Biology Dept
Kenyon College
Chapter 11.  Development: Differentiation and Determination
Fall Section Spring Section 1 Spring Section 2
Differential gene expression and development
Mechanisms of cellular determination
Pattern formation

Differential gene expression and development

The fate of a cell describes what it will become in the course of normal development. The fate of a particular cell can be discovered by labelling that cell and observing what structures it becomes a part of. When the fate of all cells of an embryo has been discovered, we can build a fate map, which is a diagram of that organism at an early stage of development that indicates the fate of each cell or region at a later stage of development.

The developmental potential, or potency, of a cell describes the range of different cell types it CAN become. The zygote and its very early descendents are totipotent - these cells have the potential to develop into a complete organism. Totipotency is common in plants, but is uncommon in animals after the 2-4 cell stage. As development proceeds, the developmental potential of individual cells decreases until their fate is determined.

The determination of different cell types (cell fates) involves progressive restrictions in their developmental potentials. When a cell “chooses” a particular fate, it is said to be determined, although it still "looks" just like its undetermined neighbors. Determination implies a stable change - the fate of determined cells does not change.

Differentiation follows determination, as the cell elaborates a cell-specific developmental program. Differentiation results in the presence of cell types that have clear-cut identities, such as muscle cells, nerve cells, and skin cells.

Differentiation results from differential gene expression: The specific components of a given cell provides its special characteristics. These components are either synthesized by proteins, or are themselves proteins. By expressing different subsets of genes, two cells contain different subsets of gene products (proteins).

How can we observe that cells from two tissues express different genes? Below are two blots: the Southern blot shows that tissues A and B both contain a particular gene. However, the Northern blots shows that only tissue A contains RNA transcribed from that particular gene.

Differential gene  expression is not a result of differential loss of the genetic material, DNA, except in the case of the immune system. That is, genetic information is not lost as cells become determined and begin to differentiate.

In fact, even the nuclei of adult cells contain ALL of the information needed for the construction of an entire organism, if provided with the proper cytoplasmic components. The cloning of Dolly from an adult cell is a major breakthrough, not only because of potential biotechnological applications, but because of the importance of this result for basic science: the result is the most convincing evidence for the theory of differential gene expression.

In order to clone Dolly, udder cells were removed from a Finn Dorset ewe and starved for one week to cause G0 arrest.  Nuclei from arrested Finn Dorset udder cells were fused with enucleated eggs from a Scottish Blackface ewe, and then stimulated to re-enter the cell cycle. After a few rounds of cell division, the embryo was transplanted into a surrogate Scottish Blackface mother. The sheep that was born was genetically identical to the Finn Dorset ewe, which was the source of the nucleus.

Transplantation of imaginal discs in insects and the cloning of whole plants from individual cells strengthens the conclusion that genetic information is not lost as cells become determined and begin to differentiate.

While differentiation results in specific cell types, morphogenesis is the process whereby the shape (morph) of the embryo is generated (genesis). Morphogenesis in both plants & animals involves regulated patterns of cell division and cell elongation that leads to changes in cell shape. Cell movement also plays a critical important role in animal morphogenesis.

Mechanisms of cellular determination

How do cells become different from their parent cells? How do two identical daughter cells become different from one another? How might one daughter cell become a neuron, while the other daughter cell becomes a skin cell? In some cases, determination results from the asymmetric segregation of cellular determinants. However, in most cases, determination is the result of inductive signaling between cells.

Asymmetric segregation of cellular determinants is based on the asymmetric localization of cytoplasmic molecules (usually proteins or mRNAs) within a cell before it divides. During cell division, one daughter cell receives most or all of the localized molecules, while the other daughter cell receives less (or none) of these molecules. This results in two different daughter cells, which then take on different cell fates based on differences in gene expression. The localized cytoplasmic determinants are often mRNAs encoding transcription factors, or the transcription factors themselves. Unequal segregation of cellular determinants is observed during early development of the C. elegans (see image below) and Drosophila embryos. 

P-granule segregation during the early embryonic divisions of the nematode Caenorhabditis elegans:

The image on the right shows an example of asymmetric segregation of cellular determinants in the early C. elegans embryo. All of the cells in the embryo are visible on the left side of the image, while only the P granules are visible on the right side of the image. The P granules were fluorescently labelled - they are the green "dots".

a)  A newly fertilized embryo with dispersed P granules. 

b)  P granules are localized to the posterior end of the zygote. 

c)  After the first division, P granules are present only in the smaller, posterior cell. 

d)  Another unequal division gives rise to a single cell containing P granules. 

e)  When the larva hatches, P granules are localized to the primordial germ cells. 

