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Background: Aerococcus urinae and Aerococcus sanguinicola are emerging pathogens linked with urinary tract infections. We present a case series of A. urinae and A. sanguinicola isolates characterizing the spectrum of clinical presentation, microbiological characteristics and antimicrobial sensitivities. Methods: Retrospective chart review was performed on patients who grew positive cultures for A. urinae and A. sanguinicola identified on MALDI-TOF in Saskatchewan from January to June 2023. Demographic and clinical variables, antimicrobial susceptibility and prescription were documented. Results: This cohort (n = 115) had a median age 82 years. A. urinae and A. sanguinicola infections spanned from urinary tract infection (n = 96) to urosepsis (n = 6). These infections were predominantly monomicrobial (73.9%) and were susceptible to ceftriaxone, penicillin G and vancomycin. Antimicrobials were seldom prescribed within the urinary tract infection cohort (31.2%). Conclusion: Untreated A. urinae and A. sanguinicola infections can precipitate into urosepsis. The reported antimicrobial susceptibility for these Aerococcus isolates should be utilized to provide appropriate antimicrobial coverage.
Aerococcus urinae and Aerococcus sanguinicola are bacteria that can cause urine infections. They are often overlooked and thought to be unable to cause serious blood infections, such as sepsis. We collected data on 87 cases of A. urinae and 28 cases of A. sanguinicola to show that these bacteria can cause urine and blood infections in elderly patients. We also looked at other studies and summarized that patients with serious blood infections from these bacteria often had a previous urine infection from these same bacteria. These bacteria can be resistant to a common antibiotic used to treat urine infections. It is important to test and report if these bacteria are resistant to this common antibiotic and doctors must be aware that they can cause serious blood infections if not treated with the correct antibiotics.
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We describe the case of a 57-year-old man with poorly controlled type 2 diabetes mellitus who presented with 30 days of left-sided abdominal pain. He was found to have a left adrenal abscess and underwent adrenalectomy. Intraoperative cultures grew Nocardia beijingensis, which is an uncommonly identified Nocardia species rarely affecting immunocompetent patients. We review the published literature on cases of N beijingensis among immunocompetent patients. This is the first report summarizing the diagnosis and management of N beijingensis isolated from an adrenal abscess.
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Acanthocephala is a phylum of parasitic pseudocoelomates that infect a wide range of vertebrate and invertebrate hosts and can cause zoonotic infections in humans. The zoologic literature is quite rich and diverse; however, the human-centric literature is sparse, with sporadic reports over the past 70 years. Causal agents of acanthocephaliasis in humans are reviewed as well as their biology and life cycle. This review provides the first consolidated and summarized report of human cases of acanthocephaliasis based on English language publications, including epidemiology, clinical presentation, treatment, and diagnosis and identification.
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Acantocéfalos , Helmintíase , Enteropatias Parasitárias , Parasitos , Animais , Helmintíase/diagnóstico , Interações Hospedeiro-Parasita , HumanosRESUMO
On 24 August 2020, the Centers for Disease Control and Prevention (CDC) updated its website to highlight that asymptomatic individuals, even those with exposure to a COVID-19-positive contact, do not necessarily need to be tested unless they have medical conditions associated with increased risk of severe illness from COVID-19. The CDC subsequently updated its guidance on 19 September 2020 to support testing of asymptomatic persons, including close contacts of persons with documented SARS-CoV-2 infection. In this editorial, the American Society for Microbiology Clinical and Public Health Microbiology Committee's Subcommittee on Laboratory Practices comments on testing of asymptomatic individuals relative to current medical knowledge of the virus and mitigation measures. Specific points are provided concerning such testing when undertaking contact tracing and routine surveillance. Limitations to consider when testing asymptomatic persons are covered, including the need to prioritize testing of contacts of positive COVID-19 cases. We urge the CDC to consult with primary stakeholders of COVID-19 testing when making such impactful changes in testing guidance.
