Biology subjects

Witwit, H.

Publications and source records attributed to Witwit, H..

9 recordsLinked to original sources

bioRxiv2026

Sirtuin 1 Is Required for Optimal Mammarenavirus Multiplication

Mammarenaviruses (MaAv) cause persistent infections in diverse rodent reservoirs worldwide and several are zoonotic pathogens with an important public-health burden in their endemic regions. Moreover, the globally distributed MaAv lymphocytic choriomeningitis virus (LCMV) is an underrecognized pathogen of clinical significance in congenital infections and immunocompromised individuals. The lack of FDA-approved vaccines or antivirals for MaAv infections underscores the urgent need for novel anti-MaAv therapeutic strategies. Neutral sphingomyelinase 2 (nSMase2) was recently identified as a host factor contributing to LCMV multiplication, and its inhibitor cambinol exhibits dose-dependent antiviral activity against LCMV but the underlying mechanisms remain undefined. Here, we show that cambinol disrupts multiple stages of the LCMV life cycle. Cambinol inhibits the pH-dependent fusion event mediated by MaAv glycoprotein, a step required for completion of virus cell entry. It also reduces viral ribonucleoprotein (vRNP)-directed genome replication and transcription and impairs the budding activity of the virus matrix Z protein. Cambinol also inhibits sirtuins 1 and 2 (Sirt-1 and Sirt-2), two NAD+-dependent protein deacetylases with pleiotropic roles in cellular metabolism and stress responses, raising the question of whether cambinol anti-LCMV activity reflects nSMase2 inhibition alone or also involves sirtuin-dependent pathways. LCMV multiplication was significantly reduced in SIRT1, but not SIRT2, knockout (KO) cells, uncovering a pro-viral role for Sirt-1 in the LCMV life cycle. Consistent with this finding, LCMV vRNP activity and production of infectious progeny were reduced in SIRT1 KO cells. These findings identify Sirt-1 as a host factor required for optimal LCMV multiplication. Sirt-1 inhibitors are in clinical development for oncological and neurological indications, raising the possibility of repurposing Sirt-1 inhibitors as host-directed antivirals (HDAs) against human pathogenic MaAv. Abstract figure O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/740332v1_ufig1.gif" ALT="Figure 1"> View larger version (35K): org.highwire.dtl.DTLVardef@d53c5forg.highwire.dtl.DTLVardef@16ea861org.highwire.dtl.DTLVardef@1f09addorg.highwire.dtl.DTLVardef@1472e6b_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2026

Mammarenavirus-Induced Remodeling of the Cellular Lipid Landscape Reveals Sphingolipid Metabolism as a Novel Target for Antiviral Intervention

Several mammarenaviruses (MaAv) cause severe and often life-threatening disease in humans and represent major public health threats in their endemic regions. Lassa (LASV) and Junin (JUNV) MaAv, endemic to Western Africa and the Argentine Pampas, respectively, are etiologic agents of viral hemorrhagic fevers associated with high morbidity and mortality. In addition, the globally distributed MaAv lymphocytic choriomeningitis virus (LCMV) is an underrecognized human pathogen capable of causing severe congenital disease and fatal infections in immunocompromised individuals. Despite their public health importance, no FDA-approved vaccines or virus-specific antiviral therapies exist to prevent and treat human MaAv infections. Current treatment relies on the off-label use of ribavirin whose therapeutic efficacy remains controversial. These findings underscore the urgent need to develop effective antiviral strategies against human pathogenic MaAv. Here, we investigated the impact of LCMV infection on host lipid metabolism using an integrated transcriptomic and lipidomic approach. Our data reveal extensive time-dependent remodeling of the cellular lipid landscape, with particularly prominent alterations in sphingolipid and fatty acid metabolic pathways. Functional interrogation of these pathways using pharmacological inhibitors identified acetyl-CoA carboxylase (ACC) and neutral sphingomyelinase 2 (nSMase2) as host factors contributing to efficient viral replication. Notably, inhibition of nSMase2 reduced infectious virus production by 2 logs of infectious virus. Our findings showed that LCMV reprograms host lipid metabolism to facilitate infection and identified sphingolipid turnover as a promising target for host-directed antiviral strategies against MaAv infections.

