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1.
J Med Virol ; 2020 Feb 13.
Artigo em Inglês | MEDLINE | ID: mdl-32056235

RESUMO

The outbreak of a novel coronavirus (SARS-CoV-2) since Dec 2019 in Wuhan, the major transportation hub in central China, became an emergency of major international concern. While several etiological studies have begun to reveal the specific biological features of this virus, the epidemic characteristics need to be elucidated. Notably, a long incubation time was reported to be associated with SARS-CoV-2 infection, leading to adjustments in screening and control policies. To avoid the risk of virus spread, all potentially exposed subjects are required to be isolated for 14 days, which is the longest predicted incubation time. However, based on our analysis of a larger dataset available so far, we find there is no observable difference between the incubation time for SARS-CoV-2, SARS and MERS, highlighting the need for larger and well annotated datasets. This article is protected by copyright. All rights reserved.

2.
Nat Commun ; 11(1): 183, 2020 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-31924799

RESUMO

The basolateral amygdala (BLA) and ventral hippocampal CA1 (vCA1) are cellularly and functionally diverse along their anterior-posterior and superficial-deep axes. Here, we find that anterior BLA (aBLA) and posterior BLA (pBLA) innervate deep-layer calbindin1-negative (Calb1-) and superficial-layer calbindin1-positive neurons (Calb1+) in vCA1, respectively. Photostimulation of pBLA-vCA1 inputs has an anxiolytic effect in mice, promoting approach behaviours during conflict exploratory tasks. By contrast, stimulating aBLA-vCA1 inputs induces anxiety-like behaviour resulting in fewer approaches. During conflict stages of the elevated plus maze task vCA1Calb1+ neurons are preferentially activated at the open-to-closed arm transition, and photostimulation of vCA1Calb1+ neurons at decision-making zones promotes approach with fewer retreats. In the APP/PS1 mouse model of Alzheimer's disease, which shows anxiety-like behaviour, photostimulating the pBLA-vCA1Calb1+ circuit ameliorates the anxiety in a Calb1-dependent manner. These findings suggest the pBLA-vCA1Calb1+ circuit from heterogeneous BLA-vCA1 connections drives approach behaviour to reduce anxiety-like behaviour.

3.
J Virol ; 2020 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-31969440

RESUMO

The features of HSV-1 strain 129 (H129), including natural neurotropism and anterograde transneuronal trafficking, make it a potential tool for anterograde neural circuitry tracing. Recently anterograde poly-synaptic and mono-synaptic tracers were developed from H129, and have been applied for identification of novel connections and functions of different neural circuitries. However, how H129 viral particles are transported in neurons, especially those of the central nervous system remains unclear. In this study, we constructed recombinant H129 variants with mCherry-labeled capsids and/or GFP-labeled envelopes and infected the cortical neurons to study axonal transport of H129 viral particles. We found that different types of viral particles were unevenly distributed in the nucleus, cytoplasm of the cell body and axon. Most H129 progeny particles were unenveloped capsids and transported as capsids rather than virions in the axon. Notably, capsids acquired envelopes at axonal varicosities and terminals where are the sites forming synapse connected with other neurons. Moreover, viral capsids moved more frequently in the anterograde direction in axons, with an average velocity of 0.62 ± 0.18 µm/s and maximal velocity of 1.80 ± 0.15 µm/s. We also provided evidence that axonal transport of capsids requires the kinesin-1 molecular motor. These findings support that H129-derived tracers mapping neural circuit anterogradely and possibly transsynaptically. These data will guide future modifications and improvements of the H129-based anterograde viral tracers.IMPORTANCE Anterograde transneuronal tracers derived from H129 virus are important tools for mapping neural circuit anatomic and functional connections. It is therefore critical to elucidate the transport pattern of H129 virus within neuron and between neurons. We constructed recombinant H129 variants with genetically encoded fluorescence labeled capsid protein and/or glycoprotein to visualize viral particle movement in neurons. Both electron microscopy and light microscopy data show that H129 capsids and envelopes move separately, and notably capsids are enveloped at axonal varicosity and terminal which are the sites forming synapse to connect with other neurons. Super-resolution microscopy-based colocalization analysis and inhibition of H129 particle movement by inhibitors of molecular motors support that kinesin-1 contributes to the anterograde transport of capsids. These results shed light into mechanisms for anterograde transport of H129-derived tracers' transport in axon, and transmission between neurons via synapses, explaining the anterograde labeling of neural circuits by H129-derived tracers.

