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Figure 7. CHMP5 activity in mammalian cells. (A) Confocal images of immunofluorescence staining against <t>CHMP4B</t> of Hela-Kyoto cells expressing CHMP5- mNeonGreen under isosmotic or hypertonic conditions. (B and C) Confocal images of immunofluorescence staining against LAMP1 or CHMP4B of Hela-Kyoto cells expressing CHMP5-mNeonGreen with or without LLOMe treatment. Zoom-in of the deconvoluted image in C is shown in Fig. 8 A. (D) Live-cell imaging of Hela-Kyoto cells expressing CHMP5-mNeonGreen during osmotic shock. (E) Live-cell imaging of Hela-Kyoto cells expressing CHMP5-mNeonGreen upon LLOMe treatment. (F) Quantification of experiments described in D and E. Scale bar: 10 μm; D n = 3 ROI ≥400; E n = 3 ROI ≥2,500.
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Figure 7. CHMP5 activity in mammalian cells. (A) Confocal images of immunofluorescence staining against CHMP4B of Hela-Kyoto cells expressing CHMP5- mNeonGreen under isosmotic or hypertonic conditions. (B and C) Confocal images of immunofluorescence staining against LAMP1 or CHMP4B of Hela-Kyoto cells expressing CHMP5-mNeonGreen with or without LLOMe treatment. Zoom-in of the deconvoluted image in C is shown in Fig. 8 A. (D) Live-cell imaging of Hela-Kyoto cells expressing CHMP5-mNeonGreen during osmotic shock. (E) Live-cell imaging of Hela-Kyoto cells expressing CHMP5-mNeonGreen upon LLOMe treatment. (F) Quantification of experiments described in D and E. Scale bar: 10 μm; D n = 3 ROI ≥400; E n = 3 ROI ≥2,500.

Journal: The Journal of cell biology

Article Title: Vps60 initiates alternative ESCRT-III filaments.

doi: 10.1083/jcb.202206028

Figure Lengend Snippet: Figure 7. CHMP5 activity in mammalian cells. (A) Confocal images of immunofluorescence staining against CHMP4B of Hela-Kyoto cells expressing CHMP5- mNeonGreen under isosmotic or hypertonic conditions. (B and C) Confocal images of immunofluorescence staining against LAMP1 or CHMP4B of Hela-Kyoto cells expressing CHMP5-mNeonGreen with or without LLOMe treatment. Zoom-in of the deconvoluted image in C is shown in Fig. 8 A. (D) Live-cell imaging of Hela-Kyoto cells expressing CHMP5-mNeonGreen during osmotic shock. (E) Live-cell imaging of Hela-Kyoto cells expressing CHMP5-mNeonGreen upon LLOMe treatment. (F) Quantification of experiments described in D and E. Scale bar: 10 μm; D n = 3 ROI ≥400; E n = 3 ROI ≥2,500.

Article Snippet: Rabbit monoclonal antibody against LAMP1 (1/1,000 dilution for immunofluorescence) was from Cell Signaling (9091), and rabbit polyclonal antibody against CHMP4B (1/300) was from Proteintech (13683-1-AP).

Techniques: Activity Assay, Immunofluorescence, Staining, Expressing, Live Cell Imaging

Figure 8. CHMP5 and CHMP4B polymers have distinct dynamics in cells. (A) Decon- voluted confocal images of immunofluores- cence staining against LAMP1 or CHMP4B (same as Fig. 7 C, higher magnification) of Hela- Kyoto cells expressing CHMP5-mNeonGreen upon LLOMe treatment (scale bar: 10 μm, 2 μm inlet). (B) Live-cell imaging of Hela-Kyoto cells expressing CHMP4B-GFP and CHMP5- mScarlet-I undergoing nuclear envelop refor- mation and cytokinetic abscission (scale bar: 10 μm). Arrowheads indicate the reforming nu- clear envelope and the cytokinetic bridge. (C) Cartoon of the proposed model for function of Vps60-based ESCRT-III filaments.

Journal: The Journal of cell biology

Article Title: Vps60 initiates alternative ESCRT-III filaments.

doi: 10.1083/jcb.202206028

Figure Lengend Snippet: Figure 8. CHMP5 and CHMP4B polymers have distinct dynamics in cells. (A) Decon- voluted confocal images of immunofluores- cence staining against LAMP1 or CHMP4B (same as Fig. 7 C, higher magnification) of Hela- Kyoto cells expressing CHMP5-mNeonGreen upon LLOMe treatment (scale bar: 10 μm, 2 μm inlet). (B) Live-cell imaging of Hela-Kyoto cells expressing CHMP4B-GFP and CHMP5- mScarlet-I undergoing nuclear envelop refor- mation and cytokinetic abscission (scale bar: 10 μm). Arrowheads indicate the reforming nu- clear envelope and the cytokinetic bridge. (C) Cartoon of the proposed model for function of Vps60-based ESCRT-III filaments.

Article Snippet: Rabbit monoclonal antibody against LAMP1 (1/1,000 dilution for immunofluorescence) was from Cell Signaling (9091), and rabbit polyclonal antibody against CHMP4B (1/300) was from Proteintech (13683-1-AP).

Techniques: Staining, Expressing, Live Cell Imaging