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Doenças Assintomáticas , Teste para COVID-19/métodos , COVID-19/diagnóstico , Portador Sadio/diagnóstico , Indicadores de Doenças Crônicas , Busca de Comunicante/métodos , Feminino , Humanos , Masculino , SARS-CoV-2/isolamento & purificaçãoRESUMO
Infection with dengue virus (DENV) is widespread across tropical regions and can result in severe disease. Early diagnosis is important both for patient management and to differentiate infections that present with similar symptoms, such as malaria, chikungunya, and Zika. Rapid diagnostic tests that are used presently for point-of-care detection of DENV antigens lack the sensitivity of molecular diagnostics that detect viral RNA. However, no molecular diagnostic test for DENV is available for use in field settings. In this study, we developed and validated a reverse transcription-polymerase chain reaction (RT-PCR) for the detection of DENV adapted for use in field settings. Reverse transcription-polymerase chain reaction was performed directly from plasma samples without RNA extraction. The assay detected all four serotypes of DENV spiked into blood or plasma. Our RT-PCR does not cross-react with pathogens that cause symptoms that overlap with dengue infection. The test performed equally well in a conventional laboratory qPCR instrument and a small, low-cost portable instrument that can be used in a field setting. The lower limit of detection for the assay was 1 × 104 genome copy equivalents/mL in blood. Finally, we validated our test using 126 archived patient samples. The sensitivity of our RT-PCR was 76.7% (95% CI: 65.8-87.9%) on the conventional instrument, and 78.3% (95% CI: 65.8-87.9%) on the field instrument, when compared with the RealStar Dengue RT-PCR Kit 2.0. The molecular test described here is user-friendly, low-cost, and can be used in regions with limited laboratory capabilities.
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Vírus da Dengue/classificação , Dengue/diagnóstico , Testes Imediatos , Reação em Cadeia da Polimerase/métodos , RNA Viral/sangue , Alphavirus , Animais , Chlorocebus aethiops , Humanos , Imunoensaio , Plasmodium falciparum , Plasmodium vivax , Sensibilidade e Especificidade , Testes Sorológicos/métodos , Especificidade da Espécie , Células Vero , Zika virusRESUMO
Poxviruses encode many proteins with the ability to regulate cellular signaling pathways. One such protein is the vaccinia virus innate immunity modulator E3. Multiple functions have been ascribed to E3, including modulating the cellular response to double-stranded RNA, inhibiting the NF-κB and IRF3 pathways, and dampening apoptosis. Apoptosis serves as a powerful defense against damaged and unwanted cells and is an effective defense against viral infection; many viruses therefore encode proteins that prevent or delay apoptosis. Here, we present data indicating that E3 does not directly inhibit the intrinsic apoptotic pathway; instead, it suppresses apoptosis indirectly by stimulating expression of the viral F1 apoptotic inhibitor. Our data demonstrate that E3 promotes F1 expression by blocking activation of the double-stranded RNA-activated protein kinase R (PKR). F1 mRNA is present in cells infected with E3-null virus, but the protein product does not detectably accumulate, suggesting a block at the translational level. We also show that two 3' coterminal transcripts span the F1 open reading frame (ORF), a situation previously described for the vaccinia virus mRNAs encoding the J3 and J4 proteins. One of these is a conventional monocistronic transcript of the F1L gene, while the other arises by read-through transcription from the upstream F2L gene and does not give rise to appreciable levels of F1 protein.IMPORTANCE Previous studies have shown that E3-deficient vaccinia virus triggers apoptosis of infected cells. Our study demonstrates that this proapoptotic phenotype stems, at least in part, from the failure of the mutant virus to produce adequate quantities of the viral F1 protein, which acts at the mitochondria to directly block apoptosis. Our data establish a regulatory link between the vaccinia virus proteins that suppress the innate response to double-stranded RNA and those that block the intrinsic apoptotic pathway.
