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Biophys J, October 1999, p. 1992-2002, Vol. 77, No. 4
Department of Cell Biology, Duke University Medical Center, Durham, North Carolina 27710 USA
When membrane-attached beads are pulled vertically by a
laser tweezers, a membrane tube of constant diameter (tether) is
formed. We found that the force on the bead (tether force) did not
depend on tether length over a wide range of tether lengths, which
indicates that a previously unidentified reservoir of membrane and not
stretch of the plasma membrane provides the tether membrane. Plots of tether force vs. tether length have an initial phase, an elongation phase, and an exponential phase. During the major elongation phase, tether force is constant, buffered by the "membrane reservoir." Finally, there is an abrupt exponential rise in force that brings the
tether out of the trap, indicating depletion of the membrane reservoir.
In chick embryo fibroblasts and 3T3 fibroblasts, the maximum tether
lengths that can be pulled at a velocity of 4 µm/s are 5.1 ± 0.3 and 5.0 ± 0.2 µm, respectively. To examine the importance of the actin cytoskeleton, we treated cells with cytochalasin B or D
and found that the tether lengths increased dramatically to 13.8 ± 0.8 and 12.0 ± 0.7 µm, respectively. Similarly, treatment of
the cells with colchicine and nocodazole results in more than a twofold
increase in tether length. We found that elevation of membrane tension
(through osmotic pressure, a long-term elevation of tether force, or a
number of transitory increases) increased reservoir size over the whole
cell. Using a tracking system to hold tether force on the bead constant
near its maximal length in the exponential phase, the rate of
elongation of the tethers was measured as a function of tether force
(membrane tension). The rate of elongation of tethers was linearly
dependent on the tether force and reflected an increase in size of the
reservoir. Increases in the reservoir caused by tension increases on
one side of the cell caused increases in reservoir size on the other side of the cell. Thus, we suggest that cells maintain a plasma membrane reservoir to buffer against changes in membrane tension and
that the reservoir is increased with membrane tension or disruption of
the cytoskeleton.
Biophys J, October 1999, p. 1992-2002, Vol. 77, No. 4
© 1999 by the Biophysical Society 0006-3495/99/10/1992/11 $2.00
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