Vl. Sukhorukov et al., DNA, PROTEIN, AND PLASMA-MEMBRANE INCORPORATION BY ARRESTED MAMMALIAN-CELLS, The Journal of membrane biology, 142(1), 1994, pp. 77-92
Incorporation of DNA, protein, and plasma membrane during blockage by
aphidicolin or by doxorubicin was studied by flow cytometry and electr
ororation of three cell lines (mouse-myeloma Sp2/0-Ag14, hybridoma H73
C11, and fibroblast-like L929 cells). Drug-mediated arrest at the G1-S
boundary (aphidicolin) or in G2/M (doxorubicin) did not arrest synthe
sis of either protein or total membrane area, the increases in which o
utstripped growth in cell volume and apparent cell area, respectively.
Measurements of membrane capacity in normal and hypo-osmotic media sh
owed that the drugs had not changed the fundamental bilayer, but that
an increase in the number or size of microvilli must have occurred. Ap
hidicolin-arrested cells withstood hypo-osmotic stress better than unt
reated cells could, indicating that the membrane excess can be utilize
d as a reserve during rapid cell expansion. Hypo-osmotically treated c
ell populations exhibited only about half the coefficient of variance
(CV) in membrane properties of cells at physiological osmolality. Popu
lations of arrested cells exhibited the same high CV as asynchronous c
ells, indicating that chemical arrest does not give uniformly villated
eel populations. However, the lowest CV values were given by some syn
chronized (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. Afte
r removal of doxorubicin, the cells did not divide: some continued pro
tein synthesis, grew abnormally large, and further increased their mem
brane excess. Membrane breakdown by electric pulsing (3 X 5kV/cm, 40 m
u sec 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-cel
l capacitance were reduced), presumably via endocytosis-like vesiculat
ion.