Search Results (18)

The Junín virus (JUNV) is one of the New World arenaviruses that cause severe hemorrhagic fever. Human transferrin receptor 1 (hTfR1) has been identified as the main receptor for JUNV for virus entry into host cells. To date, no treatment has been approved for JUNV. Herein, we investigated 12 anti-hTfR1 VHH (variable domain of the heavy chain of heavy-chain antibody) antibodies and confirmed their interaction with hTfR1. Most of them could bind to the hTfR1 apical domain, which is the glycoprotein 1 (GP1) binding domain of JUNV. Among them, 18N18 exhibited neutralizing activity against both the human immunodeficiency virus (HIV)-vectored lentiviral Junín pseudoviruses and the recombinant vesicular stomatitis virus (VSV)-vectored Junín pseudoviruses. We also verified that 18N18 blocked the interaction between hTfR1 and JUNV GP1. In addition, 18N18 could neutralize another New World arenavirus, the Machupo virus. Using AlphaFold 3-based simulation of 18N18–hTfR1 docking, we determined that 18N18’s binding epitope was located at the JUNV GP1 binding epitope. 18N18 represents a candidate for JUNV treatment and provides a potential approach that could be applied to New World arenaviruses. Full article

Mammalian arenaviruses are rodent-borne zoonotic viruses, some of which can cause fatal hemorrhagic diseases in humans. The first discovered arenavirus, lymphocytic choriomeningitis virus (LCMV), has a worldwide distribution and can be fatal for transplant recipients. However, no FDA-approved drugs or vaccines are currently available. In this study, using a quantitative proteomic analysis, we identified a variety of host factors that could be needed for LCMV infection, among which we found that protein disulfide isomerase A4 (PDIA4), a downstream factor of endoplasmic reticulum stress (ERS), is important for LCMV infection. Biochemical analysis revealed that LCMV glycoprotein was the main viral component accounting for PDIA4 upregulation. The inhibition of ATF6-mediated ERS could prevent the upregulation of PDIA4 that was stimulated by LCMV infection. We further found that PDIA4 can affect the LCMV viral RNA synthesis processes and release. In summary, we conclude that PDIA4 could be a new target for antiviral drugs against LCMV. Full article

The Lassa virus (LASV) causes Lassa fever, a highly infectious and lethal agent of acute viral hemorrhagic fever. At present, there are still no effective treatments available, creating an urgent need to develop novel therapeutics. Some benzimidazole compounds targeting the arenavirus envelope glycoprotein complex (GPC) are promising inhibitors of LASV. In this study, we synthesized two series of LASV inhibitors based on the benzimidazole structure. Lentiviral pseudotypes bearing the LASV GPC were established to identify virus entry inhibitors. Surface plasmon resonance (SPR) was further used to verify the binding activities of the potential compounds. Compounds 7d−Z, 7h−Z, 13c, 13d, and 13f showed relatively excellent antiviral activities with IC₅₀ values ranging from 7.58 to 15.46 nM and their SI values above 1251. These five representative compounds exhibited stronger binding affinity with low equilibrium dissociation constants (K_D < 8.25 × 10⁻⁷ M) in SPR study. The compound 7h−Z displayed the most potent antiviral activity (IC₅₀ = 7.58 nM) with a relatively high SI value (2496), which could be further studied as a lead compound. The structure–activity relationship indicated that the compounds with lipophilic and spatially larger substituents might possess higher antiviral activity and a much larger safety margin. This study will provide some good guidance for the development of highly active compounds with a novel skeleton against LASV. Full article

Human T-cell immunoglobulin mucin 1 (hTIM-1) is known to promote cellular entry of enveloped viruses. Previous studies suggested that the polymorphisms of hTIM-1 affected its function. Here, we analyzed single nucleotide variants (SNVs) of hTIM-1 to determine their ability to promote cellular entry of viruses using pseudotyped vesicular stomatitis Indiana virus (VSIV). We obtained hTIM-1 sequences from a public database (Ensembl genome browser) and identified 35 missense SNVs in 3 loops of the hTIM-1 immunoglobulin variable (IgV) domain, which had been reported to interact with the Ebola virus glycoprotein (GP) and phosphatidylserine (PS) in the viral envelope. HEK293T cells transiently expressing wildtype hTIM-1 or its SNV mutants were infected with VSIVs pseudotyped with filovirus or arenavirus GPs, and their infectivities were compared. Eleven of the thirty-five SNV substitutions reduced the efficiency of hTIM-1-mediated entry of pseudotyped VSIVs. These SNV substitutions were found not only around the PS-binding pocket but also in other regions of the molecule. Taken together, our findings suggest that some SNVs of the hTIM-1 IgV domain have impaired ability to interact with PS and/or viral GPs in the viral envelope, which may affect the hTIM-1 function to promote viral entry into cells. Full article

