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1.
J Virol ; 96(13): e0061822, 2022 07 13.
Article in English | MEDLINE | ID: covidwho-1962091

ABSTRACT

Porcine epidemic diarrhea virus (PEDV) is the globally distributed alphacoronavirus that can cause lethal watery diarrhea in piglets, causing substantial economic damage. However, the current commercial vaccines cannot effectively the existing diseases. Thus, it is of great necessity to identify the host antiviral factors and the mechanism by which the host immune system responds against PEDV infection required to be explored. The current work demonstrated that the host protein, the far upstream element-binding protein 3 (FUBP3), could be controlled by the transcription factor TCFL5, which could suppress PEDV replication through targeting and degrading the nucleocapsid (N) protein of the virus based on selective autophagy. For the ubiquitination of the N protein, FUBP3 was found to recruit the E3 ubiquitin ligase MARCH8/MARCHF8, which was then identified, transported to, and degraded in autolysosomes via NDP52/CALCOCO2 (cargo receptors), resulting in impaired viral proliferation. Additionally, FUBP3 was found to positively regulate type-I interferon (IFN-I) signaling and activate the IFN-I signaling pathway by interacting and increasing the expression of tumor necrosis factor (TNF) receptor-associated factor 3 (TRAF3). Collectively, this study showed a novel mechanism of FUBP3-mediated virus restriction, where FUBP3 was found to degrade the viral N protein and induce IFN-I production, aiming to hinder the replication of PEDV. IMPORTANCE PEDV refers to the alphacoronavirus that is found globally and has re-emerged recently, causing severe financial losses. In PEDV infection, the host activates various host restriction factors to maintain innate antiviral responses to suppress virus replication. Here, FUBP3 was detected as a new host restriction factor. FUBP3 was found to suppress PEDV replication via the degradation of the PEDV-encoded nucleocapsid (N) protein via E3 ubiquitin ligase MARCH8 as well as the cargo receptor NDP52/CALCOCO2. Additionally, FUBP3 upregulated the IFN-I signaling pathway by interacting with and increasing tumor necrosis factor (TNF) receptor-associated factor 3 (TRAF3) expression. This study further demonstrated that another layer of complexity could be added to the selective autophagy and innate immune response against PEDV infection are complicated.


Subject(s)
Coronavirus Infections , Interferon Type I , Nucleocapsid Proteins , Porcine epidemic diarrhea virus , Transcription Factors , Animals , Antiviral Agents , Cell Line , Chlorocebus aethiops , Coronavirus Infections/metabolism , Interferon Type I/genetics , Interferon Type I/metabolism , Nucleocapsid Proteins/metabolism , Porcine epidemic diarrhea virus/physiology , Swine , TNF Receptor-Associated Factor 3 , Transcription Factors/metabolism , Ubiquitin-Protein Ligases , Vero Cells
2.
J Virol ; 96(10): e0007022, 2022 05 25.
Article in English | MEDLINE | ID: covidwho-1832352

