furin protease inhibitor  (BPS Bioscience)

Journal: Science Advances

Article Title: The D614G mutation redirects SARS-CoV-2 spike to lysosomes and suppresses deleterious traits of the furin cleavage site insertion mutation

doi: 10.1126/sciadv.ade5085

Figure Lengend Snippet: ( A ) Anti-spike and anti-HSP90 immunoblots of cell lysates prepared from doxycycline-induced HTet1/S W1 cells that had been incubated with either control medium or with medium containing the furin protease inhibitor (fpi) decanoyl-RVKR-CMK (+fpi). MW markers, from top, are 250, 150, 100, 75 (pink), 50, 37, 25 (pink), 20, 15, and 10 kDa. ( B ) Histograms of cell surface anti-spike fluorescence measurements captured by flow cytometry of doxycycline-induced HTet1 cells (gray), HTet1/S W1 (gold), and HTet1/S W1 cells (brown) that had been incubated with the furin protease inhibitor (+fpi) decanoyl-RVKR-CMK. ( C ) Anti-spike and anti-HSP90 immunoblots of cell lysates prepared from equal amounts of doxycycline-induced HTet1, HTet1/S W1 , and HTet1/S W1 -CSM cells. MW markers, from top, are 250, 150, 100, 75 (pink), 50, 37, 25 (pink), 20, and 15 kDa. ( D ) Histograms of cell surface anti-spike fluorescence intensity data obtained by flow cytometry of doxycycline-induced HTet1 (gray), HTet1/S W1 (gold), and HTet1/S W1 -CSM cells (purple). ( E ) Anti-spike and anti-HSP90 immunoblots of cell lysates prepared from doxycycline-induced HTet1, HTet1/S W1 (WT), HTet1/S D614G (D614G), HTet1/S W1 -ΔN679-A688 cells (ΔN679-A688), and HTet1/S W1 -ΔR683-S689 cells (ΔR683-S689). MW markers, from top, are 250, 150, 100, 75 (pink), 50, 37, 25 (pink), 20, and 15 kDa. ( F ) Histograms of cell surface anti-spike fluorescence measurements captured by flow cytometry of doxycycline-induced HTet1 cells (gray), HTet1/S W1 cells (orange), HTet1/S D614G cells (blue), HTet1/S W1 -ΔN679-A688 cells (dark purple), and HTet1/S W1 -ΔR683-S689 (lavender). ( G ) Relative cell surface expression of spike in each of the cell lines tested in these experiments. Similar results were observed in three independent trials ( n = 3).

Article Snippet: The furin protease inhibitor decanoyl-RVKR-CMK was obtained from Tocris (#3501).

Techniques: Western Blot, Incubation, Control, Protease Inhibitor, Fluorescence, Flow Cytometry, Expressing

Journal: Science Advances

Article Title: The D614G mutation redirects SARS-CoV-2 spike to lysosomes and suppresses deleterious traits of the furin cleavage site insertion mutation

doi: 10.1126/sciadv.ade5085

Figure Lengend Snippet: Confocal immunofluorescence micrographs of doxycycline-induced cells that had been fixed, permeabilized, and stained using anti-spike antibodies specific for the C-terminal 14 amino acids of spike (green), DAPI (blue), and antibodies specific for Lamp2 (red). The cells shown here are doxycycline-induced ( A ) HTet1/S W1 ( B ) HTet1/S W1 cells incubated with the furin protease inhibitor (FPI), ( C ) HTet1/S W1 -CSM, ( D ) HTet1/S D614G , ( E ) HTet1/S W1 -ΔN679-A688, and ( F ) HTet1/S W1 -ΔR683-S689. Scale bars, 50 μm. These results are representative of two independent trials.

Article Snippet: The furin protease inhibitor decanoyl-RVKR-CMK was obtained from Tocris (#3501).

Techniques: Immunofluorescence, Staining, Incubation, Protease Inhibitor