Journal: Journal of Virology

Article Title: Bovine coronavirus enters HRT-18 cells via membrane fusion and clathrin-mediated endocytosis in a low pH-, dynamin-, cholesterol-, microtubule-, Rab7-, and Rab11-dependent manner

doi: 10.1128/jvi.01274-25

Figure Lengend Snippet: BCoV entry into HRT-18 cells depends on cathepsin. ( A ) The maximum safe concentrations of E64d were determined using the CCK-8 assay. ( B ) Western blot analysis was used to evaluate the BCoV N protein expression levels, with grayscale analysis performed and presented as a bar graph. ( C ) RT-qPCR was performed to assess the BCoV gene copy numbers. ( D ) TCID 50 assay was used to measure the BCoV viral titers in the cell supernatant. ( E ) IFA was used to detect the number of BCoV-infected cells. Scale bar = 100 µm. ( F ) RT-qPCR was used to evaluate the effect of E64d on the viral entry. ( G ) RT-qPCR was used to evaluate the effect of E64d on BCoV attachment. Data are presented as the mean ± SD of three independent experiments (not significant, P > 0.05; * P < 0.05; ** P < 0.01; *** P < 0.001).

Article Snippet: The inhibitors used in this study included SSAA09E3 (Cat HY-138102, MedChemExpress), a novel inhibitor that blocks the fusion of the viral membrane with the host cell membrane; CPZ (Cat C0982, Sigma), a clathrin-mediated endocytosis inhibitor; nystatin (Cat 475914, Sigma), a caveolae inhibitor that acts as a sterol-binding agent disrupting caveolae; blebbistatin (Cat 203391, Sigma), an inhibitor of micropinocytosis; dynasore (Cat T1848, TargetMol), a dynamin inhibitor; MβCD (Cat T4072, TargetMol), a cholesterol depletion inhibitor; chloroquine (Cat S6999, Selleck) and NH 4 Cl (Cat A9434, Sigma), a potent inhibitor of V-ATPase and a specific inhibitor of acidification of endosomal vesicles; colchicine (Cat HY-16569, MedChemExpress), which inhibits the polymerization of tubulin; E64d (Cat S7393, Selleck), a cathepsin inhibitor; and camostat (Cat HY-13512, MedChemExpress), a TMPRSS2 inhibitor.

Techniques: CCK-8 Assay, Western Blot, Expressing, Quantitative RT-PCR, Infection

Journal: The FEBS journal

Article Title: Verapamil treatment induces cytoprotective autophagy by modulating cellular metabolism.

doi: 10.1111/febs.14064

Figure Lengend Snippet: Fig. 7: Verapamil induces autophagic flux Western blots for LC3 protein levels after treatment with increasing concentrations of Verapamil (Ver; 50, 100, 200 µM; 6 h) alone or in combination with lysosomal protease inhibitors – E64d/Pepstatin A (E/PepA; 10 µM; 6 h) (A-E). Actin or ERK2 were used as a loading control. Representative images of the Western blots are shown (upper panels) together with the quantification of LC3-II fold increase in comparison to control, untreated cells (lower panels). Each performed experiment is presented as an individual data point set with mean (±SEM), n= 3 to 6.

Article Snippet: Verapamil chloride (Sigma-Aldrich, V4629), E64d (LKT Laboratories, E0003), Pepstatin A (Calbiochem, 516481), Chloroquine (Sigma-Aldrich, C6620), Sodium oxamate (Sigma-Aldrich, O2751).

Techniques: Western Blot, Control, Comparison

Journal: iScience

Article Title: Dynamic association of the intramembrane proteases SPPL2a/b and their substrates with tetraspanin-enriched microdomains

doi: 10.1016/j.isci.2023.107819

Figure Lengend Snippet:

Article Snippet: E-64d , Enzo Lifesciences , BML-PI107-0001.

Techniques: Virus, Recombinant, Agarose Gel Electrophoresis, Protease Inhibitor, Modification, Western Blot, Transfection, Extraction, Bicinchoninic Acid Protein Assay, Plasmid Preparation, Software