Limits...
Fluctuations of intracellular forces during cell protrusion.

Ji L, Lim J, Danuser G - Nat. Cell Biol. (2008)

Bottom Line: Surprisingly, the maxima in adhesion and boundary forces lag behind maximal edge advancement by about 40 s.Maximal F-actin assembly was observed about 20 s after maximal edge advancement.On the basis of these findings, we propose that protrusion events are limited by membrane tension and that the characteristic duration of a protrusion cycle is determined by the efficiency in reinforcing F-actin assembly and adhesion formation as tension increases.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.

ABSTRACT
We present a model to estimate intracellular force variations from live-cell images of actin filament (F-actin) flow during protrusion-retraction cycles of epithelial cells in a wound healing response. To establish a mechanistic relationship between force development and cytoskelal dynamics, force fluctuations were correlated with fluctuations in F-actin turnover, flow and F-actin-vinculin coupling. Our analyses suggest that force transmission at focal adhesions requires binding of vinculin to F-actin and integrin (indirectly), which is modulated at the vinculin-integrin but not the vinculin-F-actin interface. Force transmission at focal adhesions is colocalized in space and synchronized in time with transient increases in the boundary force at the cell edge. Surprisingly, the maxima in adhesion and boundary forces lag behind maximal edge advancement by about 40 s. Maximal F-actin assembly was observed about 20 s after maximal edge advancement. On the basis of these findings, we propose that protrusion events are limited by membrane tension and that the characteristic duration of a protrusion cycle is determined by the efficiency in reinforcing F-actin assembly and adhesion formation as tension increases.

Show MeSH

Related in: MedlinePlus

Coordination of predicted force transients during cell protrusion with F-actin assembly and edge movement. (a)Left panel: Boundary forces (cyan vectors) and adhesion force magnitudes (color-coded) at the leading edge of a protruding epithelial cell. 24 probing windows (overlaid boxes) were used to construct the activity maps shown in b. Zoom in: Predicted boundary (cyan) and adhesion (red) force vectors, and F-actin flow vectors (yellow) overlaid to the adhesion force magnitude (color-coded) in protruding sector of the cell edge. Right panel: Time montage of rates of F-actin polymerization (red) and depolymerization (green, top row); and of adhesion force magnitude (color-coded) and boundary forces (cyan vectors, bottom row) during a protrusion event. The two time series are shifted by 20 sec to account the delay in predicted adhesion/boundary force transients relative to rate changes in F-actin turnover (see text; see Video 4 for time-resolved force and assembly maps). (b) Activity maps of (I) cell edge movement, (II) velocity of F-actin retrograde flow, (III) predicted adhesion force, (IV) predicted boundary force, (V) rate of F-actin polymerization and depolymerization, and (VI) power (boundary force times the sum of the speeds of cell edge protrusion and retrograde flow). (c) Cross-correlation between activities. Negative time lags indicate that the first activity is delayed relative to the second activity. (d) Event sequence during a protrusion cycle. Scale bar in a: 5 μm.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2597050&req=5

Figure 5: Coordination of predicted force transients during cell protrusion with F-actin assembly and edge movement. (a)Left panel: Boundary forces (cyan vectors) and adhesion force magnitudes (color-coded) at the leading edge of a protruding epithelial cell. 24 probing windows (overlaid boxes) were used to construct the activity maps shown in b. Zoom in: Predicted boundary (cyan) and adhesion (red) force vectors, and F-actin flow vectors (yellow) overlaid to the adhesion force magnitude (color-coded) in protruding sector of the cell edge. Right panel: Time montage of rates of F-actin polymerization (red) and depolymerization (green, top row); and of adhesion force magnitude (color-coded) and boundary forces (cyan vectors, bottom row) during a protrusion event. The two time series are shifted by 20 sec to account the delay in predicted adhesion/boundary force transients relative to rate changes in F-actin turnover (see text; see Video 4 for time-resolved force and assembly maps). (b) Activity maps of (I) cell edge movement, (II) velocity of F-actin retrograde flow, (III) predicted adhesion force, (IV) predicted boundary force, (V) rate of F-actin polymerization and depolymerization, and (VI) power (boundary force times the sum of the speeds of cell edge protrusion and retrograde flow). (c) Cross-correlation between activities. Negative time lags indicate that the first activity is delayed relative to the second activity. (d) Event sequence during a protrusion cycle. Scale bar in a: 5 μm.

Mentions: Next, we applied spatiotemporal cross-correlation analysis to identify the timing between predicted boundary/adhesion forces, velocities of cell edge movement9; and rates of F-actin assembly/disassembly23. For each of these parameters an activity map was constructed, in this example using 24 probing windows (Fig. 5a). The cell exhibits a burst of forward motion in a ∼5 μm-wide sector of the leading edge, while remaining stationary or undergoing slow retraction in other sectors (Video 4). The analyses confirmed that boundary and adhesion forces are co-localized and synchronized (Fig. 5a-left, b-III/IV and c-i). Visual inspection of time-lapse sequences of adhesion/boundary force maps and maps of F-actin assembly/disassembly suggested that the forward movement of the cell edge was accompanied by transient increases in force and F-actin assembly (Fig. 5a-right). Cross-correlation analysis revealed that the increase in boundary/adhesion force lags the increase in F-actin assembly by ∼20 sec (Fig. 5a-right, b-III/IV/V and c-ii, Video 4). Variations in assembly/disassembly rates lag behind the corresponding variations in protrusion/retraction velocities also by 20 sec (Fig. 5b-I/V and c-iii). Thus, boundary force increases should be delayed by 40 sec relative to the corresponding protrusion event, as confirmed by cross-correlation (Fig. 5b-I/IV and c-iv).


