Interaction of pathogens with cells of the immune system results in activation of inflammatory gene expression. This response, while vital for immune defence, is frequently deleterious to the host due to the exaggerated production of inflammatory proteins. The scope of inflammatory responses reflects the activation state of signalling proteins upstream of inflammatory genes as well as signal-induced assembly of nuclear chromatin complexes that support mRNA expression1–4. Recognition of post-translationally modified histones by nuclear proteins that initiate mRNA transcription and support mRNA elongation is a critical step in the regulation of gene expression5–10. Here we present a novel pharmacological approach that targets inflammatory gene expression by interfering with the recognition of acetylated histones by the Bromodomain and Extra Terminal domain (BET) family of proteins. We describe a synthetic compound (I-BET) that by “mimicking” acetylated histones disrupts chromatin complexes responsible for the expression of key inflammatory genes in activated macrophages and confers protection against LPS-induced endotoxic shock and bacteria-induced sepsis. Our findings suggest that synthetic compounds specifically targeting proteins that recognize post-translationally modified histones can serve as a new generation of immunomodulatory drugs.

How transcription affects genome 3D organization is not well understood. We found that during influenza A (IAV) infection, rampant transcription rapidly reorganizes host cell chromatin interactions. These changes occur at the ends of highly transcribed genes, where global inhibition of transcription termination by IAV NS1 protein causes readthrough transcription for hundreds of kilobases. In these readthrough regions, elongating RNA polymerase II disrupts chromatin interactions by inducing cohesin displacement from CTCF sites, leading to locus decompaction. Readthrough transcription into heterochromatin regions switches them from the inert (B) to the permissive (A) chromatin compartment and enables transcription factor binding. Data from non-viral transcription stimuli show that transcription similarly affects cohesin-mediated chromatin contacts within gene bodies. Conversely, inhibition of transcription elongation allows cohesin to accumulate at previously transcribed intragenic CTCF sites and to mediate chromatin looping and compaction. Our data indicate that transcription elongation by RNA polymerase II remodels genome 3D architecture.

Mosquito-borne flaviviruses, including dengue virus (DENV) and Zika virus (ZIKV), are a growing public health concern. Systems level analysis of how flaviviruses hijack cellular processes through virus-host protein-protein interactions (PPIs) provide information about their replication and pathogenic mechanisms. We used affinity purification-mass spectrometry (AP-MS) to compare flavivirus-host interactions for two viruses (DENV and ZIKV) in two hosts (human and mosquito). Conserved virus-host PPIs revealed that the flavivirus NS5 protein suppresses interferon stimulated genes by inhibiting recruitment of the transcription complex PAF1C, and that chemical modulation of SEC61 inhibits DENV and ZIKV replication in human and mosquito cells. Finally, we identified a ZIKV-specific interaction between NS4A and ANKLE2, a gene linked to hereditary microcephaly, and showed that ZIKV NS4A causes microcephaly in Drosophila in an ANKLE2-dependent manner. Thus, comparative flavivirus-host PPI mapping provides biological insights, and when coupled with in vivo models, can be used to unravel pathogenic mechanisms.

Viral infection is commonly associated with virus-driven hijacking of host proteins. Here we describe a novel mechanism by which influenza virus affects host cells through the interaction of influenza non-structural protein 1 (NS1) with the infected cell epigenome. We show that the NS1 protein of influenza A H3N2 subtype possesses a histone-like sequence (histone mimic) that is used by the virus to target the human PAF1 transcription elongation complex (hPAF1C). We demonstrate that binding of NS1 to hPAF1C depends on the NS1 histone mimic and results in suppression of hPAF1C-mediated transcriptional elongation. Furthermore, human PAF1 has a crucial role in the antiviral response. Loss of hPAF1C binding by NS1 attenuates influenza infection, whereas hPAF1C deficiency reduces antiviral gene expression and renders cells more susceptible to viruses. We propose that the histone mimic in NS1 enables the influenza virus to affect inducible gene expression selectively, thus contributing to suppression of the antiviral response.

Transcription factors of the nuclear factor (NF)-κB/Rel family translocate into the nucleus upon degradation of the IκBs. Postinduction repression of NF-κB activity depends on NF-κB–regulated resynthesis of IκBα, which dissociates NF-κB from DNA and exports it to the cytosol. We found that after activation, p65/RelA is degraded by the proteasome in the nucleus and in a DNA binding–dependent manner. If proteasome activity is blocked, NF-κB is not promptly removed from some target genes in spite of IκBα resynthesis and sustained transcription occurs. These results indicate that proteasomal degradation of p65/RelA does not merely regulate its stability and abundance, but also actively promotes transcriptional termination.

Transcription factors of the NF-kappaB family are essential regulators of the inflammatory and immune responses. The main 'switch' in NF-kappaB activation is cytoplasmic and leads to the release of NF-kappaB proteins from IkappaB molecules, specific inhibitors that prevent their nuclear accumulation. However, it is becoming increasingly apparent that in addition to this required activation step, both recruitment of NF-kappaB to target genes and NF-kappaB-induced transcriptional events after recruitment are actively controlled. Regulated recruitment of NF-kappaB to chromatin generates kinetic complexity in NF-kappaB-dependent gene induction and 'wires' NF-kappaB-regulated gene activity to simultaneously activated pathways and transcription factors.

Epigenetic gene silencing in eukaryotes is regulated in part by lysine methylation of the core histone proteins. While histone lysine methylation is known to control gene expression through the recruitment of modification-specific effector proteins, it remains unknown whether nonhistone chromatin proteins are targets for similar modification-recognition systems. Here we show that the histone H3 methyltransferase G9a contains a conserved methylation motif with marked sequence similarity to H3 itself. As with methylation of H3 lysine 9, autocatalytic G9a methylation is necessary and sufficient to mediate in vivo interaction with the epigenetic regulator heterochromatin protein 1 (HP1), and this methyl-dependent interaction can be reversed by adjacent G9a phosphorylation. NMR analysis indicates that the HP1 chromodomain recognizes methyl-G9a through a binding mode similar to that used in recognition of methyl-H3K9, demonstrating that the chromodomain functions as a generalized methyl-lysine binding module. These data reveal histone-like modification cassettes - or "histone mimics" - as a distinct class of nonhistone methylation targets and directly extend the principles of the histone code to the regulation of nonhistone proteins.

Because of its very high affinity for DNA, NF-jB is believed to make long-lasting contacts with cognate sites and to be essential for the nucleation of very stable enhanceosomes. However, the kinetic properties of NF-jB interaction with cognate sites in vivo are unknown. Here, we show that in living cells NF-jB is immobilized onto high-affinity binding sites only transiently, and that complete NF-jB turnover on active chromatin occurs in less than 30 s. Therefore, promoter-bound NF-jB is in dynamic equilibrium with nucleoplasmic dimers; promoter occupancy and transcriptional activity oscillate synchronously with nucleoplasmic NF-jB and independently of promoter occupancy by other sequence-specific transcription factors. These data indicate that changes in the nuclear concentration of NF-jB directly impact on promoter function and that promoters sample nucleoplasmic levels of NF-jB over a timescale of seconds, thus rapidly re-tuning their activity. We propose a revision of the enhanceosome concept in this dynamic framework.