La phosphoprotéine rabique et ses partenaires viraux et cellulaires (rôle dans l'échappement viral à l'immunité innée et fonction des corps de Negri)

PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF

Rabies Virus P Protein Interacts with STAT1 and Inhibits Interferon Signal Transduction Pathways

Rabies virus P protein is a cofactor of RNA polymerase. We investigated other potential roles of P (CVS strain) by searching for cellular partners using two-hybrid screening. We isolated a cDNA encoding the signal transducer and activator of transcription 1 (STAT1) that is a critical component of interferon type I (IFN-α/β) and type II (IFN-γ) signaling. We confirmed this interaction by glutathione S-transferase-pull-down assay. Deletion mutant analysis indicated that the carboxy-terminal part of P interacted with a region containing the DNA-binding domain and the coiled-coil domain of STAT1. The expression of P protein inhibits IFN-α- and IFN-γ-induced transcriptional responses, thus impairing the IFN-induced antiviral state. Mechanistic studies indicate that P protein does not induce STAT1 degradation and does not interfere with STAT1 phosphorylation but prevents IFN-induced STAT1 nuclear accumulation. These results indicate that rabies P protein overcomes the antiviral response of the infected cells

The Nucleocytoplasmic Rabies Virus P Protein Counteracts Interferon Signaling by Inhibiting both Nuclear Accumulation and DNA Binding of STAT1

Rabies virus P protein inhibits alpha interferon (IFN-α)- and IFN-γ-stimulated Jak-STAT signaling by retaining phosphorylated STAT1 in the cytoplasm. Here, we show that P also blocks an intranuclear step that is the STAT1 binding to the DNA promoter of IFN-responsive genes. As P is a nucleocytoplasmic shuttling protein, we first investigated the effect of the cellular distribution of P on the localization of STAT1 and consequently on IFN signaling. We show that the localization of STAT1 is correlated with the localization of P: in cells expressing a nuclear form of P (the short P3 isoform or the complete P in the presence of the export inhibitor leptomycin B), STAT1 is nuclear, whereas in cells expressing a cytoplasmic form of P, STAT1 is cytoplasmic. However, the expression of nuclear forms of P inhibits the signaling of both IFN-γ and IFN-α, demonstrating that the retention of STAT1 in the cytoplasm is not the only mechanism involved in the inhibition of IFN signaling. Electrophoretic mobility shift analysis indicates that P expression in the cell extracts of infected cells or in stable cell lines prevents IFN-induced DNA binding of STAT1. The loss of the DNA binding of STAT1 and ISGF3 was also observed when purified recombinant P or P3 was added to the extracts of IFN-γ- or IFN-α-treated cells, indicating that P directly affects the DNA binding activity of STAT1. Then products of the rabies virus P gene are able to counteract IFN signaling by creating both cytoplasmic and nuclear blocks for STAT1

Mother-to-Child Transmission of Arboviruses during Breastfeeding: From Epidemiology to Cellular Mechanisms

International audienceMost viruses use several entry sites and modes of transmission to infect their host (parenteral, sexual, respiratory, oro-fecal, transplacental, transcutaneous, etc.). Some of them are known to be essentially transmitted via arthropod bites (mosquitoes, ticks, phlebotomes, sandflies, etc.), and are thus named arthropod-borne viruses, or arboviruses. During the last decades, several arboviruses have emerged or re-emerged in different countries in the form of notable outbreaks, resulting in a growing interest from scientific and medical communities as well as an increase in epidemiological studies. These studies have highlighted the existence of other modes of transmission. Among them, mother-to-child transmission (MTCT) during breastfeeding was highlighted for the vaccine strain of yellow fever virus (YFV) and Zika virus (ZIKV), and suggested for other arboviruses such as Chikungunya virus (CHIKV), dengue virus (DENV), and West Nile virus (WNV). In this review, we summarize all epidemiological and clinical clues that suggest the existence of breastfeeding as a neglected route for MTCT of arboviruses and we decipher some of the mechanisms that chronologically occur during MTCT via breastfeeding by focusing on ZIKV transmission process

Excretion of Cell-Free and Cell-Associated Zika Virus into Breast Milk of Infected Dams and Identification of Antiviral Factors

