Fig. 4. The relationship in time between BRG1 expression and the appearance of
stress-fibre-like actin filaments. (A) Immunoblots of cell lysates (20
µg/sample) of cells transfected with the pBJ5-BRG1 expression vector
(indicated above the lanes) or the pBJ5-BRG1-K798R vector (indicated above the
lanes) taken at the time points indicated above the lanes. The lane marked `0'
is untransfected SW13 cells lysed just prior to transfection. The
-tubulin levels in the same samples are shown as a control. (B) SW13
cells were transfected with pBJ5-BRG1 or the ATPase-deficient pBJ5-BRG1-K798R
on glass coverslips and samples were taken at 24, 48 and 72 hours. The cells
were double-stained with FITC-phalloidin and anti-BRG1 antiserum visualised by
TRITC-secondary antibodies after fixation. Between 30 and 50 cells expressing
the BRG1 protein or the mutated BRG1-K798R protein in the nucleus were counted
blindly in each separate experiment for each group (four separate experiments
for 24 hours and 48 hours). The percentage of BRG1/mutated
BRG1-K798R-expressing cells that had formed thick actin bundles in the cell
body was determined for each sample. The standard deviation is given for each
group; n is the number of separate experiments in the group. The two
values for the 72 hour point were 65 and 72% for BRG1-expressing cells, and 3
and 13% for BRG1-K798R-expressing cells. Immunofluorescence images of cells
transfected with the BRG1-vector stained with phalloidin (C) and with the
BRG-antiserum against the N-terminal (D) 24 hours after transfection and
stained with phalloidin (E) and the anti-BRG1 antiserum as in D (F), 48 hours
after transfection are shown. Bar, 20 µm.
