Fig. 3: Recombinant PLY enhances endocytosis in a pore-dependent manner.

a Recombinant PLY (2 HU/ml) enhances endocytosis (FM 4-64 assay), and the process is blocked by preincubation of the toxin with cholesterol. Curves are normalized to the mock-treated cells. Representative images demonstrate the fluorescent intensity at corresponding time points (pseudocolor Fire LUT (ImageJ)). Scale bar: 50 µm. b Fluorescence intensity changes after PLY challenge (2 HU/ml) in microglia and astrocytes. All curves were normalized to the mock-treated cells. c Enhanced endocytosis by PLY in nonbrain cells such as HEK293 fibroblasts. d PLY-mediated enhanced endocytosis is eliminated due to the loss of the pore-forming capacity of PLY in the ∆6 mutant (∆6 PLY) and W433F mutant (W433F PLY). Similarly, the membrane-binding domain 4 of PLY (D4) does not alter endocytosis. All values represent the mean ± SEM, each dot symbol indicates an independent experiment (a n = 8 (Chol/PLY), n = 6 (PLY); b n = 11 (mock) and n = 13 (PLY); c n = 8; d n = 5), exact p-values are indicated (if significant), Wilcoxon matched pairs test (a), one-way ANOVA with Tukey post-test (d), all tests are two-tailed. Source data are provided as a Source Data file.