Virus infection leads to localized hyperacetylation of histones H3 and H4 at the IFN-beta promoter. (1/723)

Transcriptional activation of the human interferon-beta (IFN-beta) gene by virus infection requires the assembly of a higher order nucleoprotein complex, the enhanceosome, which consists of the transcriptional activators NF-kappa B (p50/p65), ATF-2/c-jun, IRF-3 and IRF-7, architectural protein HMGI(Y), and the coactivators p300 and CBP. In this report, we show that virus infection of cells results in a dramatic hyperacetylation of histones H3 and H4 that is localized to the IFN-beta promoter. Furthermore, expressing a truncated version of IRF-3, which lacks a p300/CBP interaction domain, suppresses both histone hyperacetylation and activation of the IFN-beta gene. Thus, coactivator-mediated localized hyperacetylation of histones may play a crucial role in inducible gene expression.  (+info)

Structural and functional analysis of interferon regulatory factor 3: localization of the transactivation and autoinhibitory domains. (2/723)

The interferon regulatory factor 3 (IRF-3) gene encodes a 55-kDa protein which is expressed constitutively in all tissues. In unstimulated cells, IRF-3 is present in an inactive cytoplasmic form; following Sendai virus infection, IRF-3 is posttranslationally modified by protein phosphorylation at multiple serine and threonine residues located in the carboxy terminus. Virus-induced phosphorylation of IRF-3 leads to cytoplasmic to nuclear translocation of phosphorylated IRF-3, association with the transcriptional coactivator CBP/p300, and stimulation of DNA binding and transcriptional activities of virus-inducible genes. Using yeast and mammalian one-hybrid analysis, we now demonstrate that an extended, atypical transactivation domain is located in the C terminus of IRF-3 between amino acids (aa) 134 and 394. We also show that the C-terminal domain of IRF-3 located between aa 380 and 427 participates in the autoinhibition of IRF-3 activity via an intramolecular association with the N-terminal region between aa 98 and 240. After Sendai virus infection, an intermolecular association between IRF-3 proteins is detected, demonstrating a virus-dependent formation of IRF-3 homodimers; this interaction is also observed in the absence of virus infection with a constitutively activated form of IRF-3. Substitution of the C-terminal Ser-Thr phosphorylation sites with the phosphomimetic Asp in the region ISNSHPLSLTSDQ between amino acids 395 and 407 [IRF-3(5D)], but not the adjacent S385 and S386 residues, generates a constitutively activated DNA binding form of IRF-3. In contrast, substitution of S385 and S386 with either Ala or Asp inhibits both DNA binding and transactivation activities of the IRF-3(5D) protein. These studies thus define the transactivation domain of IRF-3, two domains that participate in the autoinhibition of IRF-3 activity, and the regulatory phosphorylation sites controlling IRF-3 dimer formation, DNA binding activity, and association with the CBP/p300 coactivator.  (+info)

Lipopolysaccharide inhibits virus-mediated induction of interferon genes by disruption of nuclear transport of interferon regulatory factors 3 and 7. (3/723)

We have studied the effects of lipopolysaccharide (LPS) on the Newcastle disease virus (NDV)-mediated induction of cytokine genes expression. Raw cells treated with LPS before or after virus infection showed down-regulation in the expression of interferon A and, to a lesser extent, interferon B genes. In contrast, induction of the interleukin (IL)-6 gene was enhanced. The effects of LPS were not a result of the suppression of virus replication, because the transcription of viral nucleocapsid gene was not affected. Consistent with these findings, LPS also suppressed the NDV-mediated induction of chloramphenicol acetyltransferase reporter gene driven by murine interferon A4 promoter in a transient transfection assay. Furthermore, LPS inhibited virus-mediated phosphorylation of interferon regulatory factor (IRF)-3 and the consequent translocation of IRF-3 from cytoplasm to nucleus. The LPS-mediated inhibition of IFNA gene expression was much weaker in infected Raw cells that constitutively overexpressed IRF-3. The nuclear translocation of IRF-7 in infected cells was also inhibited by LPS. These data suggest that LPS down-regulates the virus-mediated induction of IFNA genes by post-translationally targeting the IRF-3 and IRF-7 proteins.  (+info)

Retinoic acid resistance in NB4 APL cells is associated with lack of interferon alpha synthesis Stat1 and p48 induction. (4/723)

In the t(15;17) acute promyelocytic leukaemia (APL), all trans-retinoic (RA) treatment induces maturation leading to clinically complete but not durable remission, as RA resistance develops in the treated patients as well as in vitro. RA and interferons (IFNs) are known inhibitors of proliferation in various cells including those from APL. In this report, we show that they can act cooperatively to inhibit growth and to induce differentiation of NB4 cells but not of two RA-resistant NB4 derived cell lines, NB4-R1 and NB4-R2. However, the resistant cell lines respond to IFN. In NB4 cells, RA increases the expression of Stat1, p48 and IRF-1, three transcription factors playing a central role in the IFN response and induces the synthesis and the secretion of IFN alpha. RA-induced IFN alpha seems to play a role in inhibition of NB4 cell growth but not in their differentiation. In the resistant cells, NB4-R1 and NB4-R2, both the induction of IFN and the increase of Statl and p48 expression by RA are completely blocked. In contrast, IRF-1 mRNA and protein expressions are induced in the three cell lines. This suggests that increase of IRF-1 expression is not sufficient for IFN induction. Our results identify some defects linked to RA-resistance in APL and support the hypothesis that RA-induced Stat1 expression and IFN secretion may be one of the mechanisms mediating growth inhibition by RA.  (+info)

