Hantaan virus inhibits type I interferon response by targeting RLR signaling pathways through TRIM25.

Hantaan virus (HTNV) is responsible for hemorrhagic fever with renal syndrome (HFRS), primarily due to its ability to inhibit host innate immune responses, such as type I interferon (IFN-I). In this study, we conducted a transcriptome analysis to identify host factors regulated by HTNV nucleocapsid protein (NP) and glycoprotein. Our findings demonstrate that NP and Gc proteins inhibit host IFN-I production by manipulating the retinoic acid-induced gene I (RIG-I)-like receptor (RLR) pathways. Further analysis reveals that HTNV NP and Gc proteins target upstream molecules of MAVS, such as RIG-I and MDA-5, with Gc exhibiting stronger inhibition of IFN-I responses than NP. Mechanistically, NP and Gc proteins interact with tripartite motif protein 25 (TRIM25) to competitively inhibit its interaction with RIG-I/MDA5, suppressing RLR signaling pathways. Our study unveils a cross-talk between HTNV NP/Gc proteins and host immune response, providing valuable insights into the pathogenic mechanism of HTNV.

Animal models of mpox virus infection and disease.

Mpox (monkeypox) virus (MPXV), which causes a mild smallpox-like disease, has been endemic in Africa for several decades, with sporadic cases occurring in other parts of the world. However, the most recent outbreak of mpox mainly among men that have sex with men has affected several continents, posing serious global public health concerns. The infections exhibit a wide spectrum of clinical presentation, ranging from asymptomatic infection to mild, severe disease, especially in immunocompromised individuals, young children, and pregnant women. Some therapeutics and vaccines developed for smallpox have partial protective and therapeutic effects against MPXV historic isolates in animal models. However, the continued evolution of MPXV has produced multiple lineages, leading to significant gaps in the knowledge of their pathogenesis that constrain the development of targeted antiviral therapies and vaccines. MPXV infections in various animal models have provided a central platform for identification and comparison of diseased pathogenesis between the contemporary and historic isolates. In this review, we discuss the susceptibility of various animals to MPXV, and describe the key pathologic features of rodent, rabbit and nonhuman primate models. We also provide application examples of animal models in elucidating viral pathogenesis and evaluating effectiveness of vaccine and antiviral drugs. These animal models are essential to understand the biology of MPXV contemporary isolates and to rapidly test potential countermeasures. Finally, we list some remaining scientific questions of MPXV that can be resolved by animal models.

HDAC6 decreases H4K16 and α-tubulin acetylation during porcine oocyte maturation.

HDAC6 is an essential factor in mouse oocyte maturation. However, the roles of HDAC6 in porcine oocyte maturation are still unclear. Therefore, we analyzed the roles of HDAC6 in porcine oocyte maturation by treatment with Tubastatin A (TubA) which is an HDAC6 inhibitor. Our results showed that treatment with 10 µg/ml TubA significantly decreased the rate of porcine oocyte maturation, but it did not influence the rate of germinal vesicle breakdown (GVBD). Then, we found that TubA treatment disrupted spindle organization by increasing the α-tubulin acetylation level during porcine oocyte maturation. Moreover, TubA treatment significantly increased H4K16 acetylation, which may compromise kinetochore and microtubule (K-MT) attachment during meiosis in porcine oocytes. We also analyzed the effects of TubA on meiosis-related (H3T3pho and H3S10pho) and transcription-related histone modifications (H3K4me3, H3K9me3 and H3K4ac) during porcine oocyte maturation. The results showed that TubA treatment increased H3S10pho and H3K4ac levels, but no influence was seen in H3T3pho, H3K4me3 and H3K9me3 levels in porcine oocytes. TubA treated oocytes also showed a compromised ability to develop after parthenogenetic activation. Finally, we found that HDAC6 exhibited higher mRNA levels and lower DNA methylation levels in porcine oocytes than it did in porcine embryonic fibroblasts (PEFs). These results indicate that the low level of DNA methylation in HDAC6 promoter ensures high expression. HDAC6 regulates the deacetylation of α-tubulin and H4K16, which promotes correct spindle organization and meiotic apparatus assembly during porcine oocyte maturation. This study illustrates a new pathway by which HDAC6 modulates mammalian oocyte maturation.

