| DICTYOSTELIUM DISCOIDEUM CELLS are amoebae which divide asexually until they exhaust their bacterial food supply. Upon starvation, thousands of these amoeboid cells aggregate by chemotaxis to form a multicellular mass called a mound, which typically contains anywhere from 100,000 to 1,000,000 amoebae. The movement of cells within the mound is the focus of our research. This multicellular mass later elongates to form a slug, which ultimately differentiates into a fruiting body composed of two cell types: stalks and spores. In favorable conditions, a spore germinates to give rise to an amoeba, completing the life cycle. |
Dictyostelium myosin
II
| Myosin II is a protein found in muscle cells which plays a key role in muscle-cell contraction. Myosin II is also found in non-muscle cells, and in many of these it may play a role in cell locomotion. To understand its role in cell movement in Dictyostelium, we have examined cell motility and signaling wave patterns in mounds of three different myosin II mutants. We have observed that the vigorous, directed circular motion of cells is strikingly affected in each of the myosin mutants. We have also observed that some mutants affect the pattern of signaling waves. To understand myosin II's role in directed circular motion in the mound, we have examined the distribution of a fluorescent myosin II (GFP-myosin) in cells executing circular motion. We find that the myosin II distribution frequently undergoes a cycle of "C"-to-spot transitions at the cell's posterior, suggesting that myosin II may be involved in rear retraction. Learn more about the role of myosin II in Dictyostelium with our QuickTime movies... |
Dictyostelium lagC-null / gbf-null mutants
 Two genes required for normal
development in Dictyostelium are lagC and gbf, where
lagC codes for a putative cell surface protein and gbf codes
for a transcription factor. When either gene is absent, development in
Dictyostelium typically fails to proceed beyond the mound stage.
Instead of forming a slug or fruiting body, the mounds disperse, then later
reform, and then again disperse, and so on. Slugs and fruiting bodies eventually
form only in a small fraction of cells lacking lagC. (See figure
at right.) To understand why the mounds fail to proceed through development,
we have examined both cell motion and signaling wave patterns in these
mounds. (QuickTime movies of this are available.) We have found that the
mutant mounds exhibit both abnormal motion and aberrant signaling wave
patterns, and that treatment with adenosine largely rescues the aberrant
wave patterns but not the abnormal cell motion. (QuickTime movies of this
are available.) Learn
more about lagC-null and gbf-null with our QuickTime movies... |
Dictyostelium prestalk / prespore cell differentiation
| The Dictyostelium fruiting body is composed of two cell types: stalks and spores.
Precursors to these cells, called prestalk and prespore, are already present in the
mound. Initially these prestalk and prespore cells are distributed randomly, but then
they organize to form a pattern: prestalk cells cluster in the mound's tip, and prespore
cells are found beneath them. This segregation of cell types into two distinct populations
occurs during development in many organisms. Such “cell sorting” is thought to occur
by one of two general mechanisms. In one model, cells move directionally in response to
guidance signals that are used to direct cells to a particular location. In the alternate model, cells move randomly and form increasingly larger clusters based on chance contacts with like cells of similar adhesive preferences. These two sorting models make distinct predictions about how cells move during the segregation process. To test these models in Dictyostelium, we have examined cell movement of prestalk and prespore cells during mound development. We have found that prestalk cells exhibit directional movement, supporting a model in which guidance signals are important for proper cell sorting. We have also found that prestalk cells move independently and do not
form small clusters as predicted by models in which adhesion mediates the sorting process. Our in vivo observations of cell sorting have also demonstrated that prestalk cells cluster first elsewhere in the mound, and then also move directionally as a unit to the mound’s apex.
See online movies. |
Dictyostelium Quantification of Motion
| Using time-lapse 3D optical-sectioning microscopy, we have undertaken a systematic analysis of individual cell movements within a developing cell mass, the Dictyostelium mound. We examined in detail two laboratory strains. Mounds from one strain (AX-2) tended to form a standing slug that fruited directly, while mounds from the other strain (KAX-3) tended to form a reclining slug that migrated before fruiting. Under standard conditions, we found very different kinds of cell movement in mounds of the two strains. In AX-2 mounds, cells moved slowly with trajectories that were primarily random or radially inward and upward. In contrast, in KAX-3 mounds, cells moved 5x faster, with trajectories that were random in the early mound, and circular in the late mound. We showed that these apparent strain differences in directional motion were actually correlated with the mode of development, as well as the strain. By altering conditions, we induced predominantly standing slugs that fruited directly in KAX-3 and predominantly reclining slugs that migrated before fruiting in AX-2, and we also observed a corresponding switch in motile behavior: KAX-3 motion was now random and radial whereas AX-2 motion was now rotational. Thus for these strains, directional motion predicts the formation of a standing slug that fruits vs. a reclining slug that migrates. To study the regulation of cell movement, we also made AX-2/KAX-3 chimeric mounds. We found that a minority of KAX-3 cells (<10%) could induce rapid rotational movement in all cells in the mound, implying that the KAX-3 cells could non-autonomously regulate the movement of the AX-2 cells. Although the predominant directional cell motions in each strain were consistent with chemotactic response to the dark-field waves, we also found small but significant subpopulations of cells that moved differently, suggesting that in addition to the dark-field wave signal there are other key factors regulating cell motion in mounds. More to come... |