Extended Data Fig. 9: ERGIC compartmentalization is dependent on microtubules.
From: A dual role of ERGIC-localized Rabs in TMED10-mediated unconventional protein secretion
a, Immunofluorescence and SIM analysis of GFP-ERGIC53 and Tubulin in HeLa cells with or without Nocodazole treatment (10 μM) using SPY555-tubulin. Scale bar, 5 μm. b, Immunofluorescence and SIM analysis of GFP-ERGIC53 and TMED10-V5 stably expressed in HeLa cells with or without Nocodazole treatment (10 μM) using anti-V5 antibody. Scale bar, 5 μm in overview and 1 μm in the zoomed in. Diagram showing the percentage of TMED10 separated from ERGIC53 (mean ± SD). P values are indicated (two-tailed t test, n=52 cells, ns, no significant difference). c, Secretion of mIL-1β in HEK293T cells transfected with mIL-1β-FLAG plasmid together with GFP/GFP–Rab2A plasmids with or without Nocodazole. Diagram showing the quantification of IL-1β secretion (mean ± SD). P values are indicated (two-tailed t test, n=3 experiments, ns, no significant difference). d, Immunofluorescence and SIM analysis of GFP-ERGIC53 and TMED10-V5 stably expressed in HeLa cells with or without Cytochalasin B treatment (10 μM) using anti-V5 antibody. Actin was labelled with Phalloidin. Scale bar, 5 μm in overview and 2 μm in the zoomed in. e, Immunofluorescence and SIM analysis of GFP-ERGIC53 and TMED10-V5 stably expressed in HeLa cells with or without Latrunculin A treatment (0.4 μM) using anti-V5 antibody. Actin was labelled with Phalloidin. Scale bar, 5 μm in overview and 2 μm in the zoomed in. f, Secretion of mIL-1β in HEK293T cells transfected with mIL-1β-FLAG plasmid together with GFP/GFP–Rab2A plasmids with or without Cytochalasin B (10 μM). The data are representative of three independent experiments. g, Secretion of mIL-1β in HEK293T cells transfected with mIL-1β-FLAG plasmid together with GFP/GFP–Rab2A plasmids with or without Latrunculin A (0.4 μM). f,g, The data are representative of three independent experiments. h, Immunofluorescence and SIM analysis of GFP-ERGIC53 and TMED10-V5 stably expressed in HeLa cells with or without BFA treatment (0.5 ng/μL) using anti-V5 antibody and anti-Rab1A/1B/2A antibody. Scale bar, 5 μm in overview and 2 μm in the zoomed in. Diagram showing the percentage of TMED10 in ERGIC53 (mean ± SD). P values are indicated (two-tailed t test, n=50 cells, ns, no significant difference). Exact P values are presented in Source numerical data. Source numerical data and unprocessed blots are available in Source data.