4.
Pain ; 161(2): 416-428, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31651582

RESUMO

Chronic pain and anxiety symptoms are frequently encountered clinically, but the neural circuit mechanisms underlying the comorbid anxiety symptoms in pain (CASP) in context of chronic pain remain unclear. Using viral neuronal tracing in mice, we identified a previously unknown pathway whereby glutamatergic neurons from layer 5 of the hindlimb primary somatosensory cortex (S1) (Glu), a well-known brain region involved in pain processing, project to GABAergic neurons in the caudal dorsolateral striatum (GABA). In a persistent inflammatory pain model induced by complete Freund's adjuvant injection, enhanced excitation of the Glu→GABA pathway was found in mice exhibiting CASP. Reversing this pathway using chemogenetic or optogenetic approaches alleviated CASP. In addition, the optical activation of Glu terminals in the cDLS produced anxiety-like behaviors in naive mice. Overall, the current study demonstrates the putative importance of a novel Glu→GABA pathway in controlling at least some aspects of CASP.

5.
Cell Rep ; 29(12): 3847-3858.e5, 2019 Dec 17.
Artigo em Inglês | MEDLINE | ID: mdl-31851918

RESUMO

While comorbid pain in depression (CP) occurs at a high rate worldwide, the neural connections underlying the core symptoms of CP have yet to be elucidated. Here, we define a pathway whereby GABAergic neurons from the central nucleus of the amygdala (GABACeA) project to glutamatergic neurons in the parafascicular nucleus (GluPF). These GluPF neurons relay directly to neurons in the second somatosensory cortex (S2), a well-known area involved in pain signal processing. Enhanced inhibition of the GABACeA→GluPF→S2 pathway is found in mice exhibiting CP symptoms. Reversing this pathway using chemogenetic or optogenetic approaches alleviates CP symptoms. Together, the current study demonstrates the putative importance of the GABACeA→GluPF→S2 pathway in controlling at least some aspects of CP.

6.
J Biol Chem ; 294(49): 18742-18755, 2019 Dec 06.
Artigo em Inglês | MEDLINE | ID: mdl-31666336

RESUMO

The centrosome is a cytoplasmic nonenveloped organelle functioning as one of the microtubule-organizing centers and composing a centriole center surrounded by pericentriolar material (PCM) granules. PCM consists of many centrosomal proteins, including PCM1 and centrosomal protein 131 (CEP131), and helps maintain centrosome stability. Zika virus (ZIKV) is a flavivirus of the family Flaviviridae whose RNA and viral particles are replicated in the cytoplasm. However, how ZIKV interacts with host cell components during its productive infection stage is incompletely understood. Here, using several primate cell lines, we report that ZIKV infection disrupts and disperses the PCM granules. We demonstrate that PCM1- and CEP131-containing granules are dispersed in ZIKV-infected cells, whereas the centrioles remain intact. We found that ZIKV does not significantly alter cellular skeletal proteins, and, hence, these proteins may not be involved in the interaction between ZIKV and centrosomal proteins. Moreover, ZIKV infection decreased PCM1 and CEP131 protein, but not mRNA, levels. We further found that the protease inhibitor MG132 prevents the decrease in PCM1 and CEP131 levels and centriolar satellite dispersion. Therefore, we hypothesized that ZIKV infection induces proteasomal PCM1 and CEP131 degradation and thereby disrupts the PCM granules. Supporting this hypothesis, we show that ZIKV infection increases levels of mind bomb 1 (MIB1), previously demonstrated to be an E3 ubiquitin ligase for PCM1 and CEP131 and that ZIKV fails to degrade or disperse PCM in MIB1-ko cells. Our results imply that ZIKV infection activates MIB1-mediated ubiquitination that degrades PCM1 and CEP131, leading to PCM granule dispersion.