Replication-competent reporter-expressing viruses are crucial tools in molecular virology with applications that range from antiviral screening to live-cell imaging of protein spatiotemporal dynamics. However, there is currently little information available regarding viable strategies to develop reporter-expressing arenaviruses. To address this, we used Tacaribe virus (TCRV), an apathogenic BSL2 arenavirus, to assess the feasibility of different reporter expression approaches. We first generated trisegmented TCRV viruses with either the glycoprotein (GP) or nucleoprotein (NP) replaced by a reporter (GFP, mCherry, or nanoluciferase). These viruses were all viable, but showed marked differences in brightness and attenuation. Next, we generated terminal fusions with each of the TCRV proteins (i.e., NP, GP, polymerase (L), matrix protein (Z)) either with or without a T2A self-cleavage site. We tested both the function of the reporter-fused proteins alone, and the viability of corresponding recombinant TCRVs. We successfully rescued viruses with both direct and cleavable reporter fusions at the C-terminus of Z, as well as cleavable N-terminal fusions with NP. These viruses all displayed detectable reporter activity, but were also moderately attenuated. Finally, reporter proteins were inserted into a flexible hinge region within L. These viruses were also viable and showed moderate attenuation; however, reporter expression was only detectable for the luminescent virus. These strategies provide an exciting range of new tools for research into the molecular biology of TCRV that can likely also be adapted to other arenaviruses. Full article

New World arenaviruses are rodent-transmitted viruses and include a number of pathogens that are responsible for causing severe human disease. This includes Junín virus (JUNV), which is the causative agent of Argentine hemorrhagic fever. The wild nature and mobility of the rodent reservoir host makes it difficult to control the disease, and currently passive immunization with high-titer neutralizing antibody-containing plasma from convalescent patients is the only specific therapy. However, dwindling supplies of naturally available convalescent plasma, and challenges in developing similar resources for other closely related viruses, have made the development of alternative antibody-based therapeutic approaches of critical importance. In this study, we sought to induce a neutralizing antibody response in rabbits against the receptor-binding subunit of the viral glycoprotein, GP1, and the specific peptide sequences in GP1 involved in cellular receptor contacts. While these specific receptor-interacting peptides did not efficiently induce the production of neutralizing antibodies when delivered as a particulate antigen (as part of hepatitis B virus core-like particles), we showed that recombinant JUNV GP1 purified from transfected mammalian cells induced virus-neutralizing antibodies at high titers in rabbits. Further, neutralization was observed across a range of unrelated JUNV strains, a feature that is critical for effectiveness in the field. These results underscore the potential of GP1 alone to induce a potent neutralizing antibody response and highlight the importance of epitope presentation. In addition, effective virus neutralization by rabbit antibodies supports the potential applicability of this species for the future development of immunotherapeutics (e.g., based on humanized monoclonal antibodies). Such information can be applied in the design of vaccines and immunogens for both prevention and specific therapies against this and likely also other closely related pathogenic New World arenaviruses. Full article

Lassa virus (LASV)—a member of the family Arenaviridae—causes Lassa fever in humans and is endemic in West Africa. Currently, no approved drugs are available. We screened 2480 small compounds for their potential antiviral activity using pseudotyped vesicular stomatitis virus harboring the LASV glycoprotein (VSV-LASVGP) and a related prototypic arenavirus, lymphocytic choriomeningitis virus (LCMV). Follow-up studies confirmed that CP100356 hydrochloride (CP100356), a specific P-glycoprotein (P-gp) inhibitor, suppressed VSV-LASVGP, LCMV, and LASV infection with half maximal inhibitory concentrations of 0.52, 0.54, and 0.062 μM, respectively, without significant cytotoxicity. Although CP100356 did not block receptor binding at the cell surface, it inhibited low-pH-dependent membrane fusion mediated by arenavirus glycoproteins. P-gp downregulation did not cause a significant reduction in either VSV-LASVGP or LCMV infection, suggesting that P-gp itself is unlikely to be involved in arenavirus entry. Finally, our data also indicate that CP100356 inhibits the infection by other mammarenaviruses. Thus, our findings suggest that CP100356 can be considered as an effective virus entry inhibitor for LASV and other highly pathogenic mammarenaviruses. Full article

