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:KaposiM-bM-^@M-^Ys sarcoma-associated herpesvirus (KSHV) is an oncogenic gammaherpesvirus which establishes latent infection in endothelial and B cells, as well as in primary effusion lymphoma (PEL). During latency, the viral genome exists as a circular DNA minichromosome (episome) and is packaged into chromatin analogous to human chromosomes. Only a small subset of promoters, those which drive latent RNAs, are active in latent episomes. In general, nucleosome depletion (M-bM-^@M-^\open chromatinM-bM-^@M-^]) is a hallmark of eukaryotic regulatory elements such as promoters and transcriptional enhancers or insulators. We applied formaldehyde-assisted isolation of regulatory elements (FAIRE) followed by next-generation sequencing to identify regulatory elements in the KSHV genome and integrated these data with previously identified locations of histone modifications, RNA polymerase II occupancy, and CTCF binding sites. We found that (i) regions of open chromatin were not restricted to the transcriptionally defined latent loci; (ii) open chromatin was adjacent to regions harboring activating histone modifications, even at transcriptionally inactive loci; and (iii) CTCF binding sites fell within regions of open chromatin with few exceptions, including the constitutive LANA promoter and the vIL6 promoter. FAIRE-identified nucleosome depletion was similar among B and endothelial cell lineages, suggesting a common viral genome architecture in all forms of latency. Ten total samples analyzed by FAIRE-seq from latent KSHV-infected cell lines. Two replicates were performed for BC1, KSHV-BJAB, KSHV-HUVEC, and L1-TIVE cells using the Illumina HiSeq 2000 platform. For BCBL1 cells, 1 FAIRE-seq sample and 1 non-cross-linked control BCBL1 sample was analyzed using the Illumina GAIIx