microbiology↗

bioRxiv2026

Prolyl tRNA Synthetase Is Required for Mammarenavirus Multiplication

Several mammarenaviruses (MaAv), chiefly Lassa virus (LASV) in Western Africa and Junin virus (JUNV) in the Argentinean Pampas, cause severe disease in humans and pose important public health problems in their endemic regions. In addition, the globally distributed MaAv lymphocytic choriomeningitis virus (LCMV) is an underrecognized human pathogen of clinical significance especially in congenital infections and LCMV poses a serious risk for immunocompromised individuals. There are no FDA-approved MaAv vaccines or antivirals and current anti-MaAv therapy is limited to an off-label use of ribavirin whose efficacy remains controversial. This highlights an urgent unmet need for developing antivirals against human pathogenic MaAv. Halofuginone (HF), a derivative of the natural alkaloid febrifugine, has been shown to exhibit antiviral activity against several RNA viruses. Here, we present evidence that HF exhibits a potent dose-dependent antiviral activity against LCMV, and the hemorrhagic fever causing MaAv LASV and JUNV. HF binds to the bifunctional enzyme glutamyl-prolyl-tRNA synthetase 1 (EPRS1) and specifically inhibits its prolyl-tRNA synthetase (PRS) activity, resulting in translation inhibition via the amino acid starvation (AAS) response with preferential impact on proline-rich proteins. HF anti-LCMV activity was prevented by the addition of exogenous proline supporting that inhibition of PRS activity plays a critical role on the anti-MaAv activity of HF. We found that HF did not affect LCMV cell entry, modestly (twofold) reduced the activity of the virus ribonucleoprotein (vRNP) but strongly inhibited (>90%) Z budding activity, a process involving the Z proline-rich late domain motifs.

microbiology↗

bioRxiv2025

Pandemic Response Box Screening Identified CRM1/XPO1 as an Anti-Mammarenavirus Druggable Target

Mammarenaviruses (MaAv) cause persistent infection in their natural rodent hosts across the world and via zoonotic events can cause severe disease in humans. Thus, the MaAv Lassa virus (LASV) in Western Africa and Junin virus (JUNV) in the Argentinean Pampas cause hemorrhagic fever diseases with significant case fatality rates in their endemic regions. In addition, the globally distributed MaAv lymphocytic choriomeningitis virus (LCMV) is an underrecognized human pathogen of clinical significance capable of causing devastating infections in neonates and immunocompromised individuals. Despite their impact on human health, there are currently no FDA-approved vaccines or specific antiviral treatments for MaAv infections. Existing anti-MaAv therapies are limited to the off-label use of ribavirin whose efficacy remains controversial, hence, the significance of developing novel therapeutics to combat human pathogenic MaAv. We employed a high-throughput cell-based infection assay to screen the Pandemic Response Box, a collection of 400 diverse compounds with established antimicrobial activity, for MaAv inhibitors. We identified Ro-24-7429, an antagonist of the HIV-1 Tat protein and RUNX Family Transcription Factor 1 inhibitor; WO 2006118607 A2, a dihydroorotate dehydrogenase inhibitor; and verdinexor, a novel selective inhibitor of nuclear export (SINE) targeting the CRM1/XPO1, as potent anti-MaAv compounds. Consistent with their distinct validated targets, verdinexor and WO 2006118607 A2 exhibited a very strong synergistic antiviral activity when used in combination therapy. Our findings pave the way for the development of verdinexor as a potent host-directed antiviral against MaAv, which could be integrated into the development of combination therapy with direct-or host-acting antivirals to combat human pathogenic MaAv.

microbiology↗

bioRxiv2025

N-myristoyltransferase Inhibitors as Candidate Broad-Spectrum Antivirals to Treat Viral Infections Promoted by Immunosuppression Associated with JAK Inhibitors Therapy