EGR1 functions as a new host restriction factor for SARS-CoV-2 to inhibit virus replication through the E3 ubiquitin ligase MARCH8.

IMPORTANCE: Emerging vaccine-breakthrough severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants highlight an urgent need for novel antiviral therapies. Understanding the pathogenesis of coronaviruses is critical for developing antiviral drugs. Here, we demonstrate that the SARS-CoV-2 N protein suppresses interferon (IFN) responses by reducing early growth response gene-1 (EGR1) expression. The overexpression of EGR1 inhibits SARS-CoV-2 replication by promoting IFN-regulated antiviral protein expression, which interacts with and degrades SARS-CoV-2 N protein via the E3 ubiquitin ligase MARCH8 and the cargo receptor NDP52. The MARCH8 mutants without ubiquitin ligase activity are no longer able to degrade SARS-CoV-2 N proteins, indicating that MARCH8 degrades SARS-CoV-2 N proteins dependent on its ubiquitin ligase activity. This study found a novel immune evasion mechanism of SARS-CoV-2 utilized by the N protein, which is helpful for understanding the pathogenesis of SARS-CoV-2 and guiding the design of new prevention strategies against the emerging coronaviruses.

Yezo Virus Infection in Tick-Bitten Patient and Ticks, Northeastern China.

We identified Yezo virus infection in a febrile patient who had a tick bite in northeastern China, where 0.5% of Ixodes persulcatus ticks were positive for viral RNA. Clinicians should be aware of this potential health threat and include this emerging virus in the differential diagnosis for tick-bitten patients in this region.

Flavivirus prM interacts with MDA5 and MAVS to inhibit RLR antiviral signaling.

BACKGROUND: Vector-borne flaviviruses, including tick-borne encephalitis virus (TBEV), Zika virus (ZIKV), West Nile virus (WNV), yellow fever virus (YFV), dengue virus (DENV), and Japanese encephalitis virus (JEV), pose a growing threat to public health worldwide, and have evolved complex mechanisms to overcome host antiviral innate immunity. However, the underlying mechanisms of flavivirus structural proteins to evade host immune response remain elusive. RESULTS: We showed that TBEV structural protein, pre-membrane (prM) protein, could inhibit type I interferon (IFN-I) production. Mechanically, TBEV prM interacted with both MDA5 and MAVS and interfered with the formation of MDA5-MAVS complex, thereby impeding the nuclear translocation and dimerization of IRF3 to inhibit RLR antiviral signaling. ZIKV and WNV prM was also demonstrated to interact with both MDA5 and MAVS, while dengue virus serotype 2 (DENV2) and YFV prM associated only with MDA5 or MAVS to suppress IFN-I production. In contrast, JEV prM could not suppress IFN-I production. Overexpression of TBEV and ZIKV prM significantly promoted the replication of TBEV and Sendai virus. CONCLUSION: Our findings reveal the immune evasion mechanisms of flavivirus prM, which may contribute to understanding flavivirus pathogenicity, therapeutic intervention and vaccine development.

Extensive diversity of RNA viruses in ticks revealed by metagenomics in northeastern China.

