Journal: Scientific Reports

Article Title: Vesicular stomatitis virus G protein transmembrane region is crucial for the hemi-fusion to full fusion transition

doi: 10.1038/s41598-018-28868-y

Figure Lengend Snippet: TM replacement impaired VSV G protein mediated virus infection. ( a ) WT or chimeric VSV G proteins were packed into lentiviruses at similar levels. HEK293 T cells were co-transfected with pLVTHM, pMD 2.G (WT or chimeric VSV G) and psPAX2 to generate lentivirus. Lentivirus was collected and analyzed by western blot with a VSV G antibody. The capsid protein P24 was used as a normalization control. ( b and c ) TM replacement remarkably attenuated VSV G induced lentivirus infection. HeLa cells were infected by lentivirus enveloped with WT or chimeric VSV G. A GFP gene was used as a reporter of virus infection. Lentivirus-infected cells (GFP positive) were analyzed by flow cytometry. The cell infection efficiency decreased dramatically when VSV G TM was replaced by TM from Stx1A, VAMP2 or HA (***P < 0.001). Scale bar, 30 μm. ( d – f ) TM replacement of VSV G reduced the infectivity of rVSV. ( d ) 293 T cells were co-transfected with pBS-N, pBS-P, pBS-L, pCAG-T7 and WT or mutant pVSV to produce rVSV. WT or chimeric VSV G packaging into rVSV was determined by western blot. ( e and f ) The infection efficiency of rVSV enveloped with chimeric VSV G was lower than that of WT rVSV. Plaque assays were performed by serial dilution of the indicated virus. The plaque morphologies of rVSV with WT or Syntaxin, VAMP2 TM-VSV G (10 −4 dilution) and rVSV with HA TM-VSV G (10 −3 dilution) are shown in ( f ).

Article Snippet: 1 × 10 6 /well HEK293 T cells were seeded into 12-well plate and transfected with 1.17 μg pLVTHM (Addgene 12247), 0.53 μg pMD 2.G (Addgene 12259) and 0.8 μg psPAX2 (Addgene 12260) plasmids.

Techniques: Virus, Infection, Transfection, Western Blot, Control, Flow Cytometry, Mutagenesis, Serial Dilution

Journal: Frontiers in Genetics

Article Title: Anti-Fungal Drug Anidulafungin Inhibits SARS-CoV-2 Spike-Induced Syncytia Formation by Targeting ACE2-Spike Protein Interaction

doi: 10.3389/fgene.2022.866474

Figure Lengend Snippet: (A) Mechanism of action to prevent SARS-CoV-2 entry into the target host cell. SARS-CoV-2 enters human cells after the interaction of the spike protein with the ACE2 receptor. Blocking ACE2-spike interactions by targeting ACE2 receptors with antiviral compounds is an important approach for developing novel therapeutics against SARS-CoV2. (B) Schematic overview of searching novel inhibitors at the proposed study.

Article Snippet: Human ACE2 (Addgene #1786), pLVTHM/GFP (Addgene #12247), psPAX2 (Addgene 12,260), pMD2.G (Addgene #12259) were obtained from Addgene. pAAV-CMV-GFP was obtained from L. Zentilin (Molecular Medicine Lab, ICGEB). pAAV-spike-V5 and pAA-spike-d19-V5 SARS-CoV-2 spike expression vectors were used previously ( ).

Techniques: Blocking Assay

Journal: Frontiers in Genetics

Article Title: Anti-Fungal Drug Anidulafungin Inhibits SARS-CoV-2 Spike-Induced Syncytia Formation by Targeting ACE2-Spike Protein Interaction

doi: 10.3389/fgene.2022.866474

Figure Lengend Snippet: Docked all 450 compounds with ACE2 receptor (A) . The binding interaction H-bonds (green) and the amino acid residues of ACE2 and anidulafungin at the binding site (B) . The 2D plot of the ACE2-anidulafungin binding pose (C) .

Article Snippet: Human ACE2 (Addgene #1786), pLVTHM/GFP (Addgene #12247), psPAX2 (Addgene 12,260), pMD2.G (Addgene #12259) were obtained from Addgene. pAAV-CMV-GFP was obtained from L. Zentilin (Molecular Medicine Lab, ICGEB). pAAV-spike-V5 and pAA-spike-d19-V5 SARS-CoV-2 spike expression vectors were used previously ( ).

