An inducible program of inflammatory gene expression is a hallmark of antimicrobial defenses. Recently, cellular nucleic acid-binding protein (CNBP) was identified as a regulator of nuclear ...factor-kappaB (NF-κB)-dependent proinflammatory cytokine gene expression. Here, we generated mice lacking CNBP and found that CNBP regulates a very restricted gene signature that includes IL-12β. CNBP resides in the cytosol of macrophages and translocates to the nucleus in response to diverse microbial pathogens and pathogen-derived products.
-deficient macrophages induced canonical NF-κB/Rel signaling normally but were impaired in their ability to control the activation of c-Rel, a key driver of IL-12β gene transcription. The nuclear translocation and DNA-binding activity of c-Rel required CNBP. Lastly,
-deficient mice were more susceptible to acute toxoplasmosis associated with reduced production of IL-12β, as well as a reduced T helper type 1 (Th1) cell IFN-γ response essential to controlling parasite replication. Collectively, these findings identify CNBP as important regulator of c-Rel-dependent IL-12β gene transcription and Th1 immunity.
Abstract
An inducible program of inflammatory gene expression is a hallmark of antimicrobial defenses. This response is controlled by a collaboration involving signal-dependent activation of ...transcription factors, transcriptional co-regulators, and chromatin-modifying factors. Here we have identified a highly conserved Zinc finger DNA binding protein CNBP (also called ZNF9) upregulated in myeloid cells exposed to lipopolysaccharide. CNBP resides primarily in the cytosol and upon TLR4 engagement, CNBP translocate to the nucleus. To investigate the functional consequences of these events, we generated mice lacking CNBP and characterized the role of CNBP in controlling the inducible transcriptional program using a combination of RNA-sequencing and multiplex gene expression analysis (Nanostring). In response to an array of signals such as LPS, CNBP-deficient macrophages were impaired in their ability to induce important immune genes including IL12p40 and IL6 amongst others. CNBP-deficient cells showed normal activation of TBK1 and p65. However, the nuclear translocation of c-Rel that is critical for the inducible expression of IL-12p40 was significantly compromised in cells lacking CNBP. Furthermore, Chromatin Immunoprecipitation experiments revealed that CNBP itself bound the IL12 promoter and c-Rel binding to this promoter was compromised in CNBP-deficient macrophages. Lastly, ectopic expression of wild type CNBP but not a mutant form that could not be phosphorylated together with c-Rel led to synergistic activation of an IL12 promoter driven reporter gene. Collectively, these findings identify CNBP as a novel DNA binding protein that acts as a transcriptional regulator controlling the inflammatory program in macrophages.
Multiple clinical trials have shown that the 3-hydroxyl-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors known as statins have anti-inflammatory effects. However, the underlying molecular ...mechanism remains unclear. The proinflammatory cytokine interleukin-1β (IL-1β) is synthesized as a non-active precursor. The 31-kDa pro-IL-1β is processed into the 17-kDa active form by caspase-1-activating inflammasomes. Here, we report a novel signaling pathway induced by statins, which leads to processing of pro-IL-1β into an intermediate 28-kDa form. This statin-induced IL-1β processing is independent of caspase-1- activating inflammasomes. The 28-kDa form of IL-1β cannot activate interleukin-1 receptor-1 (IL1R1) to signal inflammatory responses. Instead, it interferes with mature IL-1β signaling through IL-1R1 and therefore may dampen inflammatory responses initiated by mature IL-1β. These results may provide new clues to explain the anti-inflammatory effects of statins.
HMG-CoA reductase inhibitors (statins) have anti-inflammatory effects, the molecular mechanisms of which remain unclear.
Statin treatment of macrophages induces secretion of a 28-kDa form of IL-1β that interferes with mature IL-1β signaling.
Statins may dampen inflammation through induction of an anti-inflammatory form of IL-1β.
These observations may provide clues toward elucidating the in vivo anti-inflammatory effects of statins.
Natural antisense transcripts (NATs) are a class of long noncoding RNAs (lncRNAs) that are complementary to other protein-coding genes. Although thousands of NATs are encoded by mammalian genomes, ...their functions in innate immunity are unknown. Here, we identify and characterize a novel NAT, AS-IL1α that is partially complementary to IL-1α. Similar to IL-1α, AS-IL1α is expressed at low levels in resting macrophages and is induced following infection with
Listeria monocytogenes
or stimulation with TLR ligands (Pam
3
CSK
4
, LPS, PolyI:C). Inducible expression of IL-1α mRNA and protein were significantly reduced in macrophages expressing shRNA that target AS-IL1α. AS-IL1α was located in the nucleus and did not alter the stability of IL-1α mRNA. Instead, AS-IL1α was required for the recruitment of RNA Polymerase II (RNAPII) to the IL-1α promoter. In summary, our studies identify AS-IL1α as important regulator of IL-1α transcription during the innate immune response.
