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Figure 4


Fig. 4. Microtubules are required for apoptotic cell fragmentation. (A) Influence of cytoskeletal inhibitors on apoptotic progression in UV-treated A431 cells, measured using the fluorogenic caspase substrate, Ac.DEVD.AMC (top) and by immunoblotting for cleaved PARP (bottom: tubulin shown as a loading control). Mean values (± s.e.m.) are shown from triplicate assays. (B) Assessment of apoptotic body formation in UV-treated A431 cells. Sub-5-µm apoptotic A431 cell fragments were collected by filtration and were counted by fluorescence or phase-contrast microscopy. Experiments were performed blind and are the mean ± s.e.m. of three experiments. (C) FACs analysis of apoptotic fragmentation. The relative numbers of UV-irradiated A431 cells with sub-G1 DNA content is shown in the bar chart (bottom) as a function of the value for UV only treated cells (normalised to 100%). Values are from three independent experiments (example traces are shown). (D-F) The effect of Latrunculin A on apoptotic progression (D, PARP cleavage), cellular fragmentation by apoptotic body assay (E), and FACS (F). Statistical significance by Student's t-test in A,B,C,E,F: *P<0.5; **P<0.01; ***P<0.001 compared with levels in the UV-alone group.





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