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Interações Hospedeiro-Patógeno , Proteínas de Ligação a RNA/genética , Vaccinia virus/genética , Proteínas Virais/genética , eIF-2 Quinase/genética , Animais , Apoptose/genética , Deleção de Genes , Regulação da Expressão Gênica , Células HEK293 , Células HeLa , Humanos , Fator Regulador 3 de Interferon/genética , Fator Regulador 3 de Interferon/metabolismo , Camundongos , NF-kappa B/genética , NF-kappa B/metabolismo , Fases de Leitura Aberta , Isoformas de Proteínas/antagonistas & inibidores , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Células RAW 264.7 , RNA de Cadeia Dupla/genética , RNA de Cadeia Dupla/metabolismo , Proteínas de Ligação a RNA/metabolismo , Transdução de Sinais , Vaccinia virus/metabolismo , Proteínas Virais/antagonistas & inibidores , Proteínas Virais/metabolismo , eIF-2 Quinase/metabolismoRESUMO
Current evidence supports a protective role for virus-neutralizing antibodies in immunity against hepatitis C virus (HCV) infection. Many cross-neutralizing monoclonal antibodies have been identified. These antibodies have been shown to provide protection or to clear infection in animal models. Previous clinical trials have shown that a gpE1/gpE2 vaccine can induce antibodies that neutralize the in vitro infectivity of all the major cell culture-derived HCV (HCVcc) genotypes around the world. However, cross-neutralization appeared to favor certain genotypes, with significant but lower neutralization against others. HCV may employ epitope masking to avoid antibody-mediated neutralization. Hypervariable region 1 (HVR1) at the amino terminus of glycoprotein E2 has been shown to restrict access to many neutralizing antibodies. Consistent with this, other groups have reported that recombinant viruses lacking HVR1 are hypersensitive to neutralization. It has been proposed that gpE1/gpE2 lacking this domain could be a better vaccine antigen to induce broadly neutralizing antibodies. In this study, we examined the immunogenicity of recombinant gpE1/gpE2 lacking HVR1 (ΔHVR1). Our results indicate that wild-type (WT) and ΔHVR1 gpE1/gpE2 antigens induced antibodies targeting many well-characterized cross-genotype-neutralizing epitopes. However, while the WT gpE1/gpE2 vaccine can induce cross-genotype protection against various genotypes of HCVcc and/or HCV-pseudotyped virus (HCVpp), antisera from ΔHVR1 gpE1/gpE2-immunized animals exhibited either reduced homologous neutralization activity compared to that of the WT or heterologous neutralization activity similar to that of the WT. These data suggest that ΔHVR1 gpE1/gpE2 is not a superior vaccine antigen. Based on previously reported chimpanzee protection data using WT gpE1/gpE2 and our current findings, we are preparing a combination vaccine including wild-type recombinant gpE1/gpE2 for clinical testing in the future.IMPORTANCE An HCV vaccine is an unmet medical need. Current evidence suggests that neutralizing antibodies play an important role in virus clearance, along with cellular immune responses. Previous clinical data showed that gpE1/gpE2 can effectively induce cross-neutralizing antibodies, although they favor certain genotypes. HCV employs HVR1 within gpE2 to evade host immune control. It has been hypothesized that the removal of this domain would improve the production of cross-neutralizing antibodies. In this study, we compared the immunogenicities of WT and ΔHVR1 gpE1/gpE2 antigens as vaccine candidates. Our results indicate that the ΔHVR1 gpE1/gpE2 antigen confers no advantages in the neutralization of HCV compared with the WT antigen. Previously, we showed that this WT antigen remains the only vaccine candidate to protect chimpanzees from chronic infection, contains multiple cross-neutralizing epitopes, and is well tolerated and immunogenic in humans. The current data support the further clinical development of this vaccine antigen component.
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Hepacivirus/imunologia , Antígenos da Hepatite C/imunologia , Hepatite C/prevenção & controle , Proteínas do Envelope Viral/imunologia , Vacinas contra Hepatite Viral/imunologia , Proteínas Virais/imunologia , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Células CHO , Cricetulus , Feminino , Cobaias , Hepatite C/imunologia , Hepatite C/virologia , Anticorpos Anti-Hepatite C/imunologia , Humanos , Camundongos , Testes de Neutralização , Vacinas Sintéticas/imunologiaRESUMO
Multiple sclerosis (MS) is an inflammatory demyelinating disease of the central nervous system (CNS). Neurosteroids are reported to exert anti-inflammatory effects in several neurological disorders. We investigated the expression and actions of the neurosteroid, dehydroepiandrosterone (DHEA), and its more stable 3ß-sulphated ester, DHEA-S, in MS and associated experimental models. CNS tissues from patients with MS and animals with experimental autoimmune encephalomyelitis (EAE) displayed reduced DHEA concentrations, accompanied by diminished expression of the DHEA-synthesizing enzyme CYP17A1 in oligodendrocytes (ODCs), in association with increased expression of inflammatory genes including interferon (IFN)-γ and interleukin (IL)-1ß. CYP17A1 was expressed variably in different human neural cell types but IFN-γ exposure selectively reduced CYP17A1 detection in ODCs. DHEA-S treatment reduced IL-1ß and -6 release from activated human myeloid cells with minimal effect on lymphocyte viability. Animals with EAE receiving DHEA-S treatment showed reduced Il1b and Ifng transcript levels in spinal cord compared to vehicle-treated animals with EAE. DHEA-S treatment also preserved myelin basic protein immunoreactivity and reduced axonal loss in animals with EAE, relative to vehicle-treated EAE animals. Neurobehavioral deficits were reduced in DHEA-S-treated EAE animals compared with vehicle-treated animals with EAE. Thus, CYP17A1 expression in ODCs and its product DHEA were downregulated in the CNS during inflammatory demyelination while DHEA-S provision suppressed neuroinflammation, demyelination, and axonal injury that was evident as improved neurobehavioral performance. These findings indicate that DHEA production is an immunoregulatory pathway within the CNS and its restoration represents a novel treatment approach for neuroinflammatory diseases.
