The life cycle of dictyostelium

Another type of multicellular organization derived from unicellular organisms is found in Dictyostelium discoideum.^* The life cycle of this fascinating organism is illustrated in Figure 2.17A. In its asexual cycle, solitary haploid amoebae (called myxamoebae or “social amoebae” to distinguish them from amoeba species that always remain solitary) live on decaying logs, eating bacteria and reproducing by binary fission. When they have exhausted their food supply, tens of thousands of these myxamoebae join together to form moving streams of cells that converge at a central point. Here they pile atop one another to produce a conical mound called a tight aggregate. Subsequently, a tip arises at the top of this mound, and the tight aggregate bends over to produce the migrating slug (with the tip at the front). The slug (often given the more dignified title of pseudoplasmodium or grex) is usually 2–4 mm long and is encased in a slimy sheath. The grex begins to migrate (if the environment is dark and moist) with its anterior tip slightly raised. When it reaches an illuminated area, migration ceases, and the grex differentiates into a fruiting body composed of spore cells and a stalk. The anterior cells, representing 15–20% of the entire cellular population, form the tubed stalk. This process begins as some of the central anterior cells, the prestalk cells, begin secreting an extracellular coat and extending a tube through the grex. As the prestalk cells differentiate, they form vacuoles and enlarge, lifting up the mass of prespore cells that had made up the posterior four-fifths of the grex (Jermyn and Williams 1991). The stalk cells die, but the prespore cells, elevated above the stalk, become spore cells. These spore cells disperse, each one becoming a new myxamoeba.

Figure 2.17

Life history of Dictyostelium discoideum. Haploid spores give rise to myxamoebae, which can reproduce asexually to form more haploid myxamoebae. As the food supply diminishes, aggregation occurs at central points, and a migrating pseudoplasmodium is formed. (more...)

In addition to this asexual cycle, there is a possibility for sex in Dictyostelium. Two myxamoebae can fuse to create a giant cell, which digests all the other cells of the aggregate. When it has eaten all its neighbors, it encysts itself in a thick wall and undergoes meiotic and mitotic divisions; eventually, new myxamoebae are liberated.

Dictyostelium has been a wonderful experimental organism for developmental biologists because initially identical cells are differentiated into one of two alternative cell types, spore and stalk. It is also an organism wherein individual cells come together to form a cohesive structure composed of differentiated cell types, akin to tissue formation in more complex organisms. The aggregation of thousands of myxamoebae into a single organism is an incredible feat of organization that invites experimentation to answer questions about the mechanisms involved.

Aggregation of dictyostelium cells

The first question is, What causes the myxamoebae to aggregate? Time-lapse microcinematography has shown that no directed movement occurs during the first 4–5 hours following nutrient starvation. During the next 5 hours, however, the cells can be seen moving at about 20 μm/min for 100 seconds. This movement ceases for about 4 minutes, then resumes. Although the movement is directed toward a central point, it is not a simple radial movement. Rather, cells join with one another to form streams; the streams converge into larger streams, and eventually all streams merge at the center. Bonner (1947) and Shaffer (1953) showed that this movement is due to chemotaxis: the cells are guided to aggregation centers by a soluble substance. This substance was later identified as cyclic adenosine 3´,5´-monophosphate (cAMP) (Konijn et al. 1967; Bonner et al. 1969), the chemical structure of which is shown in Figure 2.18A.

Figure 2.18

Chemotaxis of Dictyostelium myxamoebae due to spiral waves of cAMP. (A) Chemical structure of cAMP. (B) Visualization of several cAMP “waves.” Central cells secrete cAMP at regular intervals, and each pulse diffuses outward as a concentric (more...)

Aggregation is initiated as each of the cells begins to synthesize cAMP. There are no dominant cells that begin the secretion or control the others. Rather, the sites of aggregation are determined by the distribution of myxamoebae (Keller and Segal 1970; Tyson and Murray 1989). Neighboring cells respond to cAMP in two ways: they initiate a movement toward the cAMP pulse, and they release cAMP of their own (Robertson et al. 1972; Shaffer 1975). After this, the cell is unresponsive to further cAMP pulses for several minutes. The result is a rotating spiral wave of cAMP that is propagated throughout the population of cells (Figure 2.18B-D). As each wave arrives, the cells take another step toward the center.^†

The differentiation of individual myxamoebae into either stalk (somatic) or spore (reproductive) cells is a complex matter. Raper (1940) and Bonner (1957) demonstrated that the anterior cells normally become stalk, while the remaining, posterior cells are usually destined to form spores. However, surgically removing the anterior part of the slug does not abolish its ability to form a stalk. Rather, the cells that now find themselves at the anterior end (and which originally had been destined to produce spores) now form the stalk (Raper 1940). Somehow a decision is made so that whichever cells are anterior become stalk cells and whichever are posterior become spores. This ability of cells to change their developmental fates according to their location within the whole organism and thereby compensate for missing parts is called regulation. We will see this phenomenon in many embryos, including those of mammals.

