Aire controls immunologic tolerance by inducing a battery of thymic transcripts encoding proteins characteristic of peripheral tissues. Its unusually broad effect is achieved by releasing RNA ...polymerase II paused just downstream of transcriptional start sites. We explored Aire’s collaboration with the bromodomain-containing protein, Brd4, uncovering an astonishing correspondence between those genes induced by Aire and those inhibited by a small-molecule bromodomain blocker. Aire:Brd4 binding depended on an orchestrated series of posttranslational modifications within Aire’s caspase activation and recruitment domain. This interaction attracted P-TEFb, thereby mobilizing downstream transcriptional elongation and splicing machineries. Aire:Brd4 association was critical for tolerance induction, and its disruption could account for certain point mutations that provoke human autoimmune disease. Our findings evoke the possibility of unanticipated immunologic mechanisms subtending the potent antitumor effects of bromodomain blockers.
Studies investigating the causes of autism spectrum disorder (ASD) point to genetic, as well as epigenetic, mechanisms of the disease. Identification of epigenetic processes that contribute to ASD ...development and progression is of major importance and may lead to the development of novel therapeutic strategies. Here, we identify the bromodomain and extraterminal domain-containing proteins (BETs) as epigenetic regulators of genes involved in ASD-like behaviors in mice. We found that the pharmacological suppression of BET proteins in the brain of young mice, by the novel, highly specific, brain-permeable inhibitor I-BET858 leads to selective suppression of neuronal gene expression followed by the development of an autism-like syndrome. Many of the I-BET858-affected genes have been linked to ASD in humans, thus suggesting the key role of the BET-controlled gene network in the disorder. Our studies suggest that environmental factors controlling BET proteins or their target genes may contribute to the epigenetic mechanism of ASD.
Polycomb group protein Ezh2, one of the key regulators of development in organisms from flies to mice, exerts its epigenetic function through regulation of histone methylation. Here, we report the ...existence of the cytosolic Ezh2-containing methyltransferase complex and tie the function of this complex to regulation of actin polymerization in various cell types. Genetic evidence supports the essential role of cytosolic Ezh2 in actin polymerization-dependent processes such as antigen receptor signaling in T cells and PDGF-induced dorsal circular ruffle formation in fibroblasts. Revealed function of Ezh2 points to a broader usage of lysine methylation in regulation of both nuclear and extra-nuclear signaling processes.
Activation of Toll-like receptor (TLR) signaling by microbial signatures is critical to the induction of immune responses. Such responses demand tight regulation. RP105 is a TLR homolog thought to be ...mostly B cell specific, lacking a signaling domain. We report here that RP105 expression was wide, directly mirroring that of TLR4 on antigen-presenting cells. Moreover, RP105 was a specific inhibitor of TLR4 signaling in HEK 293 cells, a function conferred by its extracellular domain. Notably, RP105 and its helper molecule, MD-1, interacted directly with the TLR4 signaling complex, inhibiting its ability to bind microbial ligand. Finally, RP105 regulated TLR4 signaling in dendritic cells as well as endotoxin responses in vivo. Thus, our results identify RP105 as a physiological negative regulator of TLR4 responses.
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.
Viral infection leads to a robust cellular response whereby the infected cell produces hundreds of molecular regulators to combat infection. Currently, non-canonical components, e.g., long noncoding ...RNAs (lncRNAs) have been added to the repertoire of immune regulators involved in the antiviral program. Interestingly, studies utilizing next-generation sequencing technologies show that a subset of the >10,000 lncRNAs in the mammalian genome contain small open reading frames (smORFs) associated with active translation, i.e., many lncRNAs are not noncoding. Here, we use genome-wide high-throughput methods to identify potential micropeptides in smORF-containing lncRNAs involved in the immune response. Using influenza as a viral infection model, we performed RNA-seq and ribosome profiling to track expression and translation of putative lncRNAs that may encode for peptides and identify tens of potential candidates. Interestingly, many of these peptides are highly conserved at the protein level, strongly suggesting biological relevance and activity. By perusing publicly available data sets, four potential peptides of interest seem common to stress induction and/or are highly conserved; potential peptides from the MMP24-AS1, ZFAS1, RP11-622K12.1, and MIR22HG genes. Interestingly, using an antibody against the potential peptide encoded by MIR22HG RNA, we show that the peptide is stably expressed in the absence of infection, and upregulated in response to infection, corroborating the prediction of the ribosome profiling results. These data show the utility of perturbation approaches in identifying potentially relevant novel molecules encoded in the genome.
The tyrosine kinase c-Src is upregulated in various human cancers irrespective of its negative regulator Csk, but the regulatory mechanisms remain unclear. Here, we show that a lipid raft-anchored ...Csk adaptor, Cbp/PAG, is directly involved in controlling the oncogenicity of c-Src. Using Csk-deficient cells that can be transformed by c-Src overexpression, we found that Cbp expression is markedly downregulated by c-Src activation and re-expression of Cbp efficiently suppresses c-Src transformation as well as tumorigenesis. Cbp-deficient cells are more susceptible to v-Src transformation than their parental cells. Upon phosphorylation, Cbp specifically binds to activated c-Src and sequesters it in lipid rafts, resulting in an efficient suppression of c-Src function independent of Csk. In some human cancer cells and tumors, Cbp is downregulated and the introduction of Cbp significantly suppresses tumorigenesis. These findings indicate a potential role for Cbp as a suppressor of c-Src-mediated tumor progression.
B cell activating factor (BAFF) signals through BAFF-R to promote mature B cell survival. Recent analyses of BAFF-induced signaling revealed direct association between augmented B cell metabolic ...fitness and activation of Akt, one of the key regulators of cell survival. The strongest and most reproducible induction of Akt occurs with significant delay (24 h) after BAFF treatment, where it precedes activation of anabolism. It was also recently shown that BAFF induces sustained Erk activation and increased turnover of the proapoptotic molecule Bim. Here we show that these BAFF-induced signaling pathways are mediated by BAFF-R and represent previously unknown arms of I kappa B kinase (IKK)1-dependent signaling. In combination with the known role of IKK1 in regulating transcription of prosurvival genes, our data underscore the central role of IKK1 in coordinating multiple BAFF-R-mediated signaling pathways controlling mature B cell homeostasis.
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