Transcriptomic changes in the hypothalamus of mice with chronic migraine: Activation of pathways associated with neuropathic inflammation and central sensitization.

Chronic migraine is a common central nervous system disorder characterized by recurrent, pulsating headaches. However, the extent and mechanisms of hypothalamic involvement in disease progression have not been thoroughly investigated. Herein, we created a chronic migraine mouse model using repeated intraperitoneal injections of nitroglycerin. We performed transcriptomic sequencing on the hypothalamus of mice with chronic migraine and control mice under normal physiological conditions, followed by differential gene set enrichment and functional analysis of the data. Additionally, we examined the intrinsic connection between chronic migraine and sleep disorders using transcriptomic sequencing data from sleep-deprived mice available in public databases. We identified 39 differentially expressed genes (DEGs) in the hypothalamus of a mouse model of chronic migraine. Functional analysis of DEGs revealed enrichment primarily in signaling transduction, immune-inflammatory responses, and the cellular microenvironment. A comparison of the transcriptomic data of sleep-deprived mice revealed two commonly expressed DEGs. Our findings indicate that the hypothalamic DEGs are primarily enriched in the PI3K/AKT/mTOR pathway and associated with the NF-κB/NLRP3/IL-1 ß pathway activation to maintain the central sensitization of the chronic migraine. Chronic migraine-induced gene expression changes in the hypothalamus may help better understand the underlying mechanisms and identify therapeutic targets.

Deletion of MGF-110-9L gene from African swine fever virus weakens autophagic degradation of TBK1 as a mechanism for enhancing type I interferon production.

African swine fever (ASF) caused by African swine fever virus (ASFV) is a devastating disease for the global pig industry and economic benefit. The limited knowledge on the pathogenesis and infection mechanisms of ASF restricts progress toward vaccine development and ASF control. Previously, we illustrated that deletion of the MGF-110-9L gene from highly virulent ASFV CN/GS/2018 strains (ASFV∆9L) results in attenuated virulence in swine, but the underlying mechanism remains unclear. In this study, we found that the difference in virulence between wild-type ASFV (wt-ASFV) and ASFV∆9L strains was mainly caused by the difference in TANK Binding Kinase 1 (TBK1) reduction. TBK1 reduction was further identified to be mediated by the autophagy pathway and this degradative process requires the up-regulation of a positive autophagy regulation molecule- Phosphatidylinositol-4-Phosphate 3-Kinase Catalytic Subunit Type 2 Beta (PIK3C2B). Moreover, TBK1 over-expression was confirmed to inhibit ASFV replication in vitro. In summary, these results indicate that wt-ASFV counteracts type I interferon (IFN) production by degrading TBK1, while ASFVΔ9L enhanced type I IFN production by weakening TBK1 reduction, clarifying the mechanism that ASFVΔ9L present the attenuated virulence in vitro.

African Swine Fever Virus I267L Is a Hemorrhage-Related Gene Based on Transcriptome Analysis.

African swine fever (ASF) is an acute and severe disease transmitted among domestic pigs and wild boars. This disease is notorious for its high mortality rate and has caused great losses to the world's pig industry in the past few years. After infection, pigs can develop symptoms such as high fever, inflammation, and acute hemorrhage, finally leading to death. African swine fever virus (ASFV) is the causal agent of ASF; it is a large DNA virus with 150-200 genes. Elucidating the functions of each gene could provide insightful information for developing prevention and control methods. Herein, to investigate the function of I267L, porcine alveolar macrophages (PAMs) infected with an I267L-deleted ASFV strain (named ∆I267L) and wild-type ASFV for 18 h and 36 h were taken for transcriptome sequencing (RNA-seq). The most distinct different gene that appeared at both 18 hpi (hours post-infection) and 36 hpi was F3; it is the key link between inflammation and coagulation cascades. KEGG analysis (Kyoto encyclopedia of genes and genomes analysis) revealed the complement and coagulation cascades were also significantly affected at 18 hpi. Genes associated with the immune response were also highly enriched with the deletion of I267L. RNA-seq results were validated through RT-qPCR. Further experiments confirmed that ASFV infection could suppress the induction of F3 through TNF-α, while I267L deletion partially impaired this suppression. These results suggest that I267L is a pathogenicity-associated gene that modulates the hemorrhages of ASF by suppressing F3 expression. This study provides new insights into the molecular mechanisms of ASFV pathogenicity and potential targets for ASFV prevention and control.

Functional characterization of African swine fever virus I329L gene by transcriptome analysis.

