267 research outputs found
Compellingly high SARS-CoV-2 susceptibility of Golden Syrian hamsters suggests multiple zoonotic infections of pet hamsters during the COVID-19 pandemic
Golden Syrian hamsters (Mesocricetus auratus) are used as a research model for severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2). Millions of Golden Syrian hamsters are also kept as pets in close contact to humans. To determine the minimum infective dose (MID) for assessing the zoonotic transmission risk, and to define the optimal infection dose for experimental studies, we orotracheally inoculated hamsters with SARS-CoV-2 doses from 1 * 105 to 1 * 10-4 tissue culture infectious dose 50 (TCID50). Body weight and virus shedding were monitored daily. 1 * 10-3 TCID50 was defined as the MID, and this was still sufficient to induce virus shedding at levels up to 102.75 TCID50/ml, equaling the estimated MID for humans. Virological and histological data revealed 1 * 102 TCID50 as the optimal dose for experimental infections. This compelling high susceptibility leading to productive infections in Golden Syrian hamsters must be considered as a potential source of SARS-CoV-2 infection for humans that come into close contact with pet hamsters
The RING-CH ligase K5 antagonizes restriction of KSHV and HIV-1 particle release by mediating ubiquitin-dependent endosomal degradation of tetherin
Tetherin (CD317/BST2) is an interferon-induced membrane protein that inhibits the release of diverse enveloped viral particles. Several mammalian viruses have evolved countermeasures that inactivate tetherin, with the prototype being the HIV-1 Vpu protein. Here we show that the human herpesvirus Kaposi's sarcoma-associated herpesvirus (KSHV) is sensitive to tetherin restriction and its activity is counteracted by the KSHV encoded RING-CH E3 ubiquitin ligase K5. Tetherin expression in KSHV-infected cells inhibits viral particle release, as does depletion of K5 protein using RNA interference. K5 induces a species-specific downregulation of human tetherin from the cell surface followed by its endosomal degradation. We show that K5 targets a single lysine (K18) in the cytoplasmic tail of tetherin for ubiquitination, leading to relocalization of tetherin to CD63-positive endosomal compartments. Tetherin degradation is dependent on ESCRT-mediated endosomal sorting, but does not require a tyrosine-based sorting signal in the tetherin cytoplasmic tail. Importantly, we also show that the ability of K5 to substitute for Vpu in HIV-1 release is entirely dependent on K18 and the RING-CH domain of K5. By contrast, while Vpu induces ubiquitination of tetherin cytoplasmic tail lysine residues, mutation of these positions has no effect on its antagonism of tetherin function, and residual tetherin is associated with the trans-Golgi network (TGN) in Vpu-expressing cells. Taken together our results demonstrate that K5 is a mechanistically distinct viral countermeasure to tetherin-mediated restriction, and that herpesvirus particle release is sensitive to this mode of antiviral inhibition
Kaposi's Sarcoma-Associated Herpesvirus ORF45 Interacts with Kinesin-2 Transporting Viral Capsid-Tegument Complexes along Microtubules
Open reading frame (ORF) 45 of Kaposi's sarcoma-associated herpesvirus (KSHV) is a tegument protein. A genetic analysis with a null mutant suggested a possible role for this protein in the events leading to viral egress. In this study, ORF45 was found to interact with KIF3A, a kinesin-2 motor protein that transports cargoes along microtubules to cell periphery in a yeast two-hybrid screen. The association was confirmed by both co-immunoprecipitation and immunoflorescence approaches in primary effusion lymphoma cells following virus reactivation. ORF45 principally mediated the docking of entire viral capsid-tegument complexes onto the cargo-binding domain of KIF3A. Microtubules served as the major highways for transportation of these complexes as evidenced by drastically reduced viral titers upon treatment of cells with a microtubule depolymerizer, nocodazole. Confocal microscopic images further revealed close association of viral particles with microtubules. Inhibition of KIF3AβORF45 interaction either by the use of a headless dominant negative (DN) mutant of KIF3A or through shRNA-mediated silencing of endogenous KIF3A expression noticeably decreased KSHV egress reflecting as appreciable reductions in the release of extracellular virions. Both these approaches, however, failed to impact HSV-1 egress, demonstrating the specificity of KIF3A in KSHV transportation. This study thus reports on transportation of KSHV viral complexes on microtubules by KIF3A, a kinesin motor thus far not implicated in virus transportation. All these findings shed light on the understudied but significant events in the KSHV life cycle, delineating a crucial role of a KSHV tegument protein in cellular transport of viral particles
