Ca. Heckman et al., PERSISTENT EFFECTS OF PHORBOL 12-MYRISTATE 13-ACETATE - POSSIBLE IMPLICATION OF VESICLE TRAFFIC, Journal of cellular physiology, 166(1), 1996, pp. 217-230
Relative to their normal counterparts, transformed epithelial cells ha
ve a distinctive and quantifiable three-dimensional shape. Biophysical
and mathematical methods are used to distinguish these extremes in ce
lls from two lines, cultured from rat liver and tracheal epithelium, r
espectively. Cells adopted a more transformed-looking configuration tr
ansiently when exposed to phorbol 12-myristate 13-acetate (PMA) (Plumm
er and Heckman, [1990] Exp. Cell Res., 188:66-74). The purpose of the
present work was to dissect the physiological processes involved in th
e shape change. Ruffling activity, known to be PMA-stimulated in other
cells, was investigated. Although the ruffles appeared less robust th
an normal, PMA stimulated ruffling activity over a 5 h period. The num
ber of sites where ruffling was initiated declined by 5 h, however, an
d suppression was seen by 10 h. Cells from both lines adopted the tran
sformed shape configuration when exposed for 2 h to monensin. When the
subset of shape features changed by this treatment was compared with
those originally changed during transformation, it was found that mone
nsin-treated cells mimicked the features of transformed cells. Its eff
ect on ruffling was, however, unlike PMA's. Thus, the phenotype was un
likely to arise from ruffling itself but might be a process driven by
ruffling. Chloroquine also stimulated cells to assume characteristics
of transformed cells. Since both it and monensin could interfere with
endosomes and with the processing of endocytosed contents, this was a
likely site of action. Experiments were done to determine whether PMA
also affected the processing of extracellular fluids. When the accumul
ation of horseradish peroxidase (HRP) was measured, the rate was found
to be higher in PMA-treated cells from 5 min, the earliest time assay
ed, onward. The results suggest that the transformed type of cell in t
hese cell lines showed a constitutive dilation and/or reorganization o
f some portion of the endosomal pathway. (C) 1996 Wiley-Liss, Inc.