8.
Nat Neurosci ; 22(10): 1649-1658, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31451801

RESUMO

Comorbid depressive symptoms (CDS) in chronic pain are a common health problem, but the neural circuit mechanisms underlying these symptoms remain unclear. Here we identify a novel pathway involving 5-hydroxytryptamine (5-HT) projections from the dorsal raphe nucleus (5-HTDRN) to somatostatin (SOM)-expressing and non-SOM interneurons in the central nucleus of the amygdala (CeA). The SOMCeA neurons project directly to the lateral habenula, an area known involved in depression. Inhibition of the 5-HTDRN→SOMCeA pathway produced depression-like behavior in a male mouse model of chronic pain. Activation of this pathway using pharmacological or optogenetic approaches reduced depression-like behavior in these mice. Human functional magnetic resonance imaging data showed that compared to healthy controls, functional connectivity between the CeA-containing centromedial amygdala and the DRN was reduced in patients with CDS but not in patients in chronic pain without depression. These findings indicate that a novel 5-HTDRN→SOMCeA→lateral habenula pathway may mediate at least some aspects of CDS.


Assuntos
Dor Crônica/patologia , Depressão/patologia , Vias Neurais/patologia , Tonsila do Cerebelo/diagnóstico por imagem , Tonsila do Cerebelo/patologia , Animais , Comportamento Animal , Dor Crônica/complicações , Dor Crônica/diagnóstico por imagem , Depressão/complicações , Depressão/diagnóstico por imagem , Núcleo Dorsal da Rafe/diagnóstico por imagem , Núcleo Dorsal da Rafe/patologia , Feminino , Habenula/diagnóstico por imagem , Habenula/patologia , Humanos , Imagem por Ressonância Magnética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Vias Neurais/diagnóstico por imagem , Neuralgia/diagnóstico por imagem , Neuralgia/patologia , Optogenética , Serotonina/metabolismo , Somatostatina/metabolismo
9.
PLoS Biol ; 17(8): e3000417, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31469831

RESUMO

Threatening sounds can elicit a series of defensive behavioral reactions in animals for survival, but the underlying neural substrates are not fully understood. Here, we demonstrate a previously unexplored neural pathway in mice that projects directly from the auditory cortex (ACx) to the lateral periaqueductal gray (lPAG) and controls noise-evoked defensive behaviors. Electrophysiological recordings showed that the lPAG could be excited by a loud noise that induced an escape-like behavior. Trans-synaptic viral tracing showed that a great number of glutamatergic neurons, rather than GABAergic neurons, in the lPAG were directly innervated by those in layer V of the ACx. Activation of this pathway by optogenetic manipulations produced a behavior in mice that mimicked the noise-evoked escape, whereas inhibition of the pathway reduced this behavior. Therefore, our newly identified descending pathway is a novel neural substrate for noise-evoked escape and is involved in controlling the threat-related behavior.

10.
PLoS Pathog ; 15(5): e1007691, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31107917

RESUMO

Cyclic GMP-AMP synthase (cGAS) senses viral DNA in the cytosol and then catalyzes synthesis of the second messenger cGAMP, which activates the ER-localized adaptor protein Mediator of IRF3 Activator (MITA) to initiate innate antiviral response. Human cytomegalovirus (HCMV) proteins can antagonize host immune responses to promote latent infection. Here, we identified HCMV UL42 as a negative regulator of cGAS/MITA-dependent antiviral response. UL42-deficiency enhances HCMV-induced production of type I interferons (IFNs) and downstream antiviral genes. Consistently, wild-type HCMV replicates more efficiently than UL42-deficient HCMV. UL42 interacts with both cGAS and MITA. UL42 inhibits DNA binding, oligomerization and enzymatic activity of cGAS. UL42 also impairs translocation of MITA from the ER to perinuclear punctate structures, which is required for MITA activation, by facilitating p62/LC3B-mediated degradation of translocon-associated protein ß (TRAPß). These results suggest that UL42 can antagonize innate immune response to HCMV by targeting the core components of viral DNA-triggered signaling pathways.