Fluctuations of intracellular forces during cell protrusion.

Ji L, Lim J, Danuser G - Nat. Cell Biol. (2008)

Coordination of predicted force transients during cell protrusion with F-actin assembly and edge movement. (a)Left panel: Boundary forces (cyan vectors) and adhesion force magnitudes (color-coded) at the leading edge of a protruding epithelial cell. 24 probing windows (overlaid boxes) were used to construct the activity maps shown in b. Zoom in: Predicted boundary (cyan) and adhesion (red) force vectors, and F-actin flow vectors (yellow) overlaid to the adhesion force magnitude (color-coded) in protruding sector of the cell edge. Right panel: Time montage of rates of F-actin polymerization (red) and depolymerization (green, top row); and of adhesion force magnitude (color-coded) and boundary forces (cyan vectors, bottom row) during a protrusion event. The two time series are shifted by 20 sec to account the delay in predicted adhesion/boundary force transients relative to rate changes in F-actin turnover (see text; see Video 4 for time-resolved force and assembly maps). (b) Activity maps of (I) cell edge movement, (II) velocity of F-actin retrograde flow, (III) predicted adhesion force, (IV) predicted boundary force, (V) rate of F-actin polymerization and depolymerization, and (VI) power (boundary force times the sum of the speeds of cell edge protrusion and retrograde flow). (c) Cross-correlation between activities. Negative time lags indicate that the first activity is delayed relative to the second activity. (d) Event sequence during a protrusion cycle. Scale bar in a: 5 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2597050&req=5

Figure 5: Coordination of predicted force transients during cell protrusion with F-actin assembly and edge movement. (a)Left panel: Boundary forces (cyan vectors) and adhesion force magnitudes (color-coded) at the leading edge of a protruding epithelial cell. 24 probing windows (overlaid boxes) were used to construct the activity maps shown in b. Zoom in: Predicted boundary (cyan) and adhesion (red) force vectors, and F-actin flow vectors (yellow) overlaid to the adhesion force magnitude (color-coded) in protruding sector of the cell edge. Right panel: Time montage of rates of F-actin polymerization (red) and depolymerization (green, top row); and of adhesion force magnitude (color-coded) and boundary forces (cyan vectors, bottom row) during a protrusion event. The two time series are shifted by 20 sec to account the delay in predicted adhesion/boundary force transients relative to rate changes in F-actin turnover (see text; see Video 4 for time-resolved force and assembly maps). (b) Activity maps of (I) cell edge movement, (II) velocity of F-actin retrograde flow, (III) predicted adhesion force, (IV) predicted boundary force, (V) rate of F-actin polymerization and depolymerization, and (VI) power (boundary force times the sum of the speeds of cell edge protrusion and retrograde flow). (c) Cross-correlation between activities. Negative time lags indicate that the first activity is delayed relative to the second activity. (d) Event sequence during a protrusion cycle. Scale bar in a: 5 μm.
Mentions: Next, we applied spatiotemporal cross-correlation analysis to identify the timing between predicted boundary/adhesion forces, velocities of cell edge movement9; and rates of F-actin assembly/disassembly23. For each of these parameters an activity map was constructed, in this example using 24 probing windows (Fig. 5a). The cell exhibits a burst of forward motion in a ∼5 μm-wide sector of the leading edge, while remaining stationary or undergoing slow retraction in other sectors (Video 4). The analyses confirmed that boundary and adhesion forces are co-localized and synchronized (Fig. 5a-left, b-III/IV and c-i). Visual inspection of time-lapse sequences of adhesion/boundary force maps and maps of F-actin assembly/disassembly suggested that the forward movement of the cell edge was accompanied by transient increases in force and F-actin assembly (Fig. 5a-right). Cross-correlation analysis revealed that the increase in boundary/adhesion force lags the increase in F-actin assembly by ∼20 sec (Fig. 5a-right, b-III/IV/V and c-ii, Video 4). Variations in assembly/disassembly rates lag behind the corresponding variations in protrusion/retraction velocities also by 20 sec (Fig. 5b-I/V and c-iii). Thus, boundary force increases should be delayed by 40 sec relative to the corresponding protrusion event, as confirmed by cross-correlation (Fig. 5b-I/IV and c-iv).

Bottom Line: Surprisingly, the maxima in adhesion and boundary forces lag behind maximal edge advancement by about 40 s.Maximal F-actin assembly was observed about 20 s after maximal edge advancement.On the basis of these findings, we propose that protrusion events are limited by membrane tension and that the characteristic duration of a protrusion cycle is determined by the efficiency in reinforcing F-actin assembly and adhesion formation as tension increases.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.

ABSTRACT
We present a model to estimate intracellular force variations from live-cell images of actin filament (F-actin) flow during protrusion-retraction cycles of epithelial cells in a wound healing response. To establish a mechanistic relationship between force development and cytoskelal dynamics, force fluctuations were correlated with fluctuations in F-actin turnover, flow and F-actin-vinculin coupling. Our analyses suggest that force transmission at focal adhesions requires binding of vinculin to F-actin and integrin (indirectly), which is modulated at the vinculin-integrin but not the vinculin-F-actin interface. Force transmission at focal adhesions is colocalized in space and synchronized in time with transient increases in the boundary force at the cell edge. Surprisingly, the maxima in adhesion and boundary forces lag behind maximal edge advancement by about 40 s. Maximal F-actin assembly was observed about 20 s after maximal edge advancement. On the basis of these findings, we propose that protrusion events are limited by membrane tension and that the characteristic duration of a protrusion cycle is determined by the efficiency in reinforcing F-actin assembly and adhesion formation as tension increases.

Show MeSH
Related in: MedlinePlus