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Sistema Nervoso Central/patologia , Citocinas/metabolismo , Esclerose Múltipla/patologia , Neurotransmissores/metabolismo , Oligodendroglia/metabolismo , Animais , Astrócitos/efeitos dos fármacos , Astrócitos/metabolismo , Estudos de Casos e Controles , Células Cultivadas , Desidroepiandrosterona/metabolismo , Desidroepiandrosterona/uso terapêutico , Modelos Animais de Doenças , Encefalomielite Autoimune Experimental/tratamento farmacológico , Encefalomielite Autoimune Experimental/metabolismo , Encefalomielite Autoimune Experimental/patologia , Feminino , Feto/citologia , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/genética , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Esteroide 17-alfa-Hidroxilase/metabolismoRESUMO
UNLABELLED: The herpes simplex virus (HSV) virion host shutoff (vhs) RNase destabilizes cellular and viral mRNAs, suppresses host protein synthesis, dampens antiviral responses, and stimulates translation of viral mRNAs. vhs mutants display a host range phenotype: translation of viral true late mRNAs is severely impaired and stress granules accumulate in HeLa cells, while translation proceeds normally in Vero cells. We found that vhs-deficient virus activates the double-stranded RNA-activated protein kinase R (PKR) much more strongly than the wild-type virus does in HeLa cells, while PKR is not activated in Vero cells, raising the possibility that PKR might play roles in stress granule induction and/or inhibiting translation in restrictive cells. We tested this possibility by evaluating the effects of inactivating PKR. Eliminating PKR in HeLa cells abolished stress granule formation but had only minor effects on viral true late protein levels. These results document an essential role for PKR in stress granule formation by a nuclear DNA virus, indicate that induction of stress granules is the consequence rather than the cause of the translational defect, and are consistent with our previous suggestion that vhs promotes translation of viral true late mRNAs by preventing mRNA overload rather than by suppressing eIF2α phosphorylation. IMPORTANCE: The herpes simplex virus vhs RNase plays multiple roles during infection, including suppressing PKR activation, inhibiting the formation of stress granules, and promoting translation of viral late mRNAs. A key question is the extent to which these activities are mechanistically connected. Our results demonstrate that PKR is essential for stress granule formation in the absence of vhs, but at best, it plays a secondary role in suppressing translation of viral mRNAs. Thus, the ability of vhs to promote translation of viral mRNAs can be largely uncoupled from PKR suppression, demonstrating that this viral RNase modulates at least two distinct aspects of RNA metabolism.
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Grânulos Citoplasmáticos/metabolismo , Herpesvirus Humano 1/metabolismo , Biossíntese de Proteínas , RNA Mensageiro/genética , RNA Viral/genética , Proteínas Virais/metabolismo , Vírion/metabolismo , eIF-2 Quinase/metabolismo , Animais , Chlorocebus aethiops , Ativação Enzimática , Fator de Iniciação 2 em Eucariotos/metabolismo , Células HEK293 , Células HeLa , Interações Hospedeiro-Patógeno , Humanos , RNA de Cadeia Dupla/genética , RNA de Cadeia Dupla/metabolismo , RNA Mensageiro/metabolismo , RNA Viral/metabolismo , Células Vero , Proteínas Virais/genética , Replicação Viral , eIF-2 Quinase/genéticaRESUMO
Apoptosis serves as a powerful defense against damaged or pathogen-infected cells. Since apoptosis is an effective defense against viral infection, many viruses including poxviruses, encode proteins to prevent or delay apoptosis. Here we show that ectromelia virus, the causative agent of mousepox encodes an anti-apoptotic protein EVM025. Here we demonstrate that expression of functional EVM025 is crucial to prevent apoptosis triggered by virus infection and staurosporine. We demonstrate that the expression of EVM025 prevents the conformational activation of the pro-apoptotic proteins Bak and Bax, allowing the maintenance of mitochondrial membrane integrity upon infection with ECTV. Additionally, EVM025 interacted with intracellular Bak. We were able to demonstrate that EVM025 ability to inhibit Bax activation is a function of its ability to inhibit the activity of an upstream BH3 only protein Bim. Collectively, our data indicates that EVM025 inhibits apoptosis by sequestering Bak and inhibiting the activity of Bak and Bax.