ABSTRACT

In global infection and serious morbidity and mortality, porcine epidemic diarrhea virus (PEDV) has been regarded as a dreadful porcine pathogen, but the existing commercial vaccines are not enough to fully protect against the epidemic strains. Therefore, it is of great necessity to feature the PEDV-host interaction and develop efficient countermeasures against viral infection. As an RNA/DNA protein, the trans-active response DNA binding protein (TARDBP) plays a variety of functions in generating and processing RNA, including transcription, splicing, transport, and mRNA stability, which have been reported to regulate viral replication. The current work aimed to detect whether and how TARDBP influences PEDV replication. Our data demonstrated that PEDV replication was significantly suppressed by TARDBP, regulated by KLF16, which targeted its promoter. We observed that through the proteasomal and autophagic degradation pathway, TARDBP inhibited PEDV replication via the binding as well as degradation of PEDV-encoded nucleocapsid (N) protein. Moreover, we found that TARDBP promoted autophagic degradation of N protein via interacting with MARCHF8, an E3 ubiquitin ligase, as well as NDP52, a cargo receptor. We also showed that TARDBP promoted host antiviral innate immune response by inducing interferon (IFN) expression through the MyD88-TRAF3-IRF3 pathway during PEDV infection. In conclusion, these data revealed a new antiviral role of TARDBP, effectively suppressing PEDV replication through degrading virus N protein via the proteasomal and autophagic degradation pathway and activating type I IFN signaling via upregulating the expression of MyD88. IMPORTANCE PEDV refers to the highly contagious enteric coronavirus that has quickly spread globally and generated substantial financial damage to the global swine industry. During virus infection, the host regulates the innate immunity and autophagy process to inhibit virus infection. However, the virus has evolved plenty of strategies with the purpose of limiting IFN-I production and autophagy processes. Here, we identified that TARDBP expression was downregulated via the transcription factor KLF16 during PEDV infection. TARDBP could inhibit PEDV replication through the combination as well as degradation of PEDV-encoded nucleocapsid (N) protein via proteasomal and autophagic degradation pathways and promoted host antiviral innate immune response by inducing IFN expression through the MyD88-TRAF3-IRF3 pathway. In sum, our data identify a novel antiviral function of TARDBP and provide a better grasp of the innate immune response and protein degradation pathway against PEDV infection.


Subject(s)
Coronavirus Infections , DNA-Binding Proteins , Interferon Type I , Porcine epidemic diarrhea virus , Virus Replication , Animals , Coronavirus Infections/veterinary , DNA-Binding Proteins/metabolism , Immunity, Innate , Interferon Regulatory Factor-3/metabolism , Interferon Type I/metabolism , Myeloid Differentiation Factor 88/metabolism , Nucleocapsid Proteins/metabolism , Porcine epidemic diarrhea virus/genetics , Porcine epidemic diarrhea virus/physiology , RNA/metabolism , Signal Transduction , Swine , TNF Receptor-Associated Factor 3/metabolism
3.
Microbiome ; 10(1): 60, 2022 04 12.
Article in English | MEDLINE | ID: covidwho-1789144

ABSTRACT

BACKGROUND: Wild birds may harbor and transmit viruses that are potentially pathogenic to humans, domestic animals, and other wildlife. RESULTS: Using the viral metagenomic approach, we investigated the virome of cloacal swab specimens collected from 3182 birds (the majority of them wild species) consisting of > 87 different species in 10 different orders within the Aves classes. The virus diversity in wild birds was higher than that in breeding birds. We acquired 707 viral genomes from 18 defined families and 4 unclassified virus groups, with 265 virus genomes sharing < 60% protein sequence identities with their best matches in GenBank comprising new virus families, genera, or species. RNA viruses containing the conserved RdRp domain with no phylogenetic affinity to currently defined virus families existed in different bird species. Genomes of the astrovirus, picornavirus, coronavirus, calicivirus, parvovirus, circovirus, retrovirus, and adenovirus families which include known avian pathogens were fully characterized. Putative cross-species transmissions were observed with viruses in wild birds showing > 95% amino acid sequence identity to previously reported viruses in domestic poultry. Genomic recombination was observed for some genomes showing discordant phylogenies based on structural and non-structural regions. Mapping the next-generation sequencing (NGS) data respectively against the 707 genomes revealed that these viruses showed distribution pattern differences among birds with different habitats (breeding or wild), orders, and sampling sites but no significant differences between birds with different behavioral features (migratory and resident). CONCLUSIONS: The existence of a highly diverse virome highlights the challenges in elucidating the evolution, etiology, and ecology of viruses in wild birds. Video Abstract.