The use of Janus kinase (JAK) inhibitors in the clinic has been expanded significantly during the last decade. However, the immunosuppressive effects of JAK inhibitors, via modulation of key innate cellular signaling pathways, can predispose treated patients to infections, and can also result in reduced control of silent infections and increased risk of reactivation of opportunistic infections. Thus, the JAK inhibitor ruxolitinib, approved for the treatment of myelofibrosis and polycythemia vera, has been shown to exerts a proviral activity during infection with different viruses. Therefore, the clinical relevance of developing antiviral treatments that can be effective in the presence of JAK inhibitors. N-terminal myristoyl transferase (NMT) inhibitors have been shown to exhibit potent antiviral activity against different viruses. Here we document that in the presence of ruxolitinib, NMT inhibitors retain their potent antiviral activity against different viruses, including HSV-1. Our findings support that NMT inhibitors should be explored as therapeutics to treat viral infections associated with immunosuppression caused by treatments with JAK inhibitors.

microbiology↗

bioRxiv2025

Mammarenavirus Z protein myristoylation and oligomerization are not required for its dose-dependent inhibitory effect on vRNP activity

We have recently documented the use of N-Myristoyltransferase inhibitors (NMTi) as an antiviral strategy against human pathogenic mammarenaviruses, including Lassa and Junin viruses, responsible for the hemorrhagic fever (HF) diseases Lassa fever (LF) and Argentine HF (AHF), respectively. Mammarenavirus Z matrix protein has been shown to exert a dose-dependent inhibitory effect in the activity of the virus ribonucleoprotein (vRNP) complex responsible for directing replication and transcription of the viral genome. Prevention of Z myristoylation by NMTi targeted Z protein for degradation, which resulted in inhibition of virus multiplication. Here, we review the recent findings of the use NMTi as antiviral, and reconcile the mechanistic process behind this inhibitory effect, more importantly, we used NMTi and G2A-mutated Z protein as a control of myristoylation function of wild type Z protein to elucidate that the vRNP suppression is monomeric-dependent activity of Z protein abundancy and it is independent of its myristoylation function.

microbiology↗

bioRxiv2024

Cellular N-myristoyl transferases Are Required for Mammarenavirus Multiplication

The mammarenavirus matrix Z protein plays critical roles in virus assembly and cell egress, whereas heterotrimer complexes of a stable signal peptide (SSP) together with glycoprotein subunits GP1 and GP2, generated via co-and post-translational processing of the surface glycoprotein precursor GPC, form the spikes that decorate the virion surface and mediate virus cell entry via receptor-mediated endocytosis. The Z protein and SSP undergo N-terminal myristoylation by host cell N-myristoyltransferases (NMT1 and NMT2), and G2A mutations that prevent myristoylation of Z or SSP have been shown to affect Z mediated virus budding and GP2 mediated fusion activity required to complete the virus cell entry process. In the present work, we present evidence that the validated on-target specific pan NMT inhibitor DDD85464 exerts a potent antiviral activity against the prototypic mammarenavirus lymphocytic choriomeningitis virus (LCMV) that correlated with reduced Z budding activity and GP2 mediated fusion activity, as well as proteasome mediated degradation of the Z protein. The potent anti-mammarenaviral activity of DDD85646 was also observed with the hemorrhagic fever causing mammarenaviruses Junin (JUNV) and Lassa (LASV) viruses. Our results support exploration of NMT inhibition as a broad-spectrum antiviral against human pathogenic mammarenaviruses.

microbiology↗

bioRxiv2023

Activation of Protein Kinase R (PKR) Plays a Pro-Viral Role in Mammarenavirus Infected Cells

