mRNA of resting and stimulated murine bone marrow derived macrophages was extracted and subjected to by multiparallel sequencing. Experiments carried out in untreated cells as well as in cells treated for 4hrs (IFNg, IL4, TNFa, TGFb1, IL1b, MALP2, CpG)
Project description:According to current models, transcription factors (TFs) activated by extracellular stimuli operate in the context of a pre-established enhancer repertoire induced and maintained by lineage-specific TFs. Here, we uncovered the existence of latent enhancers, defined as regions of the genome that in terminally differentiated cells are poorly accessible and lack the histone marks characteristic of enhancers, but readily acquire these features in response to extracellular cues. Stimulation of resting macrophages caused simultaneous binding of stimulus-activated TFs and lineage-determining TFs to these regions, enabling deposition of enhancer-specific features. Once unveiled, these enhancers did not return to a latent state even when stimulation ceased; instead, they persisted and mediated a faster and stronger response upon restimulation. We suggest that stimulus-specific expansion of the available cis-regulatory repertoire provides an epigenomic memory of the exposure to environmental agents. Chromatin immunoprecipitations of H3 lysine 4 mono-methylated, H3 lysine 27 acetylated, H3 lysine 4 tri-methylated, the transcription factor PU.1 and total RNA polymerase II followed by multiparallel sequencing performed in murine bone marrow-derived macrophages (BMDMs). Experiments were carried out in untreated cells as well as in cells treated for 4hrs (lipopolysaccharide (LPS), IFNg, IL4, TNFa, TGFb1, IL1b, MALP2, CpG) and 24hrs (LPS).
Project description:According to current models, transcription factors (TFs) activated by extracellular stimuli operate in the context of a pre-established enhancer repertoire induced and maintained by lineage-specific TFs. Here, we uncovered the existence of latent enhancers, defined as regions of the genome that in terminally differentiated cells are poorly accessible and lack the histone marks characteristic of enhancers, but readily acquire these features in response to extracellular cues. Stimulation of resting macrophages caused simultaneous binding of stimulus-activated TFs and lineage-determining TFs to these regions, enabling deposition of enhancer-specific features. Once unveiled, these enhancers did not return to a latent state even when stimulation ceased; instead, they persisted and mediated a faster and stronger response upon restimulation. We suggest that stimulus-specific expansion of the available cis-regulatory repertoire provides an epigenomic memory of the exposure to environmental agents.
Project description:According to current models, transcription factors (TFs) activated by extracellular stimuli operate in the context of a pre-established enhancer repertoire induced and maintained by lineage-specific TFs. Here, we uncovered the existence of latent enhancers, defined as regions of the genome that in terminally differentiated cells are poorly accessible and lack the histone marks characteristic of enhancers, but readily acquire these features in response to extracellular cues. Stimulation of resting macrophages caused simultaneous binding of stimulus-activated TFs and lineage-determining TFs to these regions, enabling deposition of enhancer-specific features. Once unveiled, these enhancers did not return to a latent state even when stimulation ceased; instead, they persisted and mediated a faster and stronger response upon restimulation. We suggest that stimulus-specific expansion of the available cis-regulatory repertoire provides an epigenomic memory of the exposure to environmental agents.
Project description:According to current models, transcription factors (TFs) activated by extracellular stimuli operate in the context of a pre-established enhancer repertoire induced and maintained by lineage-specific TFs. Here, we uncovered the existence of latent enhancers, defined as regions of the genome that in terminally differentiated cells are poorly accessible and lack the histone marks characteristic of enhancers, but readily acquire these features in response to extracellular cues. Stimulation of resting macrophages caused simultaneous binding of stimulus-activated TFs and lineage-determining TFs to these regions, enabling deposition of enhancer-specific features. Once unveiled, these enhancers did not return to a latent state even when stimulation ceased; instead, they persisted and mediated a faster and stronger response upon restimulation. We suggest that stimulus-specific expansion of the available cis-regulatory repertoire provides an epigenomic memory of the exposure to environmental agents.
Project description:The integrated activity of cis-regulatory elements fine-tunes transcriptional programs of mammalian cells by recruiting cell type–specific as well as ubiquitous transcription factors (TFs). Despite their key role in modulating transcription, enhancers are still poorly characterized at the molecular level, and their limited DNA sequence conservation in evolution and variable distance from target genes make their unbiased identification challenging. The coexistence of high mono-methylation and low tri-methylation levels of lysine 4 of histone H3 is considered a signature of enhancers, but a comprehensive view of histone modifications associated to enhancers is still lacking. By combining chromatin immunoprecipitation (ChIP) with mass spectrometry, we investigated cis-regulatory regions in macrophages to comprehensively identify histone marks specifically associated with enhancers, and to profile their dynamics after transcriptional activation elicited by an inflammatory stimulation. The intersection of the proteomics data with ChIP-seq and RNA-seq analyses revealed the existence of novel subpopulations of enhancers, marked by specific histone modification signatures: specifically, H3K36me2/K4me1 marks transcribed enhancers, while H3K36me3/K4me1 and H3K79me2/K4me1 combinations mark distinct classes of intronic enhancers. Thus, our MS analysis of functionally distinct genomic regions revealed the combinatorial code of histone modifications, highlighting the potential of proteomics in addressing fundamental questions in epigenetics.