Assuntos
Antivirais/farmacologia , Infecções por Citomegalovirus/imunologia , Citomegalovirus/imunologia , Imunidade Inata/imunologia , Proteínas de Membrana/metabolismo , Nucleotidiltransferases/metabolismo , Proteínas Virais/farmacologia , Citomegalovirus/efeitos dos fármacos , Infecções por Citomegalovirus/tratamento farmacológico , Infecções por Citomegalovirus/metabolismo , Infecções por Citomegalovirus/virologia , DNA Viral/genética , DNA Viral/metabolismo , Células HEK293 , Humanos , Proteínas de Membrana/genética , Nucleotidiltransferases/genética , Transdução de Sinais
11.
J Neurovirol ; 25(4): 525-539, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31144288

RESUMO

Herpes simplex virus 1 (HSV-1) is a predominant cause of herpes simplex encephalitis (HSE), leading to a high mortality rate and severe neurological sequelae worldwide. HSE is typically accompanied by the blood-brain barrier (BBB) disruption, but the underlying mechanisms are unclear. To explore the disruption mechanisms of the BBB, quantitative analysis of the cellular proteome was carried out to investigate the proteomic changes that occur after infection. In this study, bEnd.3 cells were infected with HSV-1, followed by liquid chromatography-tandem mass spectrometry. A total of 6761 proteins were identified in three independent mass spectrometry analyses. Compared to the uninfected cells, 386 and 293 differentially expressed proteins were markedly upregulated or downregulated, respectively. Bioinformatic analysis showed that the activator protein-1 factor, including Fos, Jun, and ATF family proteins and cell adhesion molecules were significantly changed. Further validation of the changes observed for these proteins was carried out by western blotting and quantitative real-time PCR. Transendothelial electrical resistance (TEER) studies were performed to explore the effects of ATF3, Fra1, or JunB overexpression on the function of bEnd.3 cells. Characterization of the differential expression of these proteins in bEnd.3 cells will facilitate further exploration of BBB disruption upon HSV-1 infection.

12.
Virol Sin ; 34(3): 270-277, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30989428

RESUMO

As a universal pathogen leading to neonatal defects and transplant failure, human cytomegalovirus (HCMV) has strict species specificity and this has prevented the development of a suitable animal model for the pathogenesis study. The mechanism of cross-species barrier remains elusive and there are so far no non-human cell culture models that support HCMV replication. The Chinese tree shrew (Tupaia belangeri chinensis) is a small laboratory animal and evolutionary closely related with primates. We investigated the susceptibility of primary tree shrew dermis fibroblasts (TSDF) to HCMV infection. Infection with a GFP-expressing HCMV virus resulted in green fluorescence in infected cells with the expression of IE1, UL44 and pp28. The titers of cell-free viruses reached 103 PFU/mL at 96 hpi, compared to titers of 104 PFU/mL observed in primary human foreskin fibroblasts. Our results suggested that TSDF was semi-permissive for HCMV infection. The TSDF model could be further used to investigate key factors influencing cross-species multiplication of HCMV.


Assuntos
Citomegalovirus/fisiologia , Derme/virologia , Fibroblastos/virologia , Musaranhos , Animais , Células Cultivadas , Cromossomos Artificiais Bacterianos , Derme/citologia , Modelos Animais de Doenças , Fluorescência , Prepúcio do Pênis/citologia , Prepúcio do Pênis/virologia , Proteínas de Fluorescência Verde , Humanos , Masculino , Especificidade da Espécie , Replicação Viral
13.
PLoS Pathog ; 15(4): e1007680, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30943264

