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First published online April 23, 2007
doi: 10.1242/10.1242/jcs.002527

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Traction force microscopy in Dictyostelium reveals distinct roles for myosin II motor and actin-crosslinking activity in polarized cell movement

¹ Department of Molecular and Cell Biology, University of Connecticut, Storrs, CT 06269, USA
² Department of Biomedical Engineering, Boston University, Boston, MA 02215, USA

View larger version (44K): [in this window] [in a new window]   Fig. 1. The motile behavior of Dictyostelium moving on gelatin. (A-C) Stacked cell outlines, overlaid with a centroid track, from three representative cells of each type; wild-type, mlcE– and mhcA– cells. Examples of cells moving randomly at fast (1), intermediate (2) and slow (3) speeds, for each cell type, over a 3-minute period are shown. (A'-C') Rose plots, showing the total distance traveled and the directionality of movement for each cell type during a 3-minute period. Bar, 5 µm. (D-F) Summary histograms of the mean relative changes in instantaneous speed (D), persistence (E) and spread area (F) with respect to the wild-type value that was set to 100% for wild-type (black bar), mlcE+ (gray bar), mlcE– (striped bar) and mhcA– cells (open bar, n=9). Asterisks indicate statistically significant differences compared with wild-type cells (P<0.05), using a Student's t-test, assuming unequal variances.

View larger version (29K): [in this window] [in a new window]   Fig. 2. Characteristic distribution of traction stress for wild-type, mlcE– and mhcA– cells. Color-coded vector maps of the traction stresses generated by wild-type (A), mlcE– (B), and mhcA– (C) cells, moving in the direction indicated (arrow). The length and orientation of arrows in the vector map represent the magnitude and direction of traction stresses. Note that the traction vector scale bar (horizontal arrow) in each map represents a traction stress that differs by approximately an order of magnitude between each cell type. Colored regions represent areas of traction stress within the same magnitude range. Areas of high (90th percentile) traction stress are represented by red and purple colors, while areas of low traction stress are shown in gray, blue and light green.

View larger version (28K): [in this window] [in a new window]   Fig. 3. Changes in distribution of traction stress generated by a wild-type Dictyostelium cell during a `rapid recoil' retraction. (A-F) A time series of traction vector maps obtained from a wild-type cell moving in the general direction indicated (arrow), corresponding to one cycle of movement. The highest 90th percentile traction stresses (red and purple regions) are located in a crescent-shaped region at the rear of the cell (A,B,C) prior to retraction (R in C) at the rear (arrow) then disappear abruptly following retraction (D-F). Regions of low traction stress (white to green regions) are consistently found at the front of the cell (A-F). Once retraction has occurred these regions allow for protrusion to begin (P in E,F). Bar, 3 µm. (G) Plots of the 90th percentile traction stress (red), instantaneous speed (green), cell area (blue) and shape factor (yellow) corresponding to panels A-F. The rapid recoil retraction (R) occurs between 9-12 seconds (C,D) as indicated by the two vertical dotted lines.

View larger version (39K): [in this window] [in a new window]   Fig. 4. Changes in distribution of traction stress generated by a wild-type Dictyostelium cell during a `slow recoil' retraction. (A-L) A time series of traction vector maps obtained from a wild-type cell moving in the general direction indicated (arrow). Traction stresses increase gradually (A-C), particularly at the rear (arrow in C), prior to retraction (R in C) and decrease during retraction (C-G). Protrusion (P in C) occurs simultaneously with retraction (C-G) at the front of cell where there are regions of low traction stresses. Bar, 3 µm. (M) Plots of the 90th percentile traction stress (red), instantaneous speed (green), cell area (blue) and shape factor (yellow) corresponding to panels A-L. A-G constitute one cycle of movement that includes the first slow recoil retraction (R) which occurs between 9 and 21 seconds, as indicated by the vertical dotted lines. H-L show the beginning of a second cycle of movement, prior to the onset of a slow recoil retraction (not shown). Note the gradual increase in traction stress at the rear and its similarity to panels A-D.

View larger version (33K): [in this window] [in a new window]   Fig. 5. Changes in distribution of traction stress generated by a mlcE– Dictyostelium cell during a `slow recoil' retraction. (A-H) A time series of traction vector maps obtained from a mlcE– cell moving in the general direction indicated (arrow), during one cycle of movement. An asymmetrical distribution of high traction stress develops very slowly at the rear and along the lateral cell edges (A,D) prior to retraction (R in E, arrow). During the slow recoil retraction, traction stress decreases gradually together with a progressive loss in front-rear asymmetry (E-H), while protrusion (P in E) is occurring at the cell front, where traction stresses are low. Bar, 3 µm. (I) Plots of the 90th percentile traction stress (red), instantaneous speed (green), cell area (blue) and shape factor (yellow) corresponding to panels A-H. The slow recoil retraction (R) occurs between 28–52 seconds, as indicated by the vertical dotted lines.

View larger version (36K): [in this window] [in a new window]   Fig. 6. Changes in distribution of traction stress generated by a mhcA– Dictyostelium cell during a `slow recoil' retraction. (A-J) A time series of traction vector maps obtained from a mhcA– cell moving in the general direction indicated (arrow), during one cycle of movement. An asymmetrical distribution of traction stress occurs slowly (A-E), but more rapidly than in mlcE– cells. Regions of highest 90th percentile traction stress (purple, red) enlarge at the rear and extend along the lateral cell edges. During retraction (R in E) at the rear (arrow) traction stresses gradually decrease, particularly at the rear (E-J), while at the same time protrusion (P in E) is occurring at the front where low tractions are present (E-J). Bar, 3 µm. (K) Plots of the 90th percentile traction stress (red), instantaneous speed (green), cell area (blue), shape factor (yellow) corresponding to panels A-J. The slow recoil retraction (R) occurs between 15 to 27 seconds, as indicated by the vertical dotted lines. Note that for illustrative purposes the color scale represents a range of magnitudes that is 25 times less than the one used for the wild-type cells in Figs 3 and 4.

View larger version (24K): [in this window] [in a new window]   Fig. 7. Diagram of the hypothetical relationship between the spatio-temporal distribution of traction stress and a cycle of movement for each cell type. For each cell type, colored areas represent the average 90th percentile traction stress that is highest for the wild-type (dark blue), intermediate for mlcE– (light blue) and very low for mhcA– cells (gray). Periods of development, loss and regeneration of traction stress asymmetry are indicated by horizontal bars. The total number of cycles occurring within a period of observation (3 minutes) is expressed as cycles per minute. Graph illustrating differences in the rate of development (blue), retraction (red) and regeneration (green) of traction stress asymmetry for the wild-type, mlcE– and mhcA– cells. The gray line represents a period in which mhcA– cells lack traction stress asymmetry, except where marked (asterisk).