Rowejosephsen0571

The Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is a highly contagious virus that underlies the current COVID-19 pandemic. SARS-CoV-2 is thought to disable various features of host immunity and cellular defense. The SARS-CoV-2 nonstructural protein 1 (Nsp1) is known to inhibit host protein translation and could be a target for antiviral therapy against COVID-19. However, how SARS-CoV-2 circumvents this translational blockage for the production of its own proteins is an open question. Here, we report a bipartite mechanism of SARS-CoV-2 Nsp1 which operates by (1) hijacking the host ribosome via direct interaction of its C-terminal domain (CT) with the 40S ribosomal subunit and (2) specifically lifting this inhibition for SARS-CoV-2 via a direct interaction of its N-terminal domain (NT) with the 5' untranslated region (5' UTR) of SARS-CoV-2 mRNA. We show that while Nsp1-CT is sufficient for binding to 40S and inhibition of host protein translation, the 5' UTR of SARS-CoV-2 mRNA removes this inhibition by binding to Nsp1-NT, suggesting that the Nsp1-NT-UTR interaction is incompatible with the Nsp1-CT-40S interaction. Indeed, lengthening the linker between Nsp1-NT and Nsp1-CT of Nsp1 progressively reduced the ability of SARS-CoV-2 5' UTR to escape the translational inhibition, supporting that the incompatibility is likely steric in nature. The short SL1 region of the 5' UTR is required for viral mRNA translation in the presence of Nsp1. Thus, our data provide a comprehensive view on how Nsp1 switches infected cells from host mRNA translation to SARS-CoV-2 mRNA translation, and that Nsp1 and 5' UTR may be targeted for anti-COVID-19 therapeutics.Viruses rely on the host translation machinery to synthesize their own proteins. MK-0859 nmr Consequently, they have evolved varied mechanisms to co-opt host translation for their survival. SARS-CoV-2 relies on a non-structural protein, NSP1, for shutting down host translation. Despite this, it is currently unknown how viral proteins and host factors critical for viral replication can escape a global shutdown of host translation. Here, using a novel FACS-based assay called MeTAFlow, we report a dose-dependent reduction in both nascent protein synthesis and mRNA abundance in cells expressing NSP1. link2 We perform RNA-Seq and matched ribosome profiling experiments to identify gene-specific changes both at the mRNA expression and translation level. We discover a functionally-coherent subset of human genes preferentially translated in the context of NSP1 expression. These genes include the translation machinery components, RNA binding proteins, and others important for viral pathogenicity. Importantly, we also uncover potential mechanisms of preferential translation through the presence of shared sites for specific RNA binding proteins and a remarkable enrichment for 5' terminal oligo-pyrimidine tracts. Collectively, the present study suggests fine tuning of host gene expression and translation by NSP1 despite its global repressive effect on host protein synthesis.Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the causative agent of coronavirus disease 2019 (COVID-19), enters the host cells through two main pathways, both involving key interactions between viral envelope-anchored spike glycoprotein of the novel coronavirus and the host receptor, angiotensin-converting enzyme 2 (ACE2). To date, SARS-CoV-2 has infected up to 26 million people worldwide; yet, there is no clinically approved drug or vaccine available. Therefore, a rapid and coordinated effort to re-purpose clinically approved drugs that prevent or disrupt these critical entry pathways of SARS-CoV-2 spike glycoprotein interaction with human ACE2, could potentially accelerate the identification and clinical advancement of prophylactic and/or treatment options against COVID-19, thus providing possible countermeasures against viral entry, pathogenesis and survival. Herein, we discovered that Ambroxol hydrochloride (AMB), and its progenitor, Bromhexine hydrochloride (BHH), both clinically approved drugs are potent effective modulators of the key interaction between the receptor binding domain (RBD) of SARS-CoV-2 spike protein and human ACE2. We also found that both compounds inhibited SARS-CoV-2 infection-induced cytopathic effect at micromolar concentrations. Therefore, in addition to the known TMPRSS2 activity of BHH; we report for the first time that the BHH and AMB pharmacophore has the capacity to target and modulate yet another key protein-protein interaction essential for the two known SARS-CoV-2 entry pathways into host cells. Altogether, the potent efficacy, excellent safety and pharmacologic profile of both drugs along with their affordability and availability, makes them promising candidates for drug repurposing as possible prophylactic and/or treatment options against SARS-CoV-2 infection.The ability to detect recombination in pathogen genomes is crucial to the accuracy of phylogenetic analysis and consequently to forecasting the spread of infectious diseases and to developing therapeutics and public health policies. However, previous methods for detecting recombination and reassortment events cannot handle the computational requirements of analyzing tens of thousands of genomes, a scenario that has now emerged in the effort to track the spread of the SARS-CoV-2 virus. Furthermore, the low divergence of near-identical genomes sequenced in short periods of time presents a statistical challenge not addressed by available methods. In this work we present Bolotie, an efficient method designed to detect recombination and reassortment events between clades of viral genomes. We applied our method to a large collection of SARS-CoV-2 genomes and discovered hundreds of isolates that are likely of a recombinant origin. In cases where raw sequencing data was available, we were able to rule out the possibility that these samples represented co-infections by analyzing the underlying sequence reads. Our findings further show that several recombinants appear to have persisted in the population.We present a comprehensive vaccine strategy for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) by combining antigen optimization and nanoparticle display. We first developed a receptor binding domain (RBD)-specific antibody column for purification and displayed the RBD on self-assembling protein nanoparticles (SApNPs) using the SpyTag/SpyCatcher system. We then identified the heptad repeat 2 (HR2) stalk as a major cause of spike metastability, designed an HR2-deleted glycine-capped spike (S2GΔHR2), and displayed S2GΔHR2 on three SApNPs with high yield, purity, and antigenicity. Compared to the RBD, the RBD-ferritin SApNP elicited a more potent murine neutralizing antibody (NAb) response on par with the spike. S2GΔHR2 elicited two-fold-higher NAb titers than the proline-capped spike (S2P), while S2GΔHR2 SApNPs derived from multilayered E2p and I3-01v9 60-mers elicited up to 10-fold higher NAb titers. The S2GΔHR2-presenting I3-01v9 SApNP also induced critically needed T-cell immunity, thereby providing a next-generation vaccine candidate to battle the COVID-19 pandemic.