RESUMO

Mediator of IRF3 activation (MITA, also known as STING and ERIS) is an essential adaptor protein for cytoplasmic DNA-triggered signaling and involved in innate immune responses, autoimmunity and tumorigenesis. The activity of MITA is critically regulated by ubiquitination and deubiquitination. Here, we report that USP49 interacts with and deubiquitinates MITA after HSV-1 infection, thereby turning down cellular antiviral responses. Knockdown or knockout of USP49 potentiated HSV-1-, cytoplasmic DNA- or cGAMP-induced production of type I interferons (IFNs) and proinflammatory cytokines and impairs HSV-1 replication. Consistently, Usp49-/- mice exhibit resistance to lethal HSV-1 infection and attenuated HSV-1 replication compared to Usp49+/+ mice. Mechanistically, USP49 removes K63-linked ubiquitin chains from MITA after HSV-1 infection which inhibits the aggregation of MITA and the subsequent recruitment of TBK1 to the signaling complex. These findings suggest a critical role of USP49 in terminating innate antiviral responses and provide insights into the complex regulatory mechanisms of MITA activation.


Assuntos
Herpes Simples/prevenção & controle , Imunidade Inata/imunologia , Lisina/metabolismo , Proteínas de Membrana/metabolismo , Ubiquitina Tiolesterase/metabolismo , Animais , Antivirais , Células HEK293 , Herpes Simples/imunologia , Herpes Simples/virologia , Herpesvirus Humano 1 , Humanos , Lisina/química , Proteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transdução de Sinais , Células THP-1 , Ubiquitina Tiolesterase/genética , Ubiquitinação , Replicação Viral
14.
J Virol ; 93(11)2019 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-30867312

RESUMO

Innate immunity is the first line of host defense against viral invasion. The induction of type I interferons (IFNs) and inflammatory cytokines is essential to host antiviral immune responses, which are also key targets of viral immune evasion. Human cytomegalovirus (HCMV) can establish long-term latent infections, in which immune evasion is a pivotal step. In this study, we identified HCMV protein UL44, a DNA polymerase processivity factor, as an inhibitor of the interferon regulatory factor 3 (IRF3)- and NF-κB-dependent antiviral response. Ectopic expression of UL44 inhibited HCMV-triggered induction of downstream effector genes and enhanced viral replication. Conversely, knockdown of UL44 potentiated HCMV-triggered induction of downstream antiviral genes. UL44 interacted with IRF3 and p65, and it inhibited the binding of IRF3 and NF-κB to the promoters of their downstream antiviral genes. These findings reveal an important mechanism of immune evasion by HCMV at the transcriptional level.IMPORTANCE Induction of type I IFNs and inflammatory cytokines plays pivotal roles in host antiviral innate immune responses. Viruses have evolved various mechanisms to interfere with these processes. HCMV causes severe ailments in immunodeficient populations and is a major cause of birth defects. It has been shown that HCMV antagonizes host innate immune defenses, which is important for establishing immune evasion and latent infection. In this study, we identified the HCMV DNA polymerase subunit UL44 as a suppressor of antiviral innate immune responses. Overexpression of UL44 impaired HCMV-triggered induction of type I IFNs and other antiviral genes and thus potentiated viral replication, whereas UL44 deficiency showed opposite effects. Mechanistic studies indicated that UL44 acts by inhibiting the binding of IRF3 and NF-κB to the promoters of downstream antiviral genes. These findings defined an important mechanism of HCMV immune evasion at the transcriptional level, which may provide a therapeutic target for the treatment of HCMV infection.

15.
J Immunol ; 202(8): 2397-2406, 2019 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-30814308

RESUMO

Mediator of IRF3 activation ([MITA] also known as STING) is a direct sensor of cyclic dinucleotide and critically mediates cytoplasmic DNA--triggered innate immune signaling. The activity of MITA is extensively regulated by ubiquitination and deubiquitination. In this study, we report that USP20 interacts with and removes K48-linked ubiquitin chains from MITA after HSV-1 infection, thereby stabilizing MITA and promoting cellular antiviral responses. Deletion of USP20 accelerates HSV-1-induced degradation of MITA and impairs phosphorylation of IRF3 and IκBα as well as subsequent induction of type I IFNs and proinflammatory cytokines after HSV-1 infection or cytoplasmic DNA challenge. Consistently, Usp20 -/- mice produce decreased type I IFNs and proinflammatory cytokines, exhibit increased susceptibility to lethal HSV-1 infection, and aggravated HSV-1 replication compared with Usp20 +/+ mice. In addition, complement of MITA into Usp20 -/- cells fully restores HSV-1-triggered signaling and inhibits HSV-1 infection. These findings suggest a crucial role of USP20 in maintaining the stability of MITA and promoting innate antiviral signaling.


