Reconstruction of the cell entry pathway of an extinct virus
Fig 2
Heparan sulfate facilitates HERV-K Env-mediated entry and attachment.
(A) WT HAP1, EXT1KO Neor Ctrl (transduced with a control retrovirus), EXT1KO EXT1-HA (+ addback), SLC35B2KO Neor Ctrl and SLC35B2KO SLC25B2-HA (+ addback) cells were infected with VSV-HERVK or VSV and infectivity analyzed by flow cytometry. Fold difference in percent infected cells compared to WT for VSV-HERVK was normalized to that of VSV for each condition. Error bars represent SEM for four independent experiments. (B) Relative MFI of cells from (A). Data were normalized as in (A). (C) BSRT7 cells were treated with 50mM sodium chlorate and infected with either VSV or VSV-HERVK. Fold difference in both percent infected cells (left) and MFI (right) compared to untreated cells for VSV-HERVK was normalized to that of VSV. Error bars represent SEM for three independent experiments. (D) VSV or VSV-HERVK was incubated with the indicated soluble glycosaminoglycans prior to infecting BSRT7 cells. Percent infected cells was normalized to untreated virus controls. Error bars represent SEM for three independent experiments. (E) Schematic of virus attachment experiment. Cells were blocked with BSA then incubated with both fluorescently labeled VSV-HERVK and VSV at either 37°C or 4°C. (F) Representative images from 4°C attachment experiment. Red: VSV. Green: VSV-HERVK. Blue: DAPI. Grey: calcein. (G) Results of attachment experiment. Numbers of particles/μm2 are plotted. Grey circles indicate outliers. Total number of particles counted per condition is indicated above each box.