Techniques: Binding Assay

Journal: Frontiers in Genetics

Article Title: Anti-Fungal Drug Anidulafungin Inhibits SARS-CoV-2 Spike-Induced Syncytia Formation by Targeting ACE2-Spike Protein Interaction

doi: 10.3389/fgene.2022.866474

Figure Lengend Snippet: The elucidation of MD simulation of native ACE2 and ACE2-docked complexes. (A) Representation of C-alpha conformation of RMSD. (B) Comparative RMS fluctuation plot of native ACE2 and ACE2-docked complexes. (C) Rg analysis of native ACE2 and ACE2-docked complexes. (D) SASA plot of native ACE2 and ACE2-docked complexes.

Article Snippet: Human ACE2 (Addgene #1786), pLVTHM/GFP (Addgene #12247), psPAX2 (Addgene 12,260), pMD2.G (Addgene #12259) were obtained from Addgene. pAAV-CMV-GFP was obtained from L. Zentilin (Molecular Medicine Lab, ICGEB). pAAV-spike-V5 and pAA-spike-d19-V5 SARS-CoV-2 spike expression vectors were used previously ( ).

Techniques:

Journal: Frontiers in Genetics

Article Title: Anti-Fungal Drug Anidulafungin Inhibits SARS-CoV-2 Spike-Induced Syncytia Formation by Targeting ACE2-Spike Protein Interaction

doi: 10.3389/fgene.2022.866474

Figure Lengend Snippet: The average values of RMSD, Rg, and SASA of the native ACE2 and complex containing compounds anidulafungin, lopinavir, indinavir, and MLN-4760.

Article Snippet: Human ACE2 (Addgene #1786), pLVTHM/GFP (Addgene #12247), psPAX2 (Addgene 12,260), pMD2.G (Addgene #12259) were obtained from Addgene. pAAV-CMV-GFP was obtained from L. Zentilin (Molecular Medicine Lab, ICGEB). pAAV-spike-V5 and pAA-spike-d19-V5 SARS-CoV-2 spike expression vectors were used previously ( ).

Techniques:

Journal: Frontiers in Genetics

Article Title: Anti-Fungal Drug Anidulafungin Inhibits SARS-CoV-2 Spike-Induced Syncytia Formation by Targeting ACE2-Spike Protein Interaction

doi: 10.3389/fgene.2022.866474

Figure Lengend Snippet: Free energy landscape analysis of (A) native ACE2 and complexes with Anidulafungin (B) , lopinavir (C) , indinavir (D) , and MLN-4760 (E) compounds.

Article Snippet: Human ACE2 (Addgene #1786), pLVTHM/GFP (Addgene #12247), psPAX2 (Addgene 12,260), pMD2.G (Addgene #12259) were obtained from Addgene. pAAV-CMV-GFP was obtained from L. Zentilin (Molecular Medicine Lab, ICGEB). pAAV-spike-V5 and pAA-spike-d19-V5 SARS-CoV-2 spike expression vectors were used previously ( ).

Techniques:

Journal: Frontiers in Genetics

Article Title: Anti-Fungal Drug Anidulafungin Inhibits SARS-CoV-2 Spike-Induced Syncytia Formation by Targeting ACE2-Spike Protein Interaction

doi: 10.3389/fgene.2022.866474

Figure Lengend Snippet: Anidulafungin impaired the spike-pseudotyped particle internalization. (A) Schematic representation of the SARS-Cov2 spike pseudotyped particle sentry assay. (B,C) HEK293/ACE2 cells were pre-treated with indicated drugs 1–2 h before adding the spike pseudotyped particles carrying GFP as a reporter. After 36 h, cells were immunostained with anti-GFP (green) and nuclei (blue). Representative images are in panel (B) (spike pseudotyped particles), and quantifications in panel (C) (spike pseudotyped particles). Data (mean ± SD; n = 6, Mann-Whitney U test ) are plotted as the percentage of GFP + cells normalized on the total number of cells.

Article Snippet: Human ACE2 (Addgene #1786), pLVTHM/GFP (Addgene #12247), psPAX2 (Addgene 12,260), pMD2.G (Addgene #12259) were obtained from Addgene. pAAV-CMV-GFP was obtained from L. Zentilin (Molecular Medicine Lab, ICGEB). pAAV-spike-V5 and pAA-spike-d19-V5 SARS-CoV-2 spike expression vectors were used previously ( ).

Techniques: MANN-WHITNEY