Assuntos
Endopeptidases/imunologia , Herpes Simples/imunologia , Herpesvirus Humano 1/imunologia , Proteínas de Membrana/imunologia , Proteólise , Ubiquitinação/imunologia , Animais , Endopeptidases/genética , Herpes Simples/genética , Imunidade Inata , Interferon Tipo I/genética , Interferon Tipo I/imunologia , Proteínas de Membrana/genética , Camundongos , Camundongos Knockout , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Ubiquitinação/genética
16.
Proc Natl Acad Sci U S A ; 116(9): 3799-3804, 2019 02 26.
Artigo em Inglês | MEDLINE | ID: mdl-30808765

RESUMO

Obsessive-compulsive disorder (OCD) affects ∼1 to 3% of the world's population. However, the neural mechanisms underlying the excessive checking symptoms in OCD are not fully understood. Using viral neuronal tracing in mice, we found that glutamatergic neurons from the basolateral amygdala (BLAGlu) project onto both medial prefrontal cortex glutamate (mPFCGlu) and GABA (mPFCGABA) neurons that locally innervate mPFCGlu neurons. Next, we developed an OCD checking mouse model with quinpirole-induced repetitive checking behaviors. This model demonstrated decreased glutamatergic mPFC microcircuit activity regulated by enhanced BLAGlu inputs. Optical or chemogenetic manipulations of this maladaptive circuitry restored the behavioral response. These findings were verified in a mouse functional magnetic resonance imaging (fMRI) study, in which the BLA-mPFC functional connectivity was increased in OCD mice. Together, these findings define a unique BLAGlu→mPFCGABA→Glu circuit that controls the checking symptoms of OCD.


Assuntos
Tonsila do Cerebelo/metabolismo , Complexo Nuclear Basolateral da Amígdala/metabolismo , Neurônios/metabolismo , Transtorno Obsessivo-Compulsivo/metabolismo , Tonsila do Cerebelo/diagnóstico por imagem , Tonsila do Cerebelo/fisiopatologia , Animais , Complexo Nuclear Basolateral da Amígdala/diagnóstico por imagem , Complexo Nuclear Basolateral da Amígdala/fisiopatologia , Modelos Animais de Doenças , Ácido Glutâmico/metabolismo , Humanos , Imagem por Ressonância Magnética , Camundongos , Vias Neurais/metabolismo , Vias Neurais/fisiopatologia , Neurônios/patologia , Transtorno Obsessivo-Compulsivo/diagnóstico por imagem , Transtorno Obsessivo-Compulsivo/fisiopatologia , Córtex Pré-Frontal/diagnóstico por imagem , Córtex Pré-Frontal/metabolismo , Córtex Pré-Frontal/fisiopatologia
17.
J Virol ; 92(23)2018 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-30258013