BACKGROUND: Ticks act as important vectors of infectious agents, and several emerging tick-borne viruses have recently been identified to be associated with human diseases in northeastern China. However, little is known about the tick virome in northeastern China. METHODS: Ticks collected from April 2020 to July 2021 were pooled for metagenomic analysis to investigate the virome diversity in northeastern China. RESULTS: In total, 22 RNA viruses were identified, including four each in the Nairoviridae and Phenuiviridae families, three each in the Flaviviridae, Rhabdoviridae, and Solemoviridae families, two in the Chuviridae family, and one each in the Partitiviridae, Tombusviridae families and an unclassified virus. Of these, eight viruses were of novel species, belonging to the Nairoviridae (Ji'an nairovirus and Yichun nairovirus), Phenuiviridae (Mudanjiang phlebovirus), Rhabdoviridae (Tahe rhabdovirus 1-3), Chuviridae (Yichun mivirus), and Tombusviridae (Yichun tombus-like virus) families, and five members were established human pathogens, including Alongshan virus, tick-borne encephalitis virus, Songling virus, Beiji nairovirus, and Nuomin virus. I. persulcatus ticks had significant higher number of viral species than H. japonica, H. concinna, and D. silvarum ticks. Significant differences in tick viromes were observed among Daxing'an, Xiaoxing'an and Changbai mountains. CONCLUSIONS: These findings showed an extensive diversity of RNA viruses in ticks in northeastern China, revealing potential public health threats from the emerging tick-borne viruses. Further studies are needed to explain the natural circulation and pathogenicity of these viruses.

Characterization and subcellular localization of Alongshan virus proteins.

Alongshan virus (ALSV) in the Jingmenvirus group within the family Flaviviridae is a newly discovered tick-borne virus associated with human disease, whose genome includes four segments and encodes four structural proteins (VP1a, VP1b, VP2, VP3, and VP4) and two non-structural proteins (NSP1 and NSP2). Here, we characterized the subcellular distribution and potential function of ALSV proteins in host cells. We found that viral proteins exhibited diverse subcellular distribution in multiple tissue-deriving cells and induced various morphological changes in the endoplasmic reticulum (ER), and NSP2, VP1b, VP2, and VP4 were all co-localized in the ER. The nuclear transfer and co-localization of VP4 and calnexin (a marker protein of ER), which were independent of their interaction, were unique to HepG2 cells. Expression of NSP1 could significantly reduce mitochondria quantity by inducing mitophagy. These findings would contribute to better understanding of the pathogenesis of emerging segmented flaviviruses.

The global succinylation of SARS-CoV-2-infected host cells reveals drug targets.

SARS-CoV-2, the causative agent of the COVID-19 pandemic, undergoes continuous evolution, highlighting an urgent need for development of novel antiviral therapies. Here we show a quantitative mass spectrometry-based succinylproteomics analysis of SARS-CoV-2 infection in Caco-2 cells, revealing dramatic reshape of succinylation on host and viral proteins. SARS-CoV-2 infection promotes succinylation of several key enzymes in the TCA, leading to inhibition of cellular metabolic pathways. We demonstrated that host protein succinylation is regulated by viral nonstructural protein (NSP14) through interaction with sirtuin 5 (SIRT5); overexpressed SIRT5 can effectively inhibit virus replication. We found succinylation inhibitors possess significant antiviral effects. We also found that SARS-CoV-2 nucleocapsid and membrane proteins underwent succinylation modification, which was conserved in SARS-CoV-2 and its variants. Collectively, our results uncover a regulatory mechanism of host protein posttranslational modification and cellular pathways mediated by SARS-CoV-2, which may become antiviral drug targets against COVID-19.

Tamarixetin Attenuated the Virulence of Staphylococcus aureus by Directly Targeting Caseinolytic Protease P.