Host plasma membrane protein SERINC5 is incorporated into budding retrovirus particles where it blocks subsequent entry into susceptible target cells. Three structurally unrelated proteins encoded by diverse retroviruses, human immunodeficiency virus type 1 (HIV-1) Nef, equine infectious anemia virus (EIAV) S2, and ecotropic murine leukemia virus (MLV) GlycoGag, disrupt SERINC5 antiviral activity by redirecting SERINC5 from the site of virion assembly on the plasma membrane to an internal RAB7+ endosomal compartment. Pseudotyping retroviruses with particular glycoproteins, e.g., vesicular stomatitis virus glycoprotein (VSV G), renders the infectivity of particles resistant to inhibition by virion-associated SERINC5. To better understand viral determinants for SERINC5-sensitivity, the effect of SERINC5 was assessed using HIV-1, MLV, and Mason-Pfizer monkey virus (M-PMV) virion cores, pseudotyped with glycoproteins from Arenavirus, Coronavirus, Filovirus, Rhabdovirus, Paramyxovirus, and Orthomyxovirus genera. SERINC5 restricted virions pseudotyped with glycoproteins from several retroviruses, an orthomyxovirus, a rhabdovirus, a paramyxovirus, and an arenavirus. Infectivity of particles pseudotyped with HIV-1, amphotropic-MLV (A-MLV), or influenza A virus (IAV) glycoproteins, was decreased by SERINC5, whether the core was provided by HIV-1, MLV, or M-PMV. In contrast, particles pseudotyped with glycoproteins from M-PMV, parainfluenza virus 5 (PIV5), or rabies virus (RABV) were sensitive to SERINC5, but only with particular retroviral cores. Resistance to SERINC5 did not correlate with reduced SERINC5 incorporation into particles, route of viral entry, or absolute infectivity of the pseudotyped virions. These findings indicate that some non-retroviruses may be sensitive to SERINC5 and that, in addition to the viral glycoprotein, the retroviral core influences sensitivity to SERINC5. Full article

Hemorrhagic fever viruses, among them orthohantaviruses, arenaviruses and filoviruses, are responsible for some of the most severe human diseases and represent a serious challenge for public health. The current limited therapeutic options and available vaccines make the development of novel efficacious antiviral agents an urgent need. Inhibiting viral attachment and entry is a promising strategy for the development of new treatments and to prevent all subsequent steps in virus infection. Here, we developed a fluorescence-based screening assay for the identification of new antivirals against hemorrhagic fever virus entry. We screened a phytochemical library containing 320 natural compounds using a validated VSV pseudotype platform bearing the glycoprotein of the virus of interest and encoding enhanced green fluorescent protein (EGFP). EGFP expression allows the quantitative detection of infection and the identification of compounds affecting viral entry. We identified several hits against four pseudoviruses for the orthohantaviruses Hantaan (HTNV) and Andes (ANDV), the filovirus Ebola (EBOV) and the arenavirus Lassa (LASV). Two selected inhibitors, emetine dihydrochloride and tetrandrine, were validated with infectious pathogenic HTNV in a BSL-3 laboratory. This study provides potential therapeutics against emerging virus infection, and highlights the importance of drug repurposing. Full article

Lassa virus (LASV) is a rodent-borne arenavirus circulating in West African regions that causes Lassa fever (LF). LF is normally asymptomatic at the initial infection stage, but can progress to severe disease with multiorgan collapse and hemorrhagic fever. To date, the therapeutic choices are limited, and there is no approved vaccine for avoiding LASV infection. Adenoviral vector-based vaccines represent an effective countermeasure against LASV because of their safety and adequate immunogenicity, as demonstrated in use against other emerging viral infections. Here, we constructed and characterized a novel Ad5 (E1-, E3-) vectored vaccine containing the glycoprotein precursor (GPC) of LASV. Ad5-GPC_LASV elicited both humoral and cellular immune responses in BALB/c mice. Moreover, a bioluminescent imaging-based BALB/c mouse model infected with GPC-bearing and luciferase-expressing replication-incompetent LASV pseudovirus was utilized to evaluate the vaccine efficacy. The bioluminescence intensity of immunized mice was significantly lower than that of control mice after being inoculated with LASV pseudovirus. This study suggests that Ad5-GPC_LASV represents a potential vaccine candidate against LF. Full article

Lassa virus (LASV), an arenavirus causing Lassa fever, is endemic to West Africa with up to 300,000 cases and between 5000 and 10,000 deaths per year. Rarely seen in the United States, Lassa virus is a CDC category A biological agent inasmuch deliberate aerosol exposure can have high mortality rates compared to naturally acquired infection. With the need for an animal model, specific countermeasures remain elusive as there is no FDA-approved vaccine. This natural history of aerosolized Lassa virus exposure in Macaca fascicularis was studied under continuous telemetric surveillance. The macaque response to challenge was largely analogous to severe human disease with fever, tachycardia, hypotension, and tachypnea. During initial observations, an increase trend of activated monocytes positive for viral glycoprotein was accompanied by lymphocytopenia. Disease uniformly progressed to high viremia followed by low anion gap, alkalosis, anemia, and thrombocytopenia. Hypoproteinemia occurred late in infection followed by increased levels of white blood cells, cytokines, chemokines, and biochemical markers of liver injury. Viral nucleic acids were detected in tissues of three non-survivors at endpoint, but not in the lone survivor. This study provides useful details to benchmark a pivotal model of Lassa fever in support of medical countermeasure development for both endemic disease and traditional biodefense purposes. Full article