RESUMO

To countermeasure the host cellular intrinsic defense, cytomegalovirus (CMV) and herpes simplex viruses (HSV) have evolved the ability to disperse nuclear domain 10 (ND10, aka PML body). However, mechanisms underlying their action on ND10 differ. HSV infection produces ICP0, which degrades the ND10-forming protein PML. Human CMV (HCMV) infection expresses IE1 that deSUMOylates PML to result in dispersion of ND10. It has been demonstrated that HSV ICP0 degraded only the SUMOylated PML, so we hypothesized that HCMV IE1 can protect PML from degradation by ICP0. HCMV IE1-expressing cell lines (U-251 MG-IE1 and HELF-IE1) were used for infection of HSV-1 or transfection of ICP0-expressing plasmid. Multilabeling by immunocytochemistry assay and protein examination by Western blot assay were performed to determine the resultant fate of PML caused by ICP0 in the presence or absence of HCMV IE1. Here, we report that deSUMOylation of human PML (hPML) by HCMV IE1 was incomplete, as mono-SUMOylated PML remained in the IE1-expressing cells, which is consistent with the report by E. M. Schilling, M. Scherer, N. Reuter, J. Schweininger, et al. (J Virol 91:e02049-16, 2017, https://doi.org/10.1128/JVI.02049-16). As expected, we found that IE1 protected PML from degradation by ICP0 or HSV-1 infection. An in vitro study found that IE1 with mutation of L174P failed to deSUMOylate PML and did not protect PML from degradation by ICP0; hence, we conclude that the deSUMOylation of PML is important for IE1 to protect PML from degradation by ICP0. In addition, we revealed that murine CMV failed to deSUMOylate and to protect the HSV-mediated degradation of hPML, and that HCMV failed to deSUMOylate and protect the HSV-mediated degradation of mouse PML. However, IE1-expressing cells did not enhance wild-type HSV-1 replication but significantly increased ICP0-defective HSV-1 replication at a low multiplicity of infection. Therefore, our results uncovered a host-virus functional interaction at the posttranslational level.IMPORTANCE Our finding that HCMV IE1 protected hPML from degradation by HSV ICP0 is important, because the PML body (aka ND10) is believed to be the first line of host intrinsic defense against herpesviral infection. How the infected viruses overcome the nuclear defensive structure (PML body) has not been fully understood. Herpesviral proteins, ICP0 of HSV and IE1 of CMV, have been identified to interact with PML. Here, we report that HCMV IE1 incompletely deSUMOylated PML, resulting in the mono-SUMOylated PML, which is consistent with the report of Schilling et al. (J Virol 91:e02049-16, 2017, https://doi.org/10.1128/JVI.02049-16). The mono-SUMOylated PML was subjected to degradation by HSV ICP0. However, it was protected by IE1 from degradation by ICP0 or HSV-1 infection. In contrast, IE1 with L174P mutation lost the function of deSUMOylating PML and failed to protect the degradation of the mono-SUMOylated PML. Whether the mono-SUMOylated PML has any defensive function against viral infection will be further investigated.


Assuntos
Infecções por Citomegalovirus/metabolismo , Herpes Simples/metabolismo , Proteínas Imediatamente Precoces/metabolismo , Proteína da Leucemia Promielocítica/metabolismo , Proteólise , Sumoilação , Ubiquitina-Proteína Ligases/metabolismo , Animais , Células Cultivadas , Citomegalovirus/fisiologia , Infecções por Citomegalovirus/genética , Infecções por Citomegalovirus/virologia , Herpes Simples/genética , Herpes Simples/virologia , Herpesvirus Humano 1/fisiologia , Humanos , Proteínas Imediatamente Precoces/genética , Camundongos , Proteína da Leucemia Promielocítica/química , Proteína da Leucemia Promielocítica/genética , Ubiquitina-Proteína Ligases/genética , Replicação Viral
18.
J Virol ; 92(17)2018 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-29950413

RESUMO

The mechanisms underlying neurodevelopmental damage caused by virus infections remain poorly defined. Congenital human cytomegalovirus (HCMV) infection is the leading cause of fetal brain development disorders. Previous work has linked HCMV infection to perturbations of neural cell fate, including premature differentiation of neural progenitor cells (NPCs). Here, we show that HCMV infection of NPCs results in loss of the SOX2 protein, a key pluripotency-associated transcription factor. SOX2 depletion maps to the HCMV major immediate early (IE) transcription unit and is individually mediated by the IE1 and IE2 proteins. IE1 causes SOX2 downregulation by promoting the nuclear accumulation and inhibiting the phosphorylation of STAT3, a transcriptional activator of SOX2 expression. Deranged signaling resulting in depletion of a critical stem cell protein is an unanticipated mechanism by which the viral major IE proteins may contribute to brain development disorders caused by congenital HCMV infection.IMPORTANCE Human cytomegalovirus (HCMV) infections are a leading cause of brain damage, hearing loss, and other neurological disabilities in children. We report that the HCMV proteins known as IE1 and IE2 target expression of human SOX2, a central pluripotency-associated transcription factor that governs neural progenitor cell (NPC) fate and is required for normal brain development. Both during HCMV infection and when expressed alone, IE1 causes the loss of SOX2 from NPCs. IE1 mediates SOX2 depletion by targeting STAT3, a critical upstream regulator of SOX2 expression. Our findings reveal an unanticipated mechanism by which a common virus may cause damage to the developing nervous system and suggest novel targets for medical intervention.


