Introduction To Pattern Formation

Autonomous & Conditional Cell Specification, Fields &

Morphogens

Cellular Differentiation

• All Cells have the same genes• Some genes are expressed in all cells

– Housekeeping genes

• Some genes are expressed only in specific cells– Luxury genes

• Zygotes are totipotent– Can form any cell in the organism

• Embryonic cells are pluripotent• Differentiated cells are restricted in potential

Fate Maps

Prospective Significance Prospective Potency

Autonomous (Mosaic) Specification in a Tunicate Embryo

Figure 3.8. Autonomous specification in the early tunicate embryo.When the four blastomere pairs of the 8-cell embryo are dissociated, each forms structures that it would have formed if it had remained in the embryo.

(The fate map of the tunicate shows that the left and right sides produce identical cell lineages.) (After Reverberi and Minganti 1946.)

Conditional Specification-Regulative Development in a Frog

Figure 3.11. Conditional specification. (A) What a cell becomes depends upon its position in the embryo. Its fate is determined by interactions with neighboring cells.

(B) If cells are removed from the embryo,the remaining cells can regulate and compensate for the missing part.

Autonomous vs. Conditional Specification

I. Autonomous specification (Prospective potency=Prospective significance)

Characteristic of most invertebrates.

Specification by differential acquisition of certain cytoplasmic molecules present in the egg.

Invariant cleavages produce the same lineages in each embryo of the species. Blasto mere fates are generally invariant.

Cell type specification precedes any large-scale embryonic cell migration.

Produces "mosaic" ("determinative") development: cells cannot change fate if a blasto mere is lost.

II.Conditional specification (PP>PS)

Characteristic of all vertebrates and few invertebrates.

Specification by interactions between cells. Relative positions are important.

Variable cleavages produce no invariant fate assignments to cells.

Massive cell rearrangements and migrations precede or accompany specification.

Capacity for "regulative" development: allows cells to acquire different functions

Another Way of Looking At It

• Animal development can proceed according to either the American or the European plan. Under the European plan (autonomous specification), you are what your progenitors were. Lineage is important. Under the American plan (conditional specification), the cells start off undetermined, but with certain biases. There is a great deal of mixing, lineages are not critical, and one tends to becomes what one's neighbors are - Sydney Brenner (quoted in Wilkins 1993)

The Limb Field

Figure 3.23. Prospective forelimb field of the salamander Ambystoma maculatum. The central area contains cells destined to form the limb per se (the free limb). The cells surrounding the free limb give rise to the peribrachial flank tissue and the shoulder girdle. The ring of cells outside these regions usually is not included the limb, but can form a limb if the more central tissues are extirpated. (After Stocum and Fallon 1983.)

FGF10 Expression Defines The Limb Field

Figure 16.4. FGF10 expression and action in the developing chick limb. (A) FGF10 becomes expressed in the lateral plate mesoderm in precisely those positions where limbs normally form. (B) When cells genetically constructed to secrete FGF10 are placed into the flanks of chick embryos, the FGF10 can cause the formation of an ectopic limb (arrow). (From Ohuchi et al. 1997; courtesy of S. Noji.)

Limb Bud Formation

Figure 16.3. Limb bud formation. (A) Proliferation of mesodermal cells from the somatic region of the lateral plate mesoderm causes the Limb bud in the amphibian embryo to bulge outward. These cells generate the skeletal elements of the limb. Contributions of cells from the myotome provide the source of the limb's musculature. (B) Entry of myotome cells (purple) into the limb bud. This computer reconstruction was made from sections from an in situ hybridization to the myf5 mRNA found in developing muscle cells. If you can cross your eyes, the three

dimensionality of the stereogram will become apparent. (B courtesy of J. Streicher and G. Müller.)

The Skeletal Elements of The Chick Wing

Tissue Interactions

• Induction- the presence of one tissue is required for the formation of a structure in another.– Instructive interaction: The inducing tissue

informs the fate of the responding tissue.– Permissive: The inducing tissue is necessary

for the response to occur but not sufficient to specify ultimate cell fates (Cell fate information is in the responding tissue)

Establishing the Proximal Distal Axis

Figure 16.8. Summary of experiments demonstrating the effect of the apical ectodermal ridge (AER) on the underlying mesenchyme. (Modified from Wessells 1977.)