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Apoptose/fisiologia , Vírus da Ectromelia/metabolismo , Regulação Viral da Expressão Gênica/fisiologia , Proteínas Virais/metabolismo , Animais , Linhagem Celular , Vírus da Ectromelia/genética , Fibroblastos/metabolismo , Deleção de Genes , Humanos , Camundongos , Proteínas Virais/genética , Proteína Killer-Antagonista Homóloga a bcl-2/genética , Proteína Killer-Antagonista Homóloga a bcl-2/metabolismo , Proteína X Associada a bcl-2/genética , Proteína X Associada a bcl-2/metabolismoRESUMO
Poxviruses contain large dsDNA genomes encoding numerous open reading frames that manipulate cellular signalling pathways and interfere with the host immune response. The NF-κB signalling cascade is an important mediator of innate immunity and inflammation, and is tightly regulated by ubiquitination at several key points. A critical step in NF-κB activation is the ubiquitination and degradation of the inhibitor of kappaB (IκBα), by the cellular SCFß-TRCP ubiquitin ligase complex. We show here that upon stimulation with TNFα or IL-1ß, Orthopoxvirus-infected cells displayed an accumulation of phosphorylated IκBα, indicating that NF-κB activation was inhibited during poxvirus infection. Ectromelia virus is the causative agent of lethal mousepox, a natural disease that is fatal in mice. Previously, we identified a family of four ectromelia virus genes (EVM002, EVM005, EVM154 and EVM165) that contain N-terminal ankyrin repeats and C-terminal F-box domains that interact with the cellular SCF ubiquitin ligase complex. Since degradation of IκBα is catalyzed by the SCFß-TRCP ubiquitin ligase, we investigated the role of the ectromelia virus ankyrin/F-box protein, EVM005, in the regulation of NF-κB. Expression of Flag-EVM005 inhibited both TNFα- and IL-1ß-stimulated IκBα degradation and p65 nuclear translocation. Inhibition of the NF-κB pathway by EVM005 was dependent on the F-box domain, and interaction with the SCF complex. Additionally, ectromelia virus devoid of EVM005 was shown to inhibit NF-κB activation, despite lacking the EVM005 open reading frame. Finally, ectromelia virus devoid of EVM005 was attenuated in both A/NCR and C57BL/6 mouse models, indicating that EVM005 is required for virulence and immune regulation in vivo.
