The HN protein of Newcastle disease virus induces cell apoptosis through the induction of lysosomal membrane permeabilization
Fig 5
CTSB and CTSD exacerbate NDV-induced LMP by inducing the generation of ROS.
(A) Intracellular ROS levels were measured by flow cytometry using DCFH-DA staining in HeLa cells infected with Herts/33 at 0.01 MOI for different timepoints or mock-infected. (B) HeLa cells were transfected with si-CTSB, si-CTSD, or si-NC for 48h, or mock-transfected, and intracellular ROS levels were measured by flow cytometry using DCFH-DA staining following infection with Herts/33 at 0.01 MOI or mock-infection for 12h. (C) HeLa cells were pretreated with LLoMe (500 nM), DMSO, or mock-untreated for 3h, respectively. Subsequently, the intracellular ROS levels were measured by flow cytometry using DCFH-DA staining cells following infection with Herts/33 at 0.01 MOI or mock-infection for 24h. (D) HeLa cells were treated with NAC (2 mM) for 24h after absorption with Herts/33 at 0.01 MOI for 1h. LMP levels were evaluated by confocal microscopy using indicated antibodies. Scale bars, 20 μm. Manders’ Colocalization Coefficients of galectin 3 with LAMP1 were quantified by ImageJ software and shown on the right. Error bars represent SDs for triplicate analyses of three independent experiments (A-C) or SDs for 15 cells (D). All significance analyses were assessed using one-way ANOVA with Dunnett’s multiple comparisons test.