Cell adhesion molecules in dictyostelium

How do individual cells stick together to form a cohesive organism? This problem is the same one that embryonic cells face, and the solution that evolved in the protists is the same one used by embryos: developmentally regulated cell adhesion molecules.

While growing mitotically on bacteria, Dictyostelium cells do not adhere to one another. However, once cell division stops, the cells become increasingly adhesive, reaching a plateau of maximum cohesiveness around 8 hours after starvation. The initial cell-cell adhesion is mediated by a 24,000-Da (24-kDa) glycoprotein that is absent in myxamoebae but appears shortly after division ceases (Figure 2.19; Knecht et al. 1987; Loomis 1988). This protein is synthesized from newly transcribed mRNA and becomes localized in the cell membranes of the myxamoebae. If myxamoebae are treated with antibodies that bind to and mask this protein, they will not stick to one another, and all subsequent development ceases.

Figure 2.19

Dictyostelium cells synthesize an adhesive 24-kDa glycoprotein shortly after nutrient starvation. These Dictyostelium cells were stained with a fluorescently labeled antibody that binds to the 24-kDa glycoprotein and were then observed under ultraviolet (more...)

Once this initial aggregation has occurred, it is stabilized by a second cell adhesion molecule. This 80-kDa glycoprotein is also synthesized during the aggregation phase. If it is defective or absent in the cells, small slugs will form, and their fruiting bodies will be only about one-third the normal size. Thus, the second cell adhesion system seems to be needed for retaining a large enough number of cells to form large fruiting bodies (Müller and Gerisch 1978; Loomis 1988). In addition, a third cell adhesion system is activated late in development, while the slug is migrating. This protein appears to be important in the movement of the prestalk cells to the apex of the mound (Ginger et al. 1998). Thus, Dictyostelium has evolved three developmentally regulated systems of cell-cell adhesion that are necessary for the morphogenesis of individual cells into a coherent organism. As we will see in subsequent chapters, metazoan cells also use cell adhesion molecules to form the tissues and organs of the embryo.

Dictyostelium is a “part-time multicellular organism” that does not form many cell types (Kay et al. 1989), and the more complex multicellular organisms do not form by the aggregation of formerly independent cells. Nevertheless, many of the principles of development demonstrated by this “simple” organism also appear in embryos of more complex phyla (see Loomis and Insall 1999). The ability of individual cells to sense a chemical gradient (as in the myxamoeba's response to cAMP) is very important for cell migration and morphogenesis during animal development. Moreover, the role of cell surface proteins in cell cohesiveness is seen throughout the animal kingdom, and differentiation-inducing molecules are beginning to be isolated in metazoan organisms.

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Sex and Individuality in Volvox.

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Evidence and Antibodies.

Differentiation in dictyostelium

Differentiation into stalk cell or spore cell reflects another major phenomenon of embryogenesis: the cell's selection of a developmental pathway. Cells often select a particular developmental fate when alternatives are available. A particular cell in a vertebrate embryo, for instance, can become either an epidermal skin cell or a neuron. In Dictyostelium, we see a simple dichotomous decision, because only two cell types are possible. How is it that a given cell becomes a stalk cell or a spore cell? Although the details are not fully known, a cell's fate appears to be regulated by certain diffusible molecules. The two major candidates are differentiation-inducing factor (DIF) and cAMP. DIF appears to be necessary for stalk cell differentiation. This factor, like the sex-inducing factor of Volvox, is effective at very low concentrations (10^-10 M); and, like the Volvox protein, it appears to induce differentiation into a particular type of cell. When added to isolated myxamoebae or even to prespore (posterior) cells, it causes them to form stalk cells. The synthesis of this low molecular weight lipid is genetically regulated, for there are mutant strains of Dictyostelium that form only spore precursors and no stalk cells. When DIF is added to these mutant cultures, stalk cells are able to differentiate (Kay and Jermyn 1983; Morris et al. 1987), and new prestalk-specific mRNAs are seen in the cell cytoplasm (Williams et al. 1987). While the mechanisms by which DIF induces 20% of the grex cells to become stalk tissue are still controversial (see Early et al. 1995), DIF may act by releasing calcium ions from intracellular compartments within the cell (Shaulsky and Loomis 1995).

Although DIF stimulates myxamoebae to become prestalk cells, the differentiation of prespore cells is most likely controlled by the continuing pulses of cAMP. High concentrations of cAMP initiate the expression of prespore-specific mRNAs in aggregated myxamoebae. Moreover, when slugs are placed in a medium containing an enzyme that destroys extracellular cAMP, the prespore cells lose their differentiated characteristics (Figure 2.20; Schaap and van Driel 1985; Wang et al. 1988a,b).

Figure 2.20

Chemicals controlling differentiation in Dictyostelium. (A) and (B) show the effects of placing Dictyostelium slugs into a medium containing enzymes that destroy extracellular cAMP. (A) Control grex stained for the presence of a prespore-specific protein (more...)