Functional analysis of human herpesvirus 8-encoded viral interferon regulatory factor 1 and its association with cellular interferon regulatory factors and p300. (5/723)

Human herpesvirus 8/Kaposi sarcoma-associated virus (HHV-8/KSHV) contains, in addition to genes required for viral replication, a unique set of nonstructural genes which may be part of viral mimicry and contribute to viral replication and pathogenesis in vivo. Among these, HHV-8 encodes four open reading frames (ORFs) that showed homology to the transcription factors of the interferon regulatory factor (IRF) family. The ORF K9, viral IRF 1 (vIRF-1), has been cloned, and it was shown that, when overexpressed, it down modulates the interferon-mediated transcriptional activation of the interferon-stimulated gene 15 (ISG 15) promoter, and the role of vIRF-1 in viral mimicry was implied. However, the molecular mechanism of this effect has not been clarified. Here, we extend this observation and show that vIRF-1 also downregulates the transcriptional activity of IFNA gene promoter in infected cells by interfering with the transactivating activity of cellular IRFs, including IRF-1 and IRF-3. We further show that ectopic expression of vIRF-1 in NIH 3T3 cells confers resistance to tumor necrosis factor alpha-induced apoptosis. While vIRF-1 is unable to bind DNA with the same specificity as cellular IRFs, we demonstrate by in vitro binding assay that it can associate with the family of cellular IRFs, such as IRF-1 and the interferon consensus sequence binding protein. vIRF-1 interaction domain was localized between amino acids (aa) 152 and 243. While no binding between the full-size IRF-3 and vIRF-1 could be detected by the same assay, we show that vIRF-1 also targets the carboxy-terminal region (aa 1623 to 2414) of the transcriptional coactivator p300 which could also bind IRF-3 and IRF-1. These results demonstrate that vIRF-1 can modulate the transcription of the IFNA genes by direct heterodimerization with members of the IRF family, as well as by competitive binding with cellular transcription factors to the carboxy-terminal region of p300.  (+info)

Activation of interferon regulatory factor 3 in response to DNA-damaging agents. (6/723)

Genotoxic stress triggers signal transduction pathways that mediate either the protection or apoptosis of affected cells. The interferon regulatory factors (IRFs) are involved in a wide range of host defense mechanisms against environmental stresses. Treatment with DNA-damaging agents, including doxorubicin and UV radiation, caused phosphorylation of the IRF3 transcription factor. Phosphorylation of IRF3 induced its interaction with the transcriptional co-activator cAMP-response element binding protein-binding protein. Furthermore, genotoxic stress-induced phosphorylation of IRF3 resulted in its movement from the cytoplasm to the nucleus, where it activated transcription from its binding site. These observations suggest that IRF3 plays a role in the defensive responses induced by genotoxic stress.  (+info)

Interferon regulatory factor 3 is required for viral induction of beta interferon in primary cardiac myocyte cultures. (7/723)

Viral myocarditis affects an estimated 5 to 20% of the human population. The antiviral cytokine beta interferon (IFN-beta) is critical for protection against viral myocarditis in mice. That is, nonmyocarditic reoviruses induce myocarditis in mice that lack IFN-alpha/beta, and nonmyocarditic reoviruses both induce more IFN-beta and are more sensitive to the antiviral effects of IFN-beta than myocarditic reoviruses in primary cardiac myocyte cultures. Induction of IFN-beta in certain cell types involves viral activation of the transcription factor interferon regulatory factor 3 (IRF-3). To address whether IRF-3 can induce IFN-beta in cardiac myocytes, primary cardiac myocyte cultures and control L929 cells were transfected with a plasmid constitutively expressing IRF-3. Overexpression of IRF-3 resulted in induction of IFN-beta in the absence of viral infection in both cell types. To address whether IRF-3 is required for viral induction of IFN-beta, cell cultures were transfected with a plasmid constitutively expressing a dominant negative IRF-3 protein. The dominant negative IRF-3 reduced reovirus induction of IFN-beta in control L929 cells and completely eliminated induction in primary cardiac myocyte cultures. This provides the first identification of a cardiac cellular factor required for viral induction of IFN-beta and the first report of any cell type requiring IRF-3 for this response.  (+info)

p38-dependent activation of interferon regulatory factor 3 by lipopolysaccharide. (8/723)

Interferon regulatory factor 3 (IRF3) is known to participate in the transcriptional induction of interferon (IFN) alpha and IFNbeta genes, as well as of a number of interferon-stimulated genes (ISGs), as a result of viral infection. In the present study we demonstrate the activation of IRF3 followed by ISG induction after exposure of cells to the bacterial cell wall component lipopolysaccharide. Engagement of Toll-like receptors by lipopolysaccharide triggered the nuclear translocation of IRF3, followed by its DNA binding and the subsequent induction of several interferon-regulated genes. Transcriptional activation of ISGs occurred in a protein synthesis independent manner, but was sensitive to inhibition of the stress-activated protein kinase, p38. The activation of IRF3 by viral particles or bacterial membrane components suggests that this signaling pathway might contribute to the evolutionary conserved innate immune response.  (+info)