A metatranscriptomic study of RNA viruses in cold-blooded vertebrates identified two related viruses from frogfish (Antennarius striatus) that represent a new genus Antennavirus in the family Arenaviridae (Order: Bunyavirales). Computational analyses were used to identify features common to class I viral fusion proteins (VFPs) in antennavirus glycoproteins, including an N-terminal fusion peptide, two extended alpha-helices, an intrahelical loop, and a carboxyl terminal transmembrane domain. Like mammarenavirus and hartmanivirus glycoproteins, the antennavirus glycoproteins have an intracellular zinc-binding domain and a long virion-associated stable signal peptide (SSP). The glycoproteins of reptarenaviruses are also class I VFPs, but do not contain zinc-binding domains nor do they encode SSPs. Divergent evolution from a common progenitor potentially explains similarities of antennavirus, mammarenavirus, and hartmanivirus glycoproteins, with an ancient recombination event resulting in a divergent reptarenavirus glycoprotein. Full article

Lassa virus (LASV) is an Old World arenavirus, endemic to West Africa, capable of causing hemorrhagic fever. Currently, there are no approved vaccines or effective antivirals for LASV. However, thorough understanding of the LASV glycoprotein and entry into host cells could accelerate therapeutic design. LASV entry is a two-step process involving the viral glycoprotein (GP). First, the GP subunit 1 (GP1) binds to the cell surface receptor and the viral particle is engulfed into an endosome. Next, the drop in pH triggers GP rearrangements, which ultimately leads to the GP subunit 2 (GP2) forming a six-helix-bundle (6HB). The process of GP2 forming 6HB fuses the lysosomal membrane with the LASV envelope, allowing the LASV genome to enter the host cell. The aim of this study was to identify residues in GP2 that are crucial for LASV entry. To achieve this, we performed alanine scanning mutagenesis on GP2 residues. We tested these mutant GPs for efficient GP1-GP2 cleavage, cell-to-cell membrane fusion, and transduction into cells expressing α-dystroglycan and secondary LASV receptors. In total, we identified seven GP2 mutants that were cleaved efficiently but were unable to effectively transduce cells: GP-L280A, GP-L285A/I286A, GP-I323A, GP-L394A, GP-I403A, GP-L415A, and GP-R422A. Therefore, the data suggest these residues are critical for GP2 function in LASV entry. Full article

Arenaviruses are a family of enveloped negative-stranded RNA viruses that can cause severe human disease ranging from encephalitis symptoms to fulminant hemorrhagic fever. The bi‑segmented RNA genome encodes four polypeptides: the nucleoprotein NP, the surface glycoprotein GP, the polymerase L, and the RING finger protein Z. Although it is the smallest arenavirus protein with a length of 90 to 99 amino acids and a molecular weight of approx. 11 kDa, the Z protein has multiple functions in the viral life cycle including (i) regulation of viral RNA synthesis, (ii) orchestration of viral assembly and budding, (iii) interaction with host cell proteins, and (iv) interferon antagonism. In this review, we summarize our current understanding of the structural and functional role of the Z protein in the arenavirus replication cycle. Full article

Arenaviruses include lethal human pathogens which pose serious public health threats. So far, no FDA approved vaccines are available against arenavirus infections, and therapeutic options are limited, making the identification of novel drug targets for the development of efficacious therapeutics an urgent need. Arenaviruses are comprised of two RNA genome segments and four proteins, the polymerase L, the envelope glycoprotein GP, the matrix protein Z, and the nucleoprotein NP. A crucial step in the arenavirus life-cycle is the biosynthesis and maturation of the GP precursor (GPC) by cellular signal peptidases and the cellular enzyme Subtilisin Kexin Isozyme-1 (SKI-1)/Site-1 Protease (S1P) yielding a tripartite mature GP complex formed by GP1/GP2 and a stable signal peptide (SSP). GPC cleavage by SKI-1/S1P is crucial for fusion competence and incorporation of mature GP into nascent budding virion particles. In a first part of our review, we cover basic aspects and newer developments in the biosynthesis of arenavirus GP and its molecular interaction with SKI-1/S1P. A second part will then highlight the potential of SKI-1/S1P-mediated processing of arenavirus GPC as a novel target for therapeutic intervention to combat human pathogenic arenaviruses. Full article