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Regulated actin cytoskeleton assembly at filopodium tips controls their extension and retraction.

Mallavarapu A, Mitchison T - J. Cell Biol. (1999)

Bottom Line: We sought to understand how the dynamic behavior of the actin cytoskeleton is regulated to produce extension or retraction.Both assembly and flow rate can vary with time in a single filopodium and between filopodia in a single growth cone.Regulation of assembly rate is the dominant factor in controlling filopodia behavior in our system.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Harvard University Medical School, Boston, Massachusetts 02115, USA.

ABSTRACT
The extension and retraction of filopodia in response to extracellular cues is thought to be an important initial step that determines the direction of growth cone advance. We sought to understand how the dynamic behavior of the actin cytoskeleton is regulated to produce extension or retraction. By observing the movement of fiduciary marks on actin filaments in growth cones of a neuroblastoma cell line, we found that filopodium extension and retraction are governed by a balance between the rate of actin cytoskeleton assembly at the tip and retrograde flow. Both assembly and flow rate can vary with time in a single filopodium and between filopodia in a single growth cone. Regulation of assembly rate is the dominant factor in controlling filopodia behavior in our system.

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Average assembly and flow rates grouped by rates of filopodium movement. Tip movement, assembly, and flow rates were determined for individual bouts of filopodia movement for all observations, and arbitrarily grouped on the basis of tip movement as follows. Rapidly retracting: retracting at greater than −1 μm/min. Slowly retracting: between −1 and −0.3 μm/min. Stationary: between −0.3 and +0.3 μm/min. Slowly extending: between +0.3 and +1 μm/min. Rapidly extending: extending at any rate above +1 μm/min. Error bars are SD of the mean values in each group. For analysis, the rates of tip movement, assembly, and flow were measured between each time point (every 30 s) for each filopodium and grouped. Different periods of movement of a single filopodium might contribute to several different categories. The plot pools 587 individual time point measurements taken from 75 separate filopodia from 18 different sequences. 14 were Q-rhodamine photoactivation experiments and 4 were GFP-photobleaching experiments. We were unable to detect differences between these two groups of data examined separately.
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Figure 5: Average assembly and flow rates grouped by rates of filopodium movement. Tip movement, assembly, and flow rates were determined for individual bouts of filopodia movement for all observations, and arbitrarily grouped on the basis of tip movement as follows. Rapidly retracting: retracting at greater than −1 μm/min. Slowly retracting: between −1 and −0.3 μm/min. Stationary: between −0.3 and +0.3 μm/min. Slowly extending: between +0.3 and +1 μm/min. Rapidly extending: extending at any rate above +1 μm/min. Error bars are SD of the mean values in each group. For analysis, the rates of tip movement, assembly, and flow were measured between each time point (every 30 s) for each filopodium and grouped. Different periods of movement of a single filopodium might contribute to several different categories. The plot pools 587 individual time point measurements taken from 75 separate filopodia from 18 different sequences. 14 were Q-rhodamine photoactivation experiments and 4 were GFP-photobleaching experiments. We were unable to detect differences between these two groups of data examined separately.

Mentions: The results presented above demonstrate that cytoskeleton assembly and retrograde flow rates can vary within a single filopodium over time, and that the two parameters can be regulated independently. However, the cases where the filopodium tip switched from one consistent direction of movement to another during the experiment represent only a subset (19/75) of the filopodia studied. In most cases, the filopodium under observation extended or retracted continuously, though the rate might fluctuate. To summarize the contribution of assembly and flow rates to the direction and rate of filopodium tip movement for all our experiments, we grouped all our data by filopodium tip velocity and calculated average assembly and flow rates for each group (Fig. 5). An approximately linear relationship can be discerned between average actin assembly rate and tip velocity. In contrast, average flow rate was relatively constant for different tip velocities. The only exception was a significant change in flow rate to more negative values in rapidly retracting filopodia. Thus, on average for the data collected in this study, most of the variations in filopodium motility could be attributed to differences in the rates of actin cytoskeleton assembly while the flow rate was relatively constant.


