FIGURE 4.

Cathepsin and endosome acidification inhibitors can inhibit TLR3 proteolytic processing. A, shown are Western blot (WB) results of TLR3 expressed in HEK293T cells mock-treated or treated with bafilomycin at a final concentration of 2.5 nm. The cotransfection of a plasmid expressing Unc93b1 is denoted with a + symbol. All of the images were from the same experiment and the same Western blot. However, unrelated results were removed. B, the cathepsin inhibitor z-FA-FMK can inhibit CTF accumulation while increasing the abundance of the full-length TLR3 in transiently transfected HEK293T cells. C, z-FA-FMK can inhibit TLR3 proteolytic processing in transiently transfected Huh7.5 cells. Transfection efficiency of Huh7.5 cells is lower than HEK293T cells; thus, TLR3-HA was immunoprecipitated (IP) to increase sensitivity of detection. D, z-FA-FMK can inhibit TLR3 proteolytic processing in HEK293T cells with endogenous level Unc93b1. Cell lysates were immunoprecipitated with anti-HA antibody and detected by Western blot for HA epitope. These experiments were repeated three times with the same results.