Assuntos
Citomegalovirus/crescimento & desenvolvimento , Interações Hospedeiro-Patógeno , Proteínas Imediatamente Precoces/metabolismo , Células-Tronco Neurais/patologia , Células-Tronco Neurais/virologia , Fatores de Transcrição SOXB1/metabolismo , Fator de Transcrição STAT3/metabolismo , Células Cultivadas , Humanos
19.
Cell Host Microbe ; 24(1): 69-80.e4, 2018 07 11.
Artigo em Inglês | MEDLINE | ID: mdl-29937271

RESUMO

The cytosolic DNA sensor cGAS recognizes viral DNA and synthesizes the second messenger cGAMP upon viral infection. cGAMP binds to the adaptor protein MITA/STING to activate downstream signaling events, leading to induction of type I interferons (IFNs) and antiviral effector genes. Here we identify the human cytomegalovirus (HCMV) protein UL31 as an inhibitor of cGAS. UL31 interacts directly with cGAS and disassociates DNA from cGAS, thus inhibiting cGAS enzymatic functions and reducing cGAMP production. UL31 overexpression markedly reduces antiviral responses stimulated by cytosolic DNA, while knockdown or knockout of UL31 heightens HCMV-triggered induction of type I IFNs and downstream antiviral genes. Moreover, wild-type HCMV replicates more efficiently than UL31-deficient HCMV, a phenotype that is reversed in cGAS null cells. These results highlight the importance of cGAS in the host response to HCMV as well as an important viral strategy to evade this innate immune sensor.


Assuntos
Citomegalovirus/fisiologia , Evasão da Resposta Imune/imunologia , Proteínas Nucleares/metabolismo , Nucleotidiltransferases/antagonistas & inibidores , Proteínas Virais/metabolismo , Citomegalovirus/genética , DNA Viral/genética , DNA Viral/metabolismo , Fibroblastos , Técnicas de Silenciamento de Genes , Técnicas de Inativação de Genes , Células HEK293 , Humanos , Imunidade Inata/imunologia , Interferon Tipo I/metabolismo , Proteínas Nucleares/genética , Nucleotídeos Cíclicos/metabolismo , Nucleotidiltransferases/genética , Cultura Primária de Células , Proteínas Virais/genética
20.
Front Microbiol ; 9: 1067, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29922247

RESUMO

Zika virus (ZIKV) infection is associated with severe neurological defects in fetuses and newborns, such as microcephaly. However, the underlying mechanisms remain to be elucidated. In this study, proteomic analysis on ZIKV-infected primary human fetal neural progenitor cells (NPCs) revealed that virus infection altered levels of cellular proteins involved in NPC proliferation, differentiation and migration. The transcriptional levels of some of the altered targets were also confirmed by qRT-PCR. Among the altered proteins, doublecortin (DCX) plays an important role in NPC differentiation and migration. Results showed that ZIKV infection downregulated DCX, at both mRNA and protein levels, as early as 1 day post infection (1 dpi), and lasted throughout the virus replication cycle (4 days). The downregulation of DCX was also observed in a ZIKV-infected fetal mouse brain model, which displayed decreased body weight, brain size and weight, as well as defective cortex structure. By screening the ten viral proteins of ZIKV, we found that both the expression of NS4A and NS5 were correlated with the downregulation of both mRNA and protein levels of DCX in NPCs. These data suggest that DCX is modulated following infection of the brain by ZIKV. How these observed changes of DCX expression translate in the pathological consequences of ZIKV infection and if other cellular proteins are equally involved remains to be investigated.

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