Regulated actin cytoskeleton assembly at filopodium tips controls their extension and retraction.

Mallavarapu A, Mitchison T - J. Cell Biol. (1999)

Average assembly and flow rates grouped by rates of filopodium movement. Tip movement, assembly, and flow rates were determined for individual bouts of filopodia movement for all observations, and arbitrarily grouped on the basis of tip movement as follows. Rapidly retracting: retracting at greater than −1 μm/min. Slowly retracting: between −1 and −0.3 μm/min. Stationary: between −0.3 and +0.3 μm/min. Slowly extending: between +0.3 and +1 μm/min. Rapidly extending: extending at any rate above +1 μm/min. Error bars are SD of the mean values in each group. For analysis, the rates of tip movement, assembly, and flow were measured between each time point (every 30 s) for each filopodium and grouped. Different periods of movement of a single filopodium might contribute to several different categories. The plot pools 587 individual time point measurements taken from 75 separate filopodia from 18 different sequences. 14 were Q-rhodamine photoactivation experiments and 4 were GFP-photobleaching experiments. We were unable to detect differences between these two groups of data examined separately.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2169471&req=5

Figure 5: Average assembly and flow rates grouped by rates of filopodium movement. Tip movement, assembly, and flow rates were determined for individual bouts of filopodia movement for all observations, and arbitrarily grouped on the basis of tip movement as follows. Rapidly retracting: retracting at greater than −1 μm/min. Slowly retracting: between −1 and −0.3 μm/min. Stationary: between −0.3 and +0.3 μm/min. Slowly extending: between +0.3 and +1 μm/min. Rapidly extending: extending at any rate above +1 μm/min. Error bars are SD of the mean values in each group. For analysis, the rates of tip movement, assembly, and flow were measured between each time point (every 30 s) for each filopodium and grouped. Different periods of movement of a single filopodium might contribute to several different categories. The plot pools 587 individual time point measurements taken from 75 separate filopodia from 18 different sequences. 14 were Q-rhodamine photoactivation experiments and 4 were GFP-photobleaching experiments. We were unable to detect differences between these two groups of data examined separately.
Mentions: The results presented above demonstrate that cytoskeleton assembly and retrograde flow rates can vary within a single filopodium over time, and that the two parameters can be regulated independently. However, the cases where the filopodium tip switched from one consistent direction of movement to another during the experiment represent only a subset (19/75) of the filopodia studied. In most cases, the filopodium under observation extended or retracted continuously, though the rate might fluctuate. To summarize the contribution of assembly and flow rates to the direction and rate of filopodium tip movement for all our experiments, we grouped all our data by filopodium tip velocity and calculated average assembly and flow rates for each group (Fig. 5). An approximately linear relationship can be discerned between average actin assembly rate and tip velocity. In contrast, average flow rate was relatively constant for different tip velocities. The only exception was a significant change in flow rate to more negative values in rapidly retracting filopodia. Thus, on average for the data collected in this study, most of the variations in filopodium motility could be attributed to differences in the rates of actin cytoskeleton assembly while the flow rate was relatively constant.

Bottom Line: We sought to understand how the dynamic behavior of the actin cytoskeleton is regulated to produce extension or retraction.Both assembly and flow rate can vary with time in a single filopodium and between filopodia in a single growth cone.Regulation of assembly rate is the dominant factor in controlling filopodia behavior in our system.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Harvard University Medical School, Boston, Massachusetts 02115, USA.

ABSTRACT
The extension and retraction of filopodia in response to extracellular cues is thought to be an important initial step that determines the direction of growth cone advance. We sought to understand how the dynamic behavior of the actin cytoskeleton is regulated to produce extension or retraction. By observing the movement of fiduciary marks on actin filaments in growth cones of a neuroblastoma cell line, we found that filopodium extension and retraction are governed by a balance between the rate of actin cytoskeleton assembly at the tip and retrograde flow. Both assembly and flow rate can vary with time in a single filopodium and between filopodia in a single growth cone. Regulation of assembly rate is the dominant factor in controlling filopodia behavior in our system.

Show MeSH
Related in: MedlinePlus