Interferon regulatory factors (IRFs) are essential in the innate immune response and other physiological processes. Activation of these proteins in the cytoplasm is triggered by phosphorylation of ...serine and threonine residues in a C-terminal autoinhibitory region, which stimulates dimerization, transport into the nucleus, assembly with the coactivator CBP/p300 and initiation of transcription. The crystal structure of the transactivation domain of pseudophosphorylated human IRF5 strikingly reveals a dimer in which the bulk of intersubunit interactions involve a highly extended C-terminal region. The corresponding region has previously been shown to block CBP/p300 binding to unphosphorylated IRF3. Mutation of key interface residues supports the observed dimer as the physiologically activated state of IRF5 and IRF3. Thus, phosphorylation is likely to activate IRF5 and other family members by triggering conformational rearrangements that switch the C-terminal segment from an autoinihibitory to a dimerization role.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
The mitochondrial antiviral signaling protein (MAVS) orchestrates host antiviral innate immune response to RNA virus infection. However, how MAVS signaling is controlled to eradicate virus while ...preventing self-destructive inflammation remains obscure. Here, we show that protein geranylgeranylation, a posttranslational lipid modification of proteins, limits MAVS-mediated immune signaling by targeting Rho family small guanosine triphosphatase Rac1 into the mitochondria-associated endoplasmic reticulum (ER) membranes (MAMs) at the mitochondria-ER junction. Protein geranylgeranylation and subsequent palmitoylation promote Rac1 translocation into MAMs upon viral infection. MAM-localized Rac1 limits MAVS' interaction with E3 ligase Trim31 and hence inhibits MAVS ubiquitination, aggregation, and activation. Rac1 also facilitates the recruitment of caspase-8 and cFLIP
to the MAVS signalosome and the subsequent cleavage of Ripk1 that terminates MAVS signaling. Consistently, mice with myeloid deficiency of protein geranylgeranylation showed improved survival upon influenza A virus infection. Our work revealed a critical role of protein geranylgeranylation in regulating antiviral innate immune response.
Abstract
Cytosolic DNA of microbial and eukaryotic origins elicits strong inflammatory responses characterized by the production of type I interferons and IL-1β/IL-18. The maturation of proforms of ...IL-1β and IL-18 into bioactive forms is mediated by a multiprotein complex in the cytosol called inflammasomes. Recently, AIM2 (absent in melanoma 2) protein was shown to recognize cytosolic DNA and form an inflammasome complex. To understand the role of AIM2 in innate immunity, we generated AIM2 deficient mice. Using the AIM2-deficient mice, we demonstrate that AIM2 inflammasome is an essential and non-redundant mechanism mediating IL-1β and IL-18 responses to cytosolic DNA from bacterial and viral infections. In contrast, cytosolic DNA-induced activation of NF-κB and IRF-3 are normal in AIM2-/- cells indicating that AIM2 is least likely to play a role in type I interferon production elicited by cytosolic DNA. Importantly, AIM2 deficient mice infected with vaccinia virus (VV) or mouse cytomegalovirus (mCMV) demonstrated a marked reduction in serum IL-18 levels and in number of IFN-γ producing splenic NK cells, which are critical events in the early control of VV and mCMV infection. Correspondingly, AIM2 deficiency hampered the control of virus replication in the infected mice. Overall, we demonstrate that AIM2 inflammasome is an integral component of cytosolic DNA sensing in microbial infections.
Vascular disrupting agents (VDAs) such as DMXAA (5,6-dimethylxanthenone-4-acetic acid) represent a novel approach for cancer treatment. DMXAA has potent anti-tumor activity in mice and, despite ...significant pre-clinical promise, failed human clinical trials. The anti-tumor activity of DMXAA has been linked to its ability to induce type I interferons in macrophages although the molecular mechanisms involved are poorly understood. Here we identify STING as a direct receptor for DMXAA leading to TBK1 and IRF3 signaling. Remarkably, the ability to sense DMXAA was restricted to murine STING. Human STING failed to bind to or signal in response to DMXAA. Human STING also failed to signal in response to cyclic-dinucleotides, conserved bacterial second messengers known to bind and activate murine STING signaling. Collectively, these findings detail an unexpected species-specific role for STING as a receptor for an anti-cancer drug and uncover important insights that may explain the failure of DMXAA in clinical trials for human cancer.
Although Toll-like receptor 9 (TLR9) has been implicated in cytokine and type I interferon (IFN) production during malaria in humans and mice, the high AT content of thePlasmodium falciparumgenome ...prompted us to examine the possibility that malarial DNA triggered TLR9-independent pathways. Over 6000 ATTTTTAC ("AT-rich") motifs are present in the genome ofP. falciparum, which we show here potently induce type I IFNs. Parasite DNA, parasitized erythrocytes and oligonucleotides containing the AT-rich motif induce type I IFNs via a pathway that did not involve the previously described sensors TLR9, DAI, RNA polymerase-III or IFI16/p204. Rather, AT-rich DNA sensing involved an unknown receptor that coupled to the STING, TBK1 and IRF3-IRF7 signaling pathway. Mice lacking IRF3, IRF7, the kinase TBK1 or the type I IFN receptor were resistant to otherwise lethal cerebral malaria. Collectively, these observations implicate AT-rich DNA sensing via STING, TBK1 and IRF3-IRF7 inP. falciparummalaria.
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