Figure 5.

Inhibition of lysosomal proteases does not block LLO‐dependent Ubc9 degradation. HRP was endocytosed in HeLa cells for 15 min. After different chasing period (0, 30, or 90 min), cells were incubated with DAB and H₂O₂ to cross‐link lysosomal content. (a) Quantification of cathespin activity in HeLa whole cell extracts treated or not with HRP/DAB (“CtsB” = substrate specific for cathepsin B; “Cys‐cathepsins” = substrate specific for cathepsins belonging to cysteine‐proteases) (mean ± SD from at least three independent experiments; *, p < 0.05; NS, not significant; unpaired two‐tailed Student's t test). (b) Immunoblot analysis using anti‐Ubc9 and anti‐actin antibodies of whole cell extracts from HeLa cells pretreated with HRP/DAB and incubated with 3 nM LLO for 10 or 30 min