Figure 1.

Human Niemann-Pick C1-like1 (NPC1L1) lacks filovirus receptor activity. (a) NPC1-null Chinese hamster ovary (CHO) CT43 cells were engineered to express human NPC1-flag or NPC1L1-flag. Protein expression was detected by immunoblotting with an anti-flag antibody. M_r, relative molecular weight in kilodaltons (kDa). (b) Efflux of lysosomal cholesterol mediated by NPC1-flag and NPC1L1-flag in CT43 cells was determined by filipin staining and fluorescence microscopy. Scale bar, 20 μm. (c) Infection of CT43 cells expressing NPC1-flag or NPC1L1-flag by vesicular stomatitis virus (VSV) pseudotypes bearing VSV G or filovirus glycoproteins. Asterisks indicate data points at or below the limit of detection. (d) Co-immunoprecipitation (co-IP) of NPC1 and NPC1L1 by Ebola virus (EBOV) GP. Magnetic beads coated with GP-specific monoclonal antibody KZ52 were incubated with detergent extracts containing no virus (None), uncleaved VSV-GP, or cleaved VSV-GP_CL. Control or glycoprotein-decorated beads were mixed with cell extracts containing NPC1-flag or NPC1L1-flag. Beads were then retrieved and flag-tagged proteins in the immune pellets and supernatants were detected by immunoblotting (IB).