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Vírus da Ectromelia/patogenicidade , Ectromelia Infecciosa/metabolismo , NF-kappa B/metabolismo , Proteínas Virais/metabolismo , Animais , Vírus da Ectromelia/imunologia , Vírus da Ectromelia/metabolismo , Ectromelia Infecciosa/imunologia , Citometria de Fluxo , Células HeLa , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Microscopia de Fluorescência , NF-kappa B/imunologia , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteínas Virais/imunologia , Virulência/fisiologiaRESUMO
UNLABELLED: Apoptosis is a tightly regulated process that plays a crucial role in the removal of virus-infected cells, a process controlled by both pro- and antiapoptotic members of the Bcl-2 family. The proapoptotic proteins Bak and Bax are regulated by antiapoptotic Bcl-2 proteins and are also activated by a subset of proteins known as BH3-only proteins that perform dual functions by directly activating Bak and Bax or by sequestering and neutralizing antiapoptotic family members. Numerous viruses express proteins that prevent premature host cell apoptosis. Vaccinia virus encodes F1L, an antiapoptotic protein essential for survival of infected cells that bears no discernible sequence homology to mammalian cell death inhibitors. Despite the limited sequence similarities, F1L has been shown to adopt a novel dimeric Bcl-2-like fold that enables hetero-oligomeric binding to both Bak and the proapoptotic BH3-only protein Bim that ultimately prevents Bak and Bax homo-oligomerization. However, no structural data on the mode of engagement of F1L and its Bcl-2 counterparts are available. Here we solved the crystal structures of F1L in complex with two ligands, Bim and Bak. Our structures indicate that F1L can engage two BH3 ligands simultaneously via the canonical Bcl-2 ligand binding grooves. Furthermore, by structure-guided mutagenesis, we generated point mutations within the binding pocket of F1L in order to elucidate the residues responsible for both Bim and Bak binding and prevention of apoptosis. We propose that the sequestration of Bim by F1L is primarily responsible for preventing apoptosis during vaccinia virus infection. IMPORTANCE: Numerous viruses have adapted strategies to counteract apoptosis by encoding proteins responsible for sequestering proapoptotic components. Vaccinia virus, the prototypical member of the family Orthopoxviridae, encodes a protein known as F1L that functions to prevent apoptosis by interacting with Bak and the BH3-only protein Bim. Despite recent structural advances, little is known regarding the mechanics of binding between F1L and the proapoptotic Bcl-2 family members. Utilizing three-dimensional structures of F1L bound to host proapoptotic proteins, we generated variants of F1L that neutralize Bim and/or Bak. We demonstrate that during vaccinia virus infection, engagement of Bim and Bak by F1L is crucial for subversion of host cell apoptosis.
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Proteínas Reguladoras de Apoptose/química , Proteínas Reguladoras de Apoptose/metabolismo , Proteínas de Membrana/química , Proteínas de Membrana/metabolismo , Proteínas Proto-Oncogênicas/química , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Virais/química , Proteínas Virais/metabolismo , Proteína Killer-Antagonista Homóloga a bcl-2/química , Proteína Killer-Antagonista Homóloga a bcl-2/metabolismo , Animais , Proteína 11 Semelhante a Bcl-2 , Sítios de Ligação , Linhagem Celular , Cristalografia por Raios X , Análise Mutacional de DNA , Humanos , Mutagênese Sítio-Dirigida , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Mutação Puntual , Ligação Proteica , Conformação Proteica , Proteínas Virais/genéticaRESUMO
Many viruses express inhibitors of programmed cell death (apoptosis), thereby countering host defenses that would otherwise rapidly clear infected cells. To counter this, viruses such as adenoviruses and herpesviruses express recognizable homologs of the mammalian prosurvival protein Bcl-2. In contrast, the majority of poxviruses lack viral Bcl-2 (vBcl-2) homologs that are readily identified by sequence similarities. One such virus, myxoma virus, which is the causative agent of myxomatosis, expresses a virulence factor that is a potent inhibitor of apoptosis. In spite of the scant sequence similarity to Bcl-2, myxoma virus M11L adopts an almost identical 3-dimensional fold. We used M11L as bait in a sequence similarity search for other Bcl-2-like proteins and identified six putative vBcl-2 proteins from poxviruses. Some are potent inhibitors of apoptosis, in particular sheeppox virus SPPV14, which inhibited cell death induced by multiple agents. Importantly, SPPV14 compensated for the loss of antiapoptotic F1L in vaccinia virus and acts to directly counter the cell death mediators Bax and Bak. SPPV14 also engages a unique subset of the death-promoting BH3-only ligands, including Bim, Puma, Bmf, and Hrk. This suggests that SPPV14 may have been selected for specific biological roles as a virulence factor for sheeppox virus.
