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DNA, protein, and plasma-membrane incorporation by arrested mammalian cells

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Abstract

Incorporation of DNA, protein, and plasma membrane during blockage by aphidicolin or by doxorubicin was studied by flow cytometry and electrorotation of three cell lines (mouse-myeloma Sp2/0-Ag14, hybridoma H73C11, and fibroblast-like L929 cells). Drug-mediated arrest at the G1-S boundary (aphidicolin) or in G2/M (doxorubicin) did not arrest synthesis of either protein or total membrane area, the increases in which outstripped growth in cell volume and apparent cell area, respectively. Measurements of membrane capacity in normal and hypo-osmotic media showed that the drugs had not changed the fundamental bilayer, but that an increase in the number or size of microvilli must have occurred. Aphidicolin-arrested cells withstood hypo-osmotic stress better than untreated cells could, indicating that the membrane excess can be utilized as a reserve during rapid cell expansion.

Hypo-osmotically treated cell populations exhibited only about half the coefficient of variance (CV) in membrane properties of cells at physiological osmolality. Populations of arrested cells exhibited the same high CV as asynchronous cells, indicating that chemical arrest does not give uniformly villated cell populations. However, the lowest CV values were given by some synchronized (aphidicolin-blocked, then released) populations.

Removal of aphidicolin allowed most cells to progress through S and G2, and then divide. During these processes, the membrane excess was reduced. After removal of doxorubicin, the cells did not divide: some continued protein synthesis, grew abnormally large, and further increased their membrane excess.

Membrane breakdown by electric pulsing (3 X 5kV/cm, 40 μsec decay time) of aphidicolin-synchronized L cells in G2/M led to a 22% loss of plasma membrane (both the area-specific and the whole-cell capacitance were reduced), presumably via endocytosislike vesiculation.

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  1. W. M. Arnold

    Present address: CESIT, IRL Ltd., Gracefield Research Centre, PO Box 31-310, Lower Hutt, New Zealand

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  1. Lehrstuhl für Biotechnologie, Biozentrum der Universität Würzburg, Am Hubland, D-97074, Würzburg, Germany

    V. L. Sukhorukov, C. S. Djuzenova, W. M. Arnold & U. Zimmermann

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  1. V. L. Sukhorukov
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  2. C. S. Djuzenova
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This work was supported by grants of the VDI/VDE (13 MV 0305 to W.M.A. and U.Z.), of the DARA 50WB9212 (to U.Z.), and of the Deutsche Forschungsgemeinschaft (SB 176 B5 to U.Z. and W.M.A.).

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Sukhorukov, V.L., Djuzenova, C.S., Arnold, W.M. et al. DNA, protein, and plasma-membrane incorporation by arrested mammalian cells. J. Membarin Biol. 142, 77–92 (1994). https://doi.org/10.1007/BF00233385

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