Re: How did scientists first determine the slime mould amoebae's lifecycle?

Hi Loretta,

Thanks for your question! Before I answer your question, I think a little bit of background would be helpful.

The "cellular slime mold" you refer to in your question is also known as Dictyostelium discoideum, or affectionately as "Dicty" by those who study this organism. However, it is neither slimy nor a mold. Dicty is particularly interesting because it is a unicellular organism that has a multicellular phase in its life cycle.

The life cycle of Dictyostelium discoideum
When there is an adequate food supply, Dicty exists as single haploid cells called myxamoebae. However, once food stores are depleted, these cells aggregate together to form a migrating slug, or pseudoplasmodium. This slug migrates with its anterior, or front, end slightly raised. Once the slug has stopped migrating, the cells at the anterior tip form a tubular stalk which extends downward through the slug. As this stalk forms, it lifts the remaining cells, which originally formed the posterior part of the slug. These posterior cells become the spore cells. Together, these cells form the mature fruiting body. A fruiting body consists of a spore sac, elevated off the ground by a stalk, and supported by a "suction cup-like" structure called a basal disc that is attached to the growth surface. These fruiting bodies release spores that are dispersed in the environment, with each spore possessing the ability to become a new myxamoeba. This asexual life cycle allows Dicty to endure periods of starvation as a multicellular aggregate until the food supply is replenished.

Dicty as a model organism
As you alluded to in your question, Dicty is a fascinating experimental organism for developmental biologists. How do single cells that are initially identical coalesce into a multicellular structure composed of two different cell types, spore and stalk cells? On a fundamental level, this process is similar to more complex processes, such as tissue and organ formation, in higher organisms. Thus, studies of Dicty developmental may help us understand not only how slime molds develop, but also how more complex organisms form.

A "blast" from the past: the classical experiments
So, on to your question… How did biologists figure out which cells in the slug contribute to the different parts of the fruiting body? In the classical experiments, which were conducted in the 1940s and '50s, scientists used dyes to mark cells and follow their fate. In this case, they stained slugs with a dye called neutral red. The cells of the slug take up the dye uniformly, so all cells of the slug appear red. However, this is not particularly helpful for following distinct groups of cells because these red slugs would merely give rise to completely red fruiting bodies. So, the biologists solved this problem by taking the anterior portion of a 'red' slug and attaching it to an unstained, posterior portion from another slug. The resulting chimeric slug did not seem to mind that it had been manipulated in this way--it went on to form a fruiting body as it normally would. However, scientists observed that the stalk of the fruiting body was stained red, indicating that is was derived mainly from cells in the anterior region of the slug. Thus, by doing various permutations of the above experiment, biologists learned that spore cells and stalk cells arise from the posterior and anterior portions, respectively, of the migrating slug.

Using new technology to reexamine old questions
While it is well established that anterior cells become stalk and posterior cells form spores, it is often worthwhile to repeat classical experiments using modern tools. Instead of using dyes, biologists can now use molecular tools to mark cells. One way to accomplish this is to utilize cell-specific gene expression to follow cell fate. For instance, scientists have discovered there are a number of genes expressed in cells that will contribute to the stalk or spores. These "prestalk" and "prespore" cells can be marked using a technique that visually detects mRNA for a specific gene. Therefore, in this way biologists can use gene expression as a molecular marker for specific cell populations. Prespore and prestalk cells can also be labeled by using DNA technology coupled with fluorescent proteins or enzymes that generate colored products. Collectively, the main advantage of these approaches is that they provide finer resolution--labeling cells with a dye can often be rather "sloppy". These modern tools have recapitulated what was already known from the classical studies, and also provide an opportunity to extend our knowledge of Dicty development.

I hope you find this information useful! For completeness, I have included the references for the classical Dicty experiments. However, I must admit that I did not consult these references because I could not find them at my library. Instead, I contacted a researcher in the field. But, in case you want more information, you might be able to hunt them down. I have also included a couple recent references as well as the website for the Dicty database, where you can find references galore on Dictyostelium. For more general information, you can consult any developmental biology text. Gilbert's Developmental Biology 5th edition is particularly good. Please feel free to email me with any further questions!

Nikki
nmdavis@fas.harvard.edu

classical experiments
Raper, K.B. 1940. Pseudoplasmodium formation and organization in Dictyostelium discoideum. J. Elisha Mitchell Sci. Soc. 56:241-282.

Bonner, J.T. 1957. A theory of the control of differentiation in the cellular slime molds. Q. Rev. Biol. 32:232-246.

recent publications
Hodgkinson S. 1995. GFP in Dictyostelium. Trends Genet.11:327-8

Nicol A, et al. 1999. Cell-sorting in aggregates of Dictyostelium discoideum. J. Cell Sci. 112:3923-3929.

DictyBase (formerly The Dictyostelium Virtual Library).