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Apoptose , Capripoxvirus/patogenicidade , Evasão da Resposta Imune , Proteínas Virais/metabolismo , Fatores de Virulência/metabolismo , Proteína Killer-Antagonista Homóloga a bcl-2/antagonistas & inibidores , Proteína X Associada a bcl-2/antagonistas & inibidores , Sequência de Aminoácidos , Animais , Linhagem Celular , Humanos , Camundongos , Dados de Sequência Molecular , Homologia de Sequência de AminoácidosRESUMO
Precise regulation of Major Histocompatibility class II (MHC II) genes plays important roles in initiation, propagation and termination of adaptive immune responses by controlling antigen presentation to CD4+ T cells. MHC II genes are constitutively expressed in only a few cell types and are inducibly expressed by the inflammatory response cytokine interferon gamma (IFNγ) in all nucleated cells. The regulation of MHC II is tightly controlled by a Master Regulator, the class II transactivator (CIITA), which is a general regulator of both constitutive and inducible MHC II expression. Although much is known about the transcription factors necessary for CIITA expression, less is known about the epigenetic modifications and the requisite enzymes needed to provide these transcription factors access to DNA. We show here that multiple epigenetic changes occur at the IFNγ inducible CIITA promoter within 20' of IFNγ stimulation and that these changes correlate with the opening of the promoter and the initiation of transcription. Our study links these rapidly occurring epigenetic events at the inducible CIITA promoter to decreased promoter binding of the histone methyltransferase EZH2, and shows that decreased promoter binding of EZH2 transforms this previously tightly regulated and cytokine inducible promoter into a constitutively active and dysregulated gene.
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Epigênese Genética , Histonas/metabolismo , Proteínas Nucleares/genética , Transativadores/genética , Acetilação , Imunidade Adaptativa , Proteínas de Ligação a DNA/genética , Proteína Potenciadora do Homólogo 2 de Zeste , Células HeLa , Humanos , Interferon gama/metabolismo , Interferon gama/fisiologia , Metilação , Proteínas Nucleares/química , Proteínas Nucleares/metabolismo , Complexo Repressor Polycomb 2 , Regiões Promotoras Genéticas , Transativadores/química , Transativadores/metabolismo , Fatores de Transcrição/genéticaRESUMO
BACKGROUND: Studies indicate that the 19S proteasome contributes to chromatin reorganization, independent of the role the proteasome plays in protein degradation. We have previously shown that components of the 19S proteasome are crucial for regulating inducible histone activation events in mammalian cells. The 19S ATPase Sug1 binds to histone-remodeling enzymes, and in the absence of Sug1, a subset of activating epigenetic modifications including histone H3 acetylation, H3 lysine 4 trimethylation and H3 arginine 17 dimethylation are inhibited at cytokine-inducible major histocompatibilty complex (MHC)-II and class II transactivator (CIITA) promoters, implicating Sug1 in events required to initiate mammalian transcription. RESULTS: Our previous studies indicate that H3 lysine 4 trimethylation at cytokine-inducible MHC-II and CIITA promoters is dependent on proteolytic-independent functions of 19S ATPases. In this report, we show that multiple common subunits of the mixed lineage leukemia (MLL)/complex of proteins associated with Set I (COMPASS) complexes bind to the inducible MHC-II and CIITA promoters; that overexpressing a single common MLL/COMPASS subunit significantly enhances promoter activity and MHC-II HLA-DRA expression; and that these common subunits are important for H3 lysine 4 trimethylation at MHC-II and CIITA promoters. In addition, we show that H3 lysine 27 trimethylation, which is inversely correlated with H3 lysine 4 trimethylation, is significantly elevated in the presence of diminished 19S ATPase Sug1. CONCLUSION: Taken together, these experiments suggest that the 19S proteasome plays a crucial role in the initial reorganization of events enabling the relaxation of the repressive chromatin structure surrounding inducible promoters.
RESUMO
Studies indicate that the 19S proteasome functions in the epigenetic regulation of transcription. We have shown that as in yeast, components of the 19S proteasome are crucial for regulating inducible histone acetylation events in mammalian cells. The 19S ATPase Sug1 binds to histone acetyltransferases and to acetylated histone H3 and, in the absence of Sug1, histone H3 acetylation is dramatically decreased at mammalian promoters. Research in yeast further indicates that the ortholog of Sug1, Rpt6, is a link between ubiquitination of histone H2B and H3 lysine 4 trimethylation (H3K4me3). To characterize the role that the 19S proteasome plays in regulating additional activating modifications, we examined the methylation and ubiquitination status of histones at inducible mammalian genes. We find that Sug1 is crucial for regulating histone H3K4me3 and H3R17me2 at the cytokine inducible MHC-II and CIITA promoters. In the absence of Sug1, histone H3K4me3 and H3R17me2 are dramatically decreased, but the loss of Sug1 has no significant effect on H3K36me3 or H2BK120ub. Our observation that a subunit of hCompass interacts with additional activating histone modifying enzymes, but fails to bind the CIITA promoter in the absence of Sug1, strongly implicates Sug1 in recruiting enzyme complexes responsible for initiating mammalian transcription.