WATCH MOVIESof P granule movement from Susan Strome's lab!!!

from Susan Strome's lab

Movie of assymetric cell segregation

Although there are many examples where the asymmetric segregation of cellular determinants leads to differences between daughter cells, more frequently we find that cells become different from one another as a result of inductive signals coming either from other cells or from their external environment.

There are many examples in development where an inductive signal from one group of cells influences the development of another group of cells. 

There are three main ways in which signals can be passed between cells. 

In the first mechanism, a diffusible signal is sent through the extracellular space, and is received by a cell-surface receptor, which further transmits the signals by way of second messengers.

In the second mechanism, cells directly contact each other through transmembrane proteins located on their surfaces. 

In the third mechanism, the cytoplasm of two cells is connected through gap junctions,allowing the the signal to pass directly from one cell to another cell. In plants, direct connections between cells are called plasmodesmata.

Although one of the classic models for signaling involves diffusion, there is new evidence that inductive signals may in fact be actively transported within and between cells, and that cellular projections may be involved in long distance communication between cells.

Pattern formation

How do organs develop in their proper positions? How do cells "know" where they are within a developing organism? Pattern formation concerns the processes by which cells acquire positional information.

There are two general models for how patterns form: use of a morphogen gradient, and sequential induction.

The morphogen gradientmodel involves the production and release of a diffusible chemical signal called a morphogen. Morphogen release creates a concentration gradient, with high concentrations of morphogen close to the source, and low concentrations farther away from the source. Exposure to different threshold levels of morphogen leads to different cell fates. In the example below, very high concentrations of the morphogen (above threshold 3) lead to the blue fate, medium levels of morphogen (betweens thresholds 2 and 3) lead to the red fate, and low levels of morphogen (between thresholds 1 and 2) lead to the purple fate. In this way, different amounts of one chemical signal can create a complex pattern.

What is an example of the use of a gradient in pattern formation??

The very first step in patterning the embryo of the fruit fly, Drosophila melanogaster, is a good example of pattern formation by a gradient. We'll talk more about Drosophila development next week. But for now, let's just use it as an example of this important concept.

Bicoid is a transcription factor which turns on different genes in different levels - acting as a morphogen gradient. In this way, the four genes shown in part A (tailless, empty spiracles, hunchback, and kruppel) are found in different locations within the Drosophila embryo, as a result of the amount of Bicoid protein at a particular location in the embryo.

After fertilization, bicoid mRNA from the mother fly begins to be translated into Bicoid protein in the Drosophila zygote. The computer-generated image B shows how the Bicoid protein diffuses through the egg forming a gradient. High concentrations of Bicoid protein are shown in white on the left (anterior) end of the zygote, and low concentrations are shown in blue on the right (posterior) end.

Image C shows Bicoid protein in the nuclei of a Drosophila embryo after a number of rounds of mitosis. Notice that the nuclei in the anterior end (left) have more Bicoid protein than those in the posterior end (right) .

Image D shows  Kruppel protein in orange and  Hunchback protein in green. The region where the two proteins overlap is yellow. The colors come from fluorescent dyes attached to antibodies that bind specifically to these proteins.

images B and D from Robert J. Huskey
image C from the Genetic Networks Database

The sequential inductionmodel involves the production and release of a series of chemical signals. Signal 1 leads to the blue fate and production of signal 2. Signal 2 is received by neighbor cells, and leads to the red fate and production of signal 3. Signal 3 is then received by neighbor cells, and leads to the purple fate. In contrast to the morphogen gradient model, multiple chemical signals are required to create the pattern.

What is an example of the use of sequential induction in pattern formation??

The development of the vulva (a ventral opening used for copulation and egg-laying) in the soil nematode, Caenorhabditis elegans, is a good example of pattern formation by sequential induction.


from Andreas Eizinger and Ralf J. Sommer, Science 278:452-455 (1997)

This diagram of a larval nematode shows the location of cells which will become the vulva.

Vulval precursor cells P1.p through P12.p are identical to one another before the vulva develops. Vulval pattern formation requires the production of an initial signal (open arrow) by the anchor cell (AC). This signal is received by the vulval precursor cell called P6.p. The signal from AC changes P6.p in a way that alters signaling (by a second signal) between P5.p, P6.p and P7.p (filled arrows). As a result, vulval precursor cells P5.p and P7.p attain a high level of the protein LIN-12 (gray shading), but P6.p does not. That is, the combination of both signals makes P5.p and P7.p DIFFERENT FROM P6.p, and also different from their neighbors (shown in the top figure).

from Iva Greenwald

Thanks to David Marcey for construction of some of the images shown above.