A 3D Model of the Membrane Protein Complex Formed by the White Spot Syndrome Virus Structural Proteins
Outbreaks of white spot disease have had a large negative economic impact on cultured shrimp worldwide. However, the pathogenesis of the causative virus, WSSV (whit spot syndrome virus), is not yet well understood. WSSV is a large enveloped virus. The WSSV virion has three structural layers surrounding its core DNA: an outer envelope, a tegument and a nucleocapsid. In this study, we investigated the protein-protein interactions of the major WSSV structural proteins, including several envelope and tegument proteins that are known to be involved in the infection process.In the present report, we used coimmunoprecipitation and yeast two-hybrid assays to elucidate and/or confirm all the interactions that occur among the WSSV structural (envelope and tegument) proteins VP51A, VP19, VP24, VP26 and VP28. We found that VP51A interacted directly not only with VP26 but also with VP19 and VP24. VP51A, VP19 and VP24 were also shown to have an affinity for self-interaction. Chemical cross-linking assays showed that these three self-interacting proteins could occur as dimers.From our present results in conjunction with other previously established interactions we construct a 3D model in which VP24 acts as a core protein that directly associates with VP26, VP28, VP38A, VP51A and WSV010 to form a membrane-associated protein complex. VP19 and VP37 are attached to this complex via association with VP51A and VP28, respectively. Through the VP26-VP51C interaction this envelope complex is anchored to the nucleocapsid, which is made of layers of rings formed by VP664. A 3D model of the nucleocapsid and the surrounding outer membrane is presented
Targeting of Pseudorabies Virus Structural Proteins to Axons Requires Association of the Viral Us9 Protein with Lipid Rafts
The pseudorabies virus (PRV) Us9 protein plays a central role in targeting viral capsids and glycoproteins to axons of dissociated sympathetic neurons. As a result, Us9 null mutants are defective in anterograde transmission of infection in vivo. However, it is unclear how Us9 promotes axonal sorting of so many viral proteins. It is known that the glycoproteins gB, gC, gD and gE are associated with lipid raft microdomains on the surface of infected swine kidney cells and monocytes, and are directed into the axon in a Us9-dependent manner. In this report, we determined that Us9 is associated with lipid rafts, and that this association is critical to Us9-mediated sorting of viral structural proteins. We used infected non-polarized and polarized PC12 cells, a rat pheochromocytoma cell line that acquires many of the characteristics of sympathetic neurons in the presence of nerve growth factor (NGF). In these cells, Us9 is highly enriched in detergent-resistant membranes (DRMs). Moreover, reducing the affinity of Us9 for lipid rafts inhibited anterograde transmission of infection from sympathetic neurons to epithelial cells in vitro. We conclude that association of Us9 with lipid rafts is key for efficient targeting of structural proteins to axons and, as a consequence, for directional spread of PRV from pre-synaptic to post-synaptic neurons and cells of the mammalian nervous system
A Promiscuous DNA Packaging Machine from Bacteriophage T4
Packaged viral genome can be removed from bacteriophage T4 capsid and the capsid refilled with any double-stranded DNA, single or multiple molecules, using a powerful ATP-fueled DNA packaging machine
Phylogenetic analyses of highly pathogenic avian influenza virus isolates from Germany in 2006 and 2007 suggest at least three separate introductions of H5N1 virus
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