International audienceZika virus (ZIKV) is a mosquito-borne RNA virus belonging to the Flavivirus genus of the Flaviviridae family. During the 60 years following its discovery in 1947, ZIKV caused little concern for public health as the associated infection was reported as mostly asymptomatic or inducing mild symptoms. However, since 2013, severe neurological symptoms have been associated with ZIKV infection, compelling the World Health Organization to declare a Public Health Emergency of International Concern. Among those symptoms, neurological birth defects may affect children born to mothers infected during pregnancy. Additionally, during the past 8 years, ZIKV transmission through breastfeeding has repeatedly been suggested in epidemiological studies and demonstrated on a mouse model by our team. To better understand the biological factors controlling ZIKV transmission through breastfeeding, we investigated the nature of the viral entities excreted in the breast milk of infected dams and evaluated viral transmission to breastfed pups. We show that both cell-free and cell-associated virus is excreted into breast milk and that ZIKV is efficiently transmitted to the breastfed pups. Additionally, we studied murine breast milk cell types, and identified a majority of mammary luminal cells. Finally, we investigated the effect on ZIKV infectivity of several breast milk components that are antiviral against different viruses such as lactoferrin (LF) and lactalbumin (LA), or free fatty acids (FFA). We showed no effect of LF and LA, whereas FFA inactivated the virus. These results bring new insight concerning the mechanisms of ZIKV transmission during breastfeeding and identify biological factors modulating it. These elements should be considered in risk assessment of ZIKV mother-to-child transmissio

Rabies viral mechanisms to escape the IFN system: the viral protein P interferes with IRF-3, Stat1, and PML nuclear bodies.

Interferons (IFNs) are a family of secreted proteins with antiviral, antiproliferative, and immunomodulatory activities. The different biologic actions of IFN are believed to be mediated by the products of specifically IFN-stimulated genes (ISG) in the target cells. The IFN response is the first line of defense against viral infections. Viruses, which require the cellular machinery for their replication, have evolved different ways to counteract the action of IFN by inhibiting IFN production or Jak-Stat signaling or by altering ISG products. This review focuses on the role of viral proteins from the RNA virus family, particularly rabies P protein. P protein mediates inhibition of the IFN system by different pathways: it inhibits IFN production by impairing IFN regulatory factor-3 (IRF-3) phosphorylation and IFN signaling by blocking nuclear transport of Stat1 and alters promyelocytic leukemia (PML) nuclear bodies by retaining PML in the cytoplasm

Functional characterization of Negri Bodies (NBs) in Rabies Virus-Infected cells : evidence that NBs are sites of viral transcription and replication

International audienceRabies virus infection induces the formation of cytoplasmic inclusion bodies that resemble Negri bodies found in the cytoplasm of some infected nerve cells. We have studied the morphogenesis and the role of these Negri body-like structures (NBLs) during viral infection. The results indicate that these spherical structures (one or two per cell in the initial stage of infection), composed of the viral N and P proteins, grow during the virus cycle before appearing as smaller structures at late stages of infection. We have shown that the microtubule network is not necessary for the formation of these inclusion bodies but is involved in their dynamics. In contrast, the actin network does not play any detectable role in these processes. These inclusion bodies contain Hsp70 and ubiquitinylated proteins, but they are not misfolded protein aggregates. NBLs, in fact, appear to be functional structures involved in the viral life cycle. Specifically, using in situ fluorescent hybridization techniques, we show that all viral RNAs (genome, antigenome, and every mRNA) are located inside the inclusion bodies. Significantly, short-term RNA labeling in the presence of BrUTP strongly suggests that the NBLs are the sites where viral transcription and replication take place

The Influenza Virus Protein PB1-F2 Increases Viral Pathogenesis through Neutrophil Recruitment and NK Cells Inhibition

The influenza A virus (IAV) PB1-F2 protein is a virulence factor contributing to the pathogenesis observed during IAV infections in mammals. In this study, using a mouse model, we compared the host response associated with PB1-F2 with an early transcriptomic signature that was previously associated with neutrophils and consecutively fatal IAV infections. This allowed us to show that PB1-F2 is partly involved in neutrophil-related mechanisms leading to death. Using neutropenic mice, we confirmed that the harmful effect of PB1-F2 is due to an excessive inflammation mediated by an increased neutrophil mobilization. We identified the downstream effects of this PB1-F2-exacerbated neutrophil recruitment. PB1-F2 had no impact on the lymphocyte recruitment in the airways at day 8 pi. However, functional genomics analysis and flow cytometry in broncho-alveolar lavages at 4 days pi revealed that PB1-F2 induced a NK cells deficiency. Thus, our results identify PB1-F2 as an important immune disruptive factor during the IAV infection