Many viruses, including mammarenaviruses, have evolved mechanisms to counteract different components of the host cell innate immunity, which is required to facilitate robust virus multiplication. The double strand (ds)RNA sensor protein kinase receptor (PKR) pathway plays a critical role in the cell antiviral response. Whether PKR can restrict the multiplication of the Old World mammarenavirus lymphocytic choriomeningitis virus (LCMV) and the mechanisms by which LCMV may counteract the antiviral functions of PKR have not yet been investigated. Here we present evidence that LCMV infection results in very limited levels of PKR activation, but LCMV multiplication is enhanced in the absence of PKR. In contrast, infection with a recombinant LCMV with a mutation affecting the 3-5 exonuclease (ExoN) activity of the viral nucleoprotein (NP) resulted in robust PKR activation in the absence of detectable levels of dsRNA, which was associated with severely restricted virus multiplication that was alleviated in the absence of PKR. However, pharmacological inhibition of PKR activation resulted in reduced levels of LCMV multiplication. These findings uncovered a complex role of the PKR pathway in LCMV-infected cells involving both pro-and anti- viral activities. IMPORTANCEAs with many other viruses, the prototypic Old World mammarenavirus lymphocytic choriomeningitis virus (LCMV) can interfere with the host cell innate immune response to infection, which includes the double strand (ds)RNA sensor protein kinase receptor (PKR) pathway. A detailed understanding of LCMV-PKR interactions can provide novel insights about mammarenavirus-host cell interactions and facilitate the development of effective antiviral strategies against human pathogenic mammarenaviruses. In the present work, we present evidence that LCMV multiplication is enhanced in PKR- deficient cells, but pharmacological inhibition of PKR activation unexpectedly resulted in severely restricted propagation of LCMV. Likewise, we document a robust PKR activation in LCMV-infected cells in the absence of detectable levels of dsRNA. Our findings have revealed a complex role of the PKR pathway during LCMV infection and uncovered the activation of PKR as a druggable target for the development of antiviral drugs against human pathogenic mammarenaviruses.

microbiology↗

bioRxiv2021

Inhibitors of anti-apoptotic Bcl-2 family proteins exhibit potent and broad-spectrum anti-mammarenavirus activity via cell cycle arrest at G0/G1 phase

Targeting host factors is a promising strategy to develop broad-spectrum antiviral drugs. Drugs targeting anti-apoptotic Bcl-2 family proteins that were originally developed as tumor suppressors have been reported to inhibit multiplication of different types of viruses. However, the mechanisms whereby Bcl-2 inhibitors exert their antiviral activity remain poorly understood. In this study, we have investigated the mechanisms by which obatoclax (OLX) and ABT-737 Bcl-2 inhibitors exhibited a potent antiviral activity against the mammarenavirus lymphocytic choriomeningitis virus (LCMV). OLX and ABT-737 potent anti-LCMV activity was not associated with their pro-apoptotic properties, but rather their ability of inducing cell arrest at G0/G1 phase. OLX and ABT-737 mediated inhibition of Bcl-2 correlated with reduced expression levels of thymidine kinase 1 (TK1), cyclin A2 (CCNA2), and cyclin B1 (CCNB1) cell cycle regulators. In addition, siRNA-mediated knock down of TK1, CCNA2, and CCNB1 resulted in reduced levels of LCMV multiplication. The antiviral activity exerted by Bcl-2 inhibitors correlated with reduced levels of viral RNA synthesis at early times of infection. Importantly, ABT-737 exhibited moderate efficacy in a mouse model of LCMV infection, and Bcl-2 inhibitors displayed broad-spectrum antiviral activities against different mammarenaviruses and SARS-CoV-2. Our results suggest that Bcl-2 inhibitors, actively being explored as anti-cancer therapeutics, might be repositioned as broad-spectrum antivirals. IMPORTANCEAnti-apoptotic Bcl-2 inhibitors have been shown to exert potent antiviral activities against various types of viruses via mechanisms that are currently poorly understood. This study has revealed that Bcl-2 inhibitors mediated cell cycle arrest at the G0/G1 phase, rather than their pro-apoptotic activity, plays a critical role in blocking mammarenavirus multiplication in cultured cells. In addition, we show that Bcl-2 inhibitor ABT-737 exhibited moderate anti-mammarenavirus activity in vivo, and that Bcl-2 inhibitors displayed broad-spectrum antiviral activities against different mammarenaviruses and SARS-CoV-2. Our results suggest that Bcl-2 inhibitors, actively being explored as anti-cancer therapeutics, might be repositioned as broad-spectrum antivirals.

microbiology↗