Subject(s)
RNA Viruses , Viruses , Animals , Animals, Wild , Birds , Cloaca , Phylogeny , RNA Viruses/genetics , Virome/genetics , Viruses/genetics
4.
Microbiol Spectr ; 9(2): e0090821, 2021 10 31.
Article in English | MEDLINE | ID: covidwho-1452921

ABSTRACT

Emerging coronaviruses (CoVs) can cause severe diseases in humans and animals, and, as of yet, none of the currently available broad-spectrum drugs or vaccines can effectively control these diseases. Host antiviral proteins play an important role in inhibiting viral proliferation. One of the isoforms of cytoplasmic poly(A)-binding protein (PABP), PABPC4, is an RNA-processing protein, which plays an important role in promoting gene expression by enhancing translation and mRNA stability. However, its function in viruses remains poorly understood. Here, we report that the host protein, PABPC4, could be regulated by transcription factor SP1 and broadly inhibits the replication of CoVs, covering four genera (Alphacoronavirus, Betacoronavirus, Gammacoronavirus, and Deltacoronavirus) of the Coronaviridae family by targeting the nucleocapsid (N) protein through the autophagosomes for degradation. PABPC4 recruited the E3 ubiquitin ligase MARCH8/MARCHF8 to the N protein for ubiquitination. Ubiquitinated N protein was recognized by the cargo receptor NDP52/CALCOCO2, which delivered it to the autolysosomes for degradation, resulting in impaired viral proliferation. In addition to regulating gene expression, these data demonstrate a novel antiviral function of PABPC4, which broadly suppresses CoVs by degrading the N protein via the selective autophagy pathway. This study will shed light on the development of broad anticoronaviral therapies. IMPORTANCE Emerging coronaviruses (CoVs) can cause severe diseases in humans and animals, but none of the currently available drugs or vaccines can effectively control these diseases. During viral infection, the host will activate the interferon (IFN) signaling pathways and host restriction factors in maintaining the innate antiviral responses and suppressing viral replication. This study demonstrated that the host protein, PABPC4, interacts with the nucleocapsid (N) proteins from eight CoVs covering four genera (Alphacoronavirus, Betacoronavirus, Gammacoronavirus, and Deltacoronavirus) of the Coronaviridae family. PABPC4 could be regulated by SP1 and broadly inhibits the replication of CoVs by targeting the nucleocapsid (N) protein through the autophagosomes for degradation. This study significantly increases our understanding of the novel host restriction factor PABPC4 against CoV replication and will help develop novel antiviral strategies.


Subject(s)
Autophagy/physiology , Blood Proteins/metabolism , Coronavirus Nucleocapsid Proteins/metabolism , Coronavirus/growth & development , Poly(A)-Binding Proteins/metabolism , Virus Replication/physiology , Animals , Cell Line , Chlorocebus aethiops , HEK293 Cells , Humans , Infectious bronchitis virus/growth & development , Murine hepatitis virus/growth & development , Nuclear Proteins/metabolism , Porcine epidemic diarrhea virus/growth & development , Proteolysis , Sp1 Transcription Factor/metabolism , Swine , Ubiquitin-Protein Ligases/metabolism , Ubiquitination , Vero Cells
5.
J Med Virol ; 93(3): 1786-1791, 2021 03.
Article in English | MEDLINE | ID: covidwho-1196491

ABSTRACT

Pangolin metagenomic data obtained from public databases were used to assemble partial or complete viral genomes showing genetic relationship to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), Sendai virus, flavivirus, picornavirus, parvovirus, and genomovirus, respectively. Most of these virus genomes showed genomic recombination signals. Phylogeny based on the SARS-CoV-2-related virus sequences assembled in this study and those recently published indicated that pangolin SARS-CoV-2-related viruses were clustered into two sub-lineages according to geographic sampling sites. These findings suggest the need for further pangolin samples, from different countries, to be collected and analyzed for coronavirus to elucidate whether pangolins are intermittent hosts for SARS-CoV-2.