African swine fever (ASF) is an acute, severe, and highly contagious disease caused by the African swine fever virus (ASFV), which infects domestic pigs and wild boars. The incidence and mortality rates of swine infected with virulent strains of ASFV can reach up to 100%. The large genome, its complex structure, multiple genotypes, and a lack of understanding regarding ASFV gene function are serious obstacles to the development of safe and effective vaccines. Here, ASFV I329L was identified as a relatively conserved gene that is expressed during the late stage of infection. A recombinant virus with I329L gene deletion (ASFV CN/GS/2018-ΔI329L) was produced by replacing I329L with an enhanced green fluorescent protein (EGFP) cassette. In order to explore the function of the ASFV I329L gene, transcriptome sequencing (RNA-seq) was performed on porcine alveolar macrophages (PAMs) infected with ASFV CN/GS/2018 and ASFV CN/GS/2018-ΔI329L. GO functional and KEGG pathway analyses were performed to analyze differentially expressed genes, and different alternative splicing (AS) events were also analyzed. We compared the sequencing data for each sample with the ASFV CN/GS/2018 reference sequence. Interestingly, we found 3 and 1 up-regulated genes and 12 and 19 down-regulated genes at 12 and 24 h post-infection, respectively. In addition, we verified the expression of 5 up-regulated and 5 down-regulated genes by RT-qPCR, and the results were consistent with those obtained based on RNA-seq. In summary, the results obtained in this study provide new insights for further elucidation of ASFV proteins and ASFV-host interactions. These findings will contribute to implementing a comprehensive strategy for controlling the spread of ASF.

African swine fever virus MGF-360-10L is a novel and crucial virulence factor that mediates ubiquitination and degradation of JAK1 by recruiting the E3 ubiquitin ligase HERC5.

African swine fever virus (ASFV) causes acute hemorrhagic infectious disease in pigs. The ASFV genome encodes various proteins that enable the virus to escape innate immunity; however, the underlying mechanisms are poorly understood. The present study found that ASFV MGF-360-10L significantly inhibits interferon (IFN)-ß-triggered STAT1/2 promoter activation and the production of downstream IFN-stimulated genes (ISGs). ASFV MGF-360-10L deletion (ASFV-Δ10L) replication was impaired compared with the parental ASFV CN/GS/2018 strain, and more ISGs were induced by the ASFV-Δ10L in porcine alveolar macrophages in vitro. We found that MGF-360-10L mainly targets JAK1 and mediates its degradation in a dose-dependent manner. Meanwhile, MGF-360-10L also mediates the K48-linked ubiquitination of JAK1 at lysine residues 245 and 269 by recruiting the E3 ubiquitin ligase HERC5 (HECT and RLD domain-containing E3 ubiquitin protein ligase 5). The virulence of ASFV-Δ10L was significantly lower than that of the parental strain in vivo, which indicates that MGF-360-10L is a novel virulence factor of ASFV. Our findings elaborate the novel mechanism of MGF-360-10L on the STAT1/2 signaling pathway, expanding our understanding of the inhibition of host innate immunity by ASFV-encoded proteins and providing novel insights that could contribute to the development of African swine fever vaccines. IMPORTANCE African swine fever outbreaks remain a concern in some areas. There is no effective drug or commercial vaccine to prevent African swine fever virus (ASFV) infection. In the present study, we found that overexpression of MGF-360-10L strongly inhibited the interferon (IFN)-ß-induced STAT1/2 signaling pathway and the production of IFN-stimulated genes (ISGs). Furthermore, we demonstrated that MGF-360-10L mediates the degradation and K48-linked ubiquitination of JAK1 by recruiting the E3 ubiquitin ligase HERC5. The virulence of ASFV with MGF-360-10L deletion was significantly less than parental ASFV CN/GS/2018. Our study identified a new virulence factor and revealed a novel mechanism by which MGF-360-10L inhibits the immune response, thus providing new insights into the vaccination strategies against ASFV.

Research progress on the proteins involved in African swine fever virus infection and replication.

African swine fever (ASF) is an acute, hemorrhagic and highly contagious infectious disease caused by African swine fever virus (ASFV), which infects domestic pigs or wild boars. It is characterized by short course of disease, high fever and hemorrhagic lesions, with mortality of up to 100% from acute infection. Up to now, the lack of commercial vaccines and effective drugs has seriously threatened the healthy economic development of the global pig industry. ASFV is a double-stranded DNA virus and genome varies between about 170-194 kb, which encodes 150-200 viral proteins, including 68 structural proteins and more than 100 non-structural proteins. In recent years, although the research on structure and function of ASFV-encoded proteins has been deepened, the structure and infection process of ASFV are still not clear. This review summarizes the main process of ASFV infection, replication and functions of related viral proteins to provide scientific basis and theoretical basis for ASFV research and vaccine development.