Staphylococcus aureus, especially drug-resistant S. aureus infections, is a worldwide healthcare challenge. There is a growing focus on antivirulence therapy against S. aureus. Caseinolytic protease p (ClpP) is a protein hydrolase essential for pathogenicity in S. aureus. A flavonoid compound, tamarixetin, which was screened in this work, was specifically able to inhibit the hydrolytic activity of ClpP on the fluorescent substrate Suc-LY-AMC with an IC50 of 49.73 µM, without affecting the growth of methicillin-resistant S. aureus strain USA300 and was without obvious cytotoxicity. Further assays found that tamarixetin inhibited the transcription of hla, agr, RNAIII, pvl, PSM-α, and spa genes as well as suppressed the protein expression levels of Hla and PVL. Moreover, tamarixetin was observed to dramatically inhibit the hemolytic activity of hla in S. aureus. Consistent with that of S. aureus USA300-ΔclpP, tamarixetin was shown to increase urease expression. The thermal shift and cellular thermal shift assays showed that tamarixetin markedly changed the thermal stability of ClpP. The dissociation constant (KD) value of tamarixetin with ClpP was 2.52 × 10-6 M measured by surface plasmon resonance. The molecular docking and ClpP point mutation results also demonstrated that tamarixetin had a strong interaction with ClpP. In vivo study showed that tamarixetin was effective in protecting mice from S. aureus pneumonia by increasing survival, reducing lung tissue load, and slowing down the infiltration of inflammatory factors. In addition, tamarixetin was able to enhance the antibacterial activity of cefotaxime in combination. In conclusion, tamarixetin was promising as a ClpP inhibitor for S. aureus infections.

Hinokiflavone Attenuates the Virulence of Methicillin-Resistant Staphylococcus aureus by Targeting Caseinolytic Protease P.

Drug-resistant bacteria was the third leading cause of death worldwide in 2019, which sounds like a cautionary note for global public health. Therefore, developing novel strategies to combat Methicillin-resistant Staphylococcus aureus (MRSA) infections is the need of the hour. Caseinolytic protease P (ClpP) represents pivotal microbial degradation machinery in MRSA involved in bacterial homeostasis and pathogenicity, considered an ideal target for combating S. aureus infections. Herein, we identified a natural compound, hinokiflavone, that inhibited the activity of ClpP of MRSA strain USA300 with an IC50 of 34.36 µg/mL. Further assays showed that hinokiflavone reduced the virulence of S. aureus by inhibiting multiple virulence factors expression. Results obtained from cellular thermal transfer assay (CETSA), thermal shift assay (TSA), local surface plasmon resonance (LSPR) and molecular docking (MD) assay enunciated that hinokiflavone directly bonded to ClpP with confirmed docking sites, including SER-22, LYS-26 and ARG-28. In vivo, the evaluation of anti-infective activity showed that hinokiflavone in combination with vancomycin effectively protected mice from MRSA-induced fatal pneumonia, which was more potent than vancomycin alone. As mentioned above, hinokiflavone, as an inhibitor of ClpP, could be further developed into a promising adjuvant against S. aureus infections.

A dual-role of SARS-CoV-2 nucleocapsid protein in regulating innate immune response.

The recently emerged severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which is the causative agent of ongoing global pandemic of COVID-19, may trigger immunosuppression in the early stage and overactive immune response in the late stage of infection; However, the underlying mechanisms are not well understood. Here we demonstrated that the SARS-CoV-2 nucleocapsid (N) protein dually regulated innate immune responses, i.e., the low-dose N protein suppressed type I interferon (IFN-I) signaling and inflammatory cytokines, whereas high-dose N protein promoted IFN-I signaling and inflammatory cytokines. Mechanistically, the SARS-CoV-2 N protein dually regulated the phosphorylation and nuclear translocation of IRF3, STAT1, and STAT2. Additionally, low-dose N protein combined with TRIM25 could suppress the ubiquitination and activation of retinoic acid-inducible gene I (RIG-I). Our findings revealed a regulatory mechanism of innate immune responses by the SARS-CoV-2 N protein, which would contribute to understanding the pathogenesis of SARS-CoV-2 and other SARS-like coronaviruses, and development of more effective strategies for controlling COVID-19.

SARS-CoV-2 Membrane Protein Inhibits Type I Interferon Production Through Ubiquitin-Mediated Degradation of TBK1.