Subject(s)
COVID-19/virology , Genome, Viral/genetics , Metagenome/genetics , Pangolins/virology , SARS-CoV-2/genetics , Animals , Host Specificity/genetics , Metagenomics/methods , Phylogeny , Recombination, Genetic/genetics
6.
Arch Virol ; 165(12): 2847-2856, 2020 Dec.
Article in English | MEDLINE | ID: covidwho-841309

ABSTRACT

Here, we investigated the fecal, oral, blood, and skin virome of 10 laboratory rabbits using a viral metagenomic method. In the oral samples, we detected a novel polyomavirus (RabPyV), and phylogenetic analysis based on the large T antigen, VP1 and VP2 regions indicated that the novel strain might have undergone a recombination event. Recombination analysis based on related genomes confirmed that RabPyV is a multiple recombinant between rodent-like and avian-like polyomaviruses. In fecal samples, three partial or complete genome sequences of viruses belonging to the families Picobirnaviridae, Parvoviridae, Microviridae and Coronaviridae were characterized, and phylogenetic trees were constructed based on the predicted amino acid sequences of viral proteins. This study increases the amount of genetic information on viruses present in laboratory rabbits.


Subject(s)
Metagenome , Polyomavirus/isolation & purification , Rabbits/virology , Viral Proteins/genetics , Viruses/classification , Animals , Animals, Laboratory/virology , Blood/virology , Feces/virology , Genome, Viral , Mouth/virology , Phylogeny , Skin/virology , Viruses/isolation & purification , Whole Genome Sequencing
7.
Virol J ; 17(1): 46, 2020 04 03.
Article in English | MEDLINE | ID: covidwho-827237

ABSTRACT

BACKGROUND: Porcine epidemic diarrhea virus (PEDV) infection causes an acute enteric tract infectious disease characterized by vomiting, anorexia, dehydration, weight loss and high mortality in neonatal piglets. During PEDV infection, the spike protein (S) is a major virion structural protein interacting with receptors and inducing neutralizing antibodies. However, the neutralizing B-cell epitopes within PEDV S protein have not been well studied. METHODS: To accurately identify the important immunodominant region of S1, the purified truncated S1 proteins (SA, SB, SC, SD and SE) were used to immunize BALB/c mice to prepare polyclonal antibodies. The antisera titers were determined by indirect ELISA, western blot and IFA after four immunizations to find the important immunodominant region of S1, and then purified the immunodominant region of S1 protein and immunized mice to generate the special antibodies, and then used recombinant peptides to determine the B-cell epitopes of monoclonal antibodies. RESULTS: Five antisera of recombinant proteins of the spike protein region of PEDV were generated and we found that only the polyclonal antibody against part of the S1 region (signed as SE protein, residues 666-789) could recognize the native PEDV. Purified SE protein was used to immunize BALB/c mice and generate mAb 2E10. Pepscan of the SE protein demonstrated that SE16 (722SSTFNSTREL731) is the minimal linear epitope required for reactivity with the mAb 2E10. Further investigation indicated that the epitope SE16 was localized on the surface of PEDV S protein in the 3D structure. CONCLUSIONS: A mAb 2E10 that is specifically bound to PEDV was generated and identified a specific linear B-cell epitope (SE16, 722SSTFNSTREL731) of the mAb. The epitope region of PEDV S1 localized in the different regions in comparison with the earlier identified epitopes. These findings enhance the understanding of the PEDV spike protein structure for vaccine design and provide a potential use for developing diagnostic methods to detect PEDV.


Subject(s)
Epitopes, B-Lymphocyte/immunology , Immunodominant Epitopes/immunology , Porcine epidemic diarrhea virus/immunology , Spike Glycoprotein, Coronavirus/immunology , Animals , Antibodies, Monoclonal/immunology , Antibodies, Neutralizing/immunology , Chlorocebus aethiops , Female , Mice , Mice, Inbred BALB C , Porcine epidemic diarrhea virus/chemistry , Vero Cells
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