The receptor binding domain and stabilized SARS-CoV-2 spike were displayed on nanoparticles as vaccine antigens and elicited potent immune responses.

The receptor binding domain and stabilized SARS-CoV-2 spike were displayed on nanoparticles as vaccine antigens and elicited potent immune responses.Neuropilin-1 (NRP-1) is a multifunctional transmembrane receptor for ligands that affect developmental axonal growth and angiogenesis. In addition to a role in cancer, NRP-1 is a reported entry point for several viruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causal agent of coronavirus disease 2019 (COVID-19). The furin cleavage product of SARS-CoV-2 Spike protein takes advantage of the vascular endothelial growth factor A (VEGF-A) binding site on NRP-1 which accommodates a polybasic stretch ending in a C-terminal arginine. This site has long been a focus of drug discovery efforts for cancer therapeutics. We recently showed that interruption of the VEGF-A/NRP-1 signaling pathway ameliorates neuropathic pain and hypothesize that interference of this pathway by SARS-CoV-2 spike protein interferes with pain signaling. Here, we report hits from a small molecule and natural product screen of nearly 0.5 million compounds targeting the VEGF-A binding site on NRP-1. We identified nine chemical series with lead- or drug-like physico-chemical properties. Using an ELISA, we demonstrate that six compounds disrupt VEGF-A-NRP-1 binding more effectively than EG00229, a known NRP-1 inhibitor. Secondary validation in cells revealed that almost all tested compounds inhibited VEGF-A triggered VEGFR2 phosphorylation. Two compounds displayed robust inhibition of a recombinant vesicular stomatitis virus protein that utilizes the SARS-CoV-2 Spike for entry and fusion. These compounds represent a first step in a renewed effort to develop small molecule inhibitors of the VEGF-A/NRP-1 signaling for the treatment of neuropathic pain and cancer with the added potential of inhibiting SARS-CoV-2 virus entry.Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). link3 The initial interaction between Transmembrane Serine Protease 2 (TMPRSS2) primed SARS-CoV-2 spike (S) protein and host cell receptor angiotensin-converting enzyme 2 (ACE-2) is a pre-requisite step for this novel coronavirus pathogenesis. Here, we expressed a GFP-tagged SARS-CoV-2 S-Ectodomain in Tni insect cells. That contained sialic acid-enriched N- and O-glycans. Surface resonance plasmon (SPR) and Luminex assay showed that the purified S-Ectodomain binding to human ACE-2 and immunoreactivity with COVID-19 positive samples. We demonstrate that bromelain (isolated from pineapple stem and used as a dietary supplement) treatment diminishes the expression of ACE-2 and TMPRSS2 in VeroE6 cells and dramatically lowers the expression of S-Ectodomain. Importantly, bromelain treatment reduced the interaction between S-Ectodomain and VeroE6 cells. Most importantly, bromelain treatment significantly diminished the SARS-CoV-2 infection in VeroE6 cells. Altogether, our results suggest that bromelain or bromelain rich pineapple stem may be used as an antiviral against COVID-19.

Bromelain inhibits / cleaves the expression of ACE-2 and TMPRSS2Bromelain cleaves / degrades SARS-CoV-2 spike proteinBromelain inhibits S-Ectodomain binding and SARS-CoV-2 infection.

Bromelain inhibits / cleaves the expression of ACE-2 and TMPRSS2Bromelain cleaves / degrades SARS-CoV-2 spike proteinBromelain inhibits S-Ectodomain binding and SARS-CoV-2 infection.Pre-existing immune responses to seasonal endemic coronaviruses could have profound consequences for antibody responses to SARS-CoV-2, either induced in natural infection or through vaccination. Such consequences are well established in the influenza and flavivirus fields. A first step to establish whether pre-existing responses can impact SARS-CoV-2 infection is to understand the nature and extent of cross-reactivity in humans to coronaviruses. We compared serum antibody and memory B cell responses to coronavirus spike (S) proteins from pre-pandemic and SARS-CoV-2 convalescent donors using a series of binding and functional assays. We found weak evidence of pre-existing SARS-CoV-2 cross-reactive serum antibodies in pre-pandemic donors. However, we found stronger evidence of pre-existing cross-reactive memory B cells that were activated on SARS-CoV-2 infection. Monoclonal antibodies (mAbs) isolated from the donors showed varying degrees of cross-reactivity with betacoronaviruses, including SARS and endemic coronaviruses.