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is the causative pathogen of current COVID-19 pandemic, and insufficient production of type I interferon (IFN-I) is associated with the severe forms of the disease. Membrane (M) protein of SARS-CoV-2 has been reported to suppress host IFN-I production, but the underlying mechanism is not completely understood. In this study, SARS-CoV-2 M protein was confirmed to suppress the expression of IFNß and interferon-stimulated genes induced by RIG-I, MDA5, IKKϵ, and TBK1, and to inhibit IRF3 phosphorylation and dimerization caused by TBK1. SARS-CoV-2 M could interact with MDA5, TRAF3, IKKϵ, and TBK1, and induce TBK1 degradation via K48-linked ubiquitination. The reduced TBK1 further impaired the formation of TRAF3-TANK-TBK1-IKKε complex that leads to inhibition of IFN-I production. Our study revealed a novel mechanism of SARS-CoV-2 M for negative regulation of IFN-I production, which would provide deeper insight into the innate immunosuppression and pathogenicity of SARS-CoV-2.

A new nairo-like virus associated with human febrile illness in China.

ABSTRACTSeveral nairo-like viruses have been discovered in ticks in recent years, but their relevance to public health remains unknown. Here, we found a patient who had a history of tick bite and suffered from a febrile illness was infected with a previously discovered RNA virus, Beiji nairovirus (BJNV), in the nairo-like virus group of the order Bunyavirales. We isolated the virus by cell culture assay. BJNV could induce cytopathic effects in the baby hamster kidney and human hepatocellular carcinoma cells. Negative-stain electron microscopy revealed enveloped and spherical viral particles, morphologically similar to those of nairoviruses. We identified 67 patients as BJNV infection in 2017-2018. The median age of patients was 48 years (interquartile range 41-53 years); the median incubation period was 7 days (interquartile range 3-12 days). Most patients were men (70%), and a few (10%) had underlying diseases. Common symptoms of infected patients included fever (100%), headache (99%), depression (63%), coma (63%), and fatigue (54%), myalgia or arthralgia (45%); two (3%) patients became critically ill and one died. BJNV could cause growth retardation, viremia and histopathological changes in infected suckling mice. BJNV was also detected in sheep, cattle, and multiple tick species. These findings demonstrated that the newly discovered nairo-like virus may be associated with a febrile illness, with the potential vectors of ticks and reservoirs of sheep and cattle, highlighting its public health significance and necessity of further investigation in the tick-endemic areas worldwide.

HDAC11 inhibition disrupts porcine oocyte meiosis via regulating α-tubulin acetylation and histone modifications.

HDAC11, the sole member of HDAC class IV family, plays vital roles in activating mitosis and apoptosis of tumor cells, but its functions in meiosis are rarely investigated. In the present study, the effect of HDAC11 on meiosis during porcine oocytes maturation was fully studied. The results showed that HDAC11 inhibition by its specific inhibitor JB-3-22 dramatically decreased the porcine oocyte maturation rate by disturbing spindle organization and chromosomes alignment without affecting the cytoplasmic maturation. Further study indicated that HDAC11 inhibition significantly elevated the acetylation levels of α-tubulin and H4K16, which are crucial for spindle organization and chromosomes alignment. Moreover, immunofluorescence staining results showed that HDAC11 inhibition also disturbed other meiosis-related histone modifications, such as increased H3S10pho, H4K5ac and H4K12ac levels and reduced H3T3pho level. Furthermore, RNA-seq analysis results indicated that HDAC11 inhibition disturbed porcine oocytes transcriptome (157 up-regulation, 106 down-regulation). In addition, HDAC11 inhibition compromised oocytes quality and subsequent development after parthenogenetic activation, which may be caused by the aberrant nuclear maturation and transcriptome expression profile during oocytes maturation. Therefore, our results elucidate the function of HDAC11 in porcine oocytes maturation and embryos development through regulating α-tubulin acetylation, meiosis-related histone modifications and transcriptome.

Identification of a new orthonairovirus associated with human febrile illness in China.

