First published online September 2, 2003
doi: 10.1242/10.1242/jcs.00696
Shared, unique and redundant functions of three members of the class I myosins (MyoA, MyoB and MyoF) in motility and chemotaxis in Dictyostelium
David L. Falk1,
Deborah Wessels1,
Leslie Jenkins1,
Tien Pham1,
Spencer Kuhl1,
Margaret A. Titus2 and
David R. Soll1,*
1 W. M. Keck Dynamic Image Analysis Facility, Department of Biological Sciences,
The University of Iowa, Iowa City, IA 52242, USA
2 Department of Genetics, Cell Biology and Development, The University of
Minnesota, Minneapolis, MN 55455, USA
View larger version (98K):
[in a new window]
|
Fig. 1. Dictyostelium amoebae respond in a unique fashion to four phases
of the natural wave of cAMP (16, 17, 26, 31). (A) The spatial, temporal and
concentration components of a relayed wave of cAMP in a natural aggregation
territory. (B) The protocols for testing the behavior of mutant cells in
buffer (i.e., `basic motile behavior') and their responses to the different
spatial, temporal and concentration components of a wave.
|
|
View larger version (41K):
[in a new window]
|
Fig. 2. All mutant cells translocate with reduced velocity, as revealed by
perimeter tracks of representative control and mutant cells translocating on a
glass surface while being perfused with buffer. The red-coded image represents
the last cell image in each perimeter track. The interval time between images
was 8 seconds, and the total time of analysis, 10 minutes for each cell.
|
|
View larger version (33K):
[in a new window]
|
Fig. 3. The abnormal hemispherical shapes of myoF and
myoA/myoF cells translocating in buffer revealed in 3D
direct image reconstructions using 3D-DIAS software. (A) Representative
control JH10 cell. (B) Representative myoF cell. (C)
Representative myoA/myoF cell. Pseudopods were determined as
regions lacking particulate cytoplasm and are color-coded red. Time is noted
in seconds (s).
|
|
View larger version (35K):
[in a new window]
|
Fig. 4. Cells of all mutant strains are less efficient chemotactically than their
relevant control cells. Histograms of (A) myoA cell and nh6b control
cells; (B) myoB, myoA/myoB and KAX3 control cells; (C)
myoF, myoA/myoF and JH10 control cells. Arrows refer to
average chemotactic indices of respective strains.
|
|
View larger version (52K):
[in a new window]
|
Fig. 5. Cells of all mutant strains react chemotactically in a spatial gradient of
cAMP less efficiently and with reduced velocity, as revealed by perimeter
tracks of representative control and mutant cells. The arrow at the bottom of
each panel represents the direction of the increasing gradient of cAMP. The
red-coded image represents the last in a perimeter track. The interval time
between images is 8 seconds.
|
|
View larger version (49K):
[in a new window]
|
Fig. 6. All control strains responded normally to a series of four temporal waves
of cAMP generated in the absence of spatial gradients of cAMP. In the first of
each set of panels, the mean instantaneous velocity is plotted as a function
of time for 10 cells treated with four successive waves of cAMP. The velocity
surge occurs in each case in the front of the wave and ends as the wave
reaches its peak (dashed lines). In the second two of each set of panels,
difference pictures were generated during the front of the wave (phase B) and
during the peak and back of the wave (phases C and D) for representative
cells. The difference pictures for the peak of the wave (phase C) span the
first four and those for the back of the wave (phase D) span the last eight of
each set of difference pictures under the heading `Phase C and D'. Green
represents expansion zones and red contraction zones in the difference
pictures.
|
|
View larger version (60K):
[in a new window]
|
Fig. 7. Defects in responses to temporal waves were exhibited by myoB,
myoA/myoB and myoA/myoF mutant cells. The
velocity of mutant cells is plotted as a function of a series of four temporal
waves of cAMP generated in the absence of spatial gradients of cAMP. For an
explanation of panels and reconstructions, see legend to
Fig. 6.
|
|
View larger version (56K):
[in a new window]
|
Fig. 8. Vector flow analysis of monolayers of control and mutant cells in natural
aggregation territories. For each strain, a window of several hundred cells
was monitored in a monolayer after 6 hours of incubation. In the vector flow
plots, the magnitude of the vector components in the selected direction of the
aggregation center were averaged and plotted over time for one control strain
(nh6b), and mutant strains myoA, myoF and
myoA/myoF. Because myoA/myoF cells did not
aggregate, an arbitrary direction was selected. The x-axis represents
time, and the y-axis the direction and extent of cell displacement.
Displacement towards the aggregation center is positive and displacement away
from the aggregation is negative. Note that while myoA and
myoF cells initiate cyclic surges towards the aggregation center
(positive displacement) in a manner similar to control cells,
myoA/myoF cells do not initiate cyclic surges in any
consistent direction.
|
|
View larger version (17K):
[in a new window]
|
Fig. 9. myoA/myoF cells do not show any chemotaxis in natural
waves of cAMP generated by control cells in mixtures.
myoA/myoF cells were mixed with control (JH10) cells at a
ratio of 1:9, respectively. After 8 hours, cells were motion analyzed. (A)
Instantaneous velocity plots of a representative control and neighboring
mutant cell. (B) Centroid tracks of a control cell and a neighboring
myoA/myoF cell. The phases of the wave are noted along the
tracks. The small arrow denotes direction of control cell.
|
|
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Reddit 
Technorati 
Twitter What's this?
© The Company of Biologists Ltd 2003