AER Required for outgrowth P/D Axis

AER not responsible for Specification A/P Axis

AER is permissiveMesoderm is instructive

AER Maintained by Mesoderm

AER secretes FGF

The Apical Ectodermal Ridge

AER

FGF8 Signals Limb Outgrowth

Figure 16.12. FGF8 in the AER. (A) In situ hybridization showing expression of Fgf8 message in the ectoderm as the limb bud begins to form. (B) Expression of Fgf8 RNA in the apical ectodermal ridge, the source of mitotic signals to the underlying mesoderm. (C) In normal 3-day chick embryos, FGF8 is expressed in the apical ectodermal ridge of both the forelimb and hindlimb buds. It is also expressed in several other places in the embryo including the pharyngeal arches. (A and B courtesy of J. C. Izpisúa-Belmonte; C courtesy of A. López-Martínez and J. F. Fallon.)

Evidence that led to the progress zone model:

Sequential removal of the AER results in progressively less disruption of development in a P/D fashion.

The Progress Zone Model For Proximal-Distal Specification

Fig. 1. A cell´s proximo-distal positional value may depend on the time it spends in the Progress Zone. Cells continually leave the zone at the tip ofthe limb under the apical ectodermal ridge. Cells that leave early form proximal structures while cells that leave last form the tips of the digits. (Wolpert 2000)

Specification by the Progress Zone

                                                     

Figure 16.11. Control of proximal-distal specification by the cells of the progress zone. A) Extra set of ulna and radius formed when an early-bud progress zone was transplanted to a late wing bud that had already formed ulna and radius. (B) Lack of intermediate structures seen when a late-bud progress zone was transplanted to an early limb bud. The hinges indicate the locations of the grafts. (From Summerbell and Lewis 1975; photographs courtesy of D. Summerbell.)

An alternative model: Early Allocation and

Progenitor Expansion Model (Pre-specification)

• Observations consistent with this model: Cell division is seen throughout the limb bud

When AER is removed, cell death occurs about 200μm beneath the AER

If the limb bud is a fully formed rudiment that grows as it develops, then 200 μm of cell death will kill progressively smaller amounts of the rudiment.

Anterior Posterior Specification

• Limb field prior to bud emergence is already specified for anterior/posterior, dorsal/ventral

• How is this information maintained during the outgrowth of the limb?

• Classical experiments:

• Extensive series of experiments transplanting pieces of the limb bud from one region to another at different times in bud outgrowth.

• Only one region gave consistent pattern alterations:

The Zone of Polarizing Activity

Figure 16.17. When a ZPA is grafted to anterior limb bud mesoderm, duplicated digits emerge as a mirror image of the normal digits. (From Honig and Summerbell 1985; photograph courtesy of D. Summerbell.)

The “French Flag Model”

Figure 3.19. The French flag analogy for the operation of a gradient of positional information.

(A) In this model, positional information is delivered by a gradient of a diffusible morphogen from a source to a sink. The thresholds indicated on the left are cellular properties that enable the gradient to be interpreted. For example, cells becomes blue at one concentration of the morphogen, but as the concentration declines below a certain threshold, cells become white. Where the concentration falls below another threshold, cells become red. The result is a pattern of three colors.

(B) An important feature of this model is that a piece of tissue transplanted from one region of an embryo to another retains its identity (as to its origin), but differentiates according to its new positional instructions. This phenomenon is indicated schematically by reciprocal"grafts" between the flag of the United States of America and the French flag. (After Wolpert 1978.)

How Could Cells A Interpret a Morphogen Gradient

Anterior And Posterior Limb Fields Are Specified by Position

Questions Which Need to be Answered About Morphogen Gradients

1. What is the morphogen?

2. What is the source of the morphogen?

3. How do cells respond to the gradient?• How do they sense the gradient?• How do they change their fate in response

to morpogen?

4. What sets up the source and sink?

5. What happens to the gradient over time?