The genus Orthonairovirus, which is part of the family Nairoviridae, includes the important tick-transmitted pathogens Crimean-Congo hemorrhagic fever virus and Nairobi sheep disease virus, as well as many other poorly characterized viruses found in ticks, birds and mammals1,2. In this study, we identified a new orthonairovirus, Songling virus (SGLV), from patients who reported being bitten by ticks in Heilongjiang Province in northeastern China. SGLV shared similar genomic and morphological features with orthonairoviruses and phylogenetically formed a unique clade in Tamdy orthonairovirus of the Nairoviridae family. The isolated SGLV induced cytopathic effects in human hepatoma cells in vitro. SGLV infection was confirmed in 42 hospitalized patients analyzed between 2017 and 2018, with the main clinical manifestations being headache, fever, depression, fatigue and dizziness. More than two-thirds (69%) of patients generated virus-specific antibody responses in the acute phase. Taken together, these results suggest that this newly discovered orthonairovirus is associated with human febrile illness in China.

Comprehensive Transcriptome Analysis of lncRNAs Reveals the Role of lncAD in Chicken Intramuscular and Abdominal Adipogenesis.

Adipose tissue-specific distribution and deposition speed are the main factors affecting the slaughter performance and meat quality in poultry. Previous studies suggested that different adipose tissues owned various biochemical characteristics and gene expression patterns. To investigate the functional role of long noncoding RNAs (lncRNAs) during chicken intramuscular and abdominal adipogenesis, we performed transcriptome analysis by Ribo-Zero RNA-Seq technology. A total of 11247 lncRNAs were observed in the adipocytes derived from IMF and AbF in chicken. Among them, we got 1624 differentiated expressed novel lncRNAs. A large amount of lncRNAs were involved in several lipid metabolism and adipogenesis-related signaling pathways. Of these, lncRNAs, lncAD is one of the most upregulated lncRNA and was coexpressed with several genes of the PPAR signaling pathway. Here, we report that knockdown of lncAD inhibited its upstream gene TXNRD1 expression in a cis-regulation manner, thus to decrease intramuscular preadipocytes adipogenic differentiation and promoted cell proliferation. Our present study revealed huge lncRNAs profile differences between IMF- and AbF-derived preadipocyte adipogenesis. Collectively, our findings not only provide valuable evidence for the identification of adipogenic lncRNAs but also contribute to further studies about the post-transcriptional regulation mechanism underlying tissue-specific fat deposition in poultry.

Inhibition of HDAC6 by tubastatin A disrupts mouse oocyte meiosis via regulating histone modifications and mRNA expression.

Histone deacetylase 6 (HDAC6) participates in mouse oocyte maturation by deacetylating α-tubulin. However, how HDAC6 expression is regulated in oocytes remains unknown. In the present study, we discovered that mouse oocytes had a high level of HDAC6 expression and a low level of DNA methylation status in their promoter region. Then, a selective HDAC6 inhibitor, tubastatin A (Tub-A) was chosen to investigate the role of HDAC6 in oocyte maturation. Our results revealed that inhibition of HDAC6 caused meiotic progression arrest, disturbed spindle/chromosome organization, and kinetochore-microtubule attachments without impairing spindle assembly checkpoint function. Moreover, inhibition of HDAC6 not only increased the acetylation of α-tubulin but also elevated the acetylation status of H4K16 and decreased the phosphorylation level of H3T3 and H3S10. Conversely, depressed H3T3 phosphorylation by its kinase inhibitor increased the acetylation level of H4K16. Finally, single cell RNA-seq analysis revealed that the cell cycle-related genes CCNB1, CDK2, SMAD3, YWHAZ and the methylation-related genes DNMT1 and DNMT3B were strongly repressed in Tub-A treated oocytes. Taken together, our results indicate that HDAC6 plays important roles in chromosome condensation and kinetochore function via regulating several key histone modifications and messenger RNA transcription during oocyte meiosis.