LECTURE 8 – MECHANISMS FOR CELL DIFFERENTIATION IN DEVELOPMENT
COMMON OVERALL PATTERNS IN DEVELOPMENT
When comparing the mouse and human em
yo at different stages, it is noted that they are
very similar as both hatch from the blastocyst, developing in a similar manner to humans; it
also develops much quicker, allowing for effective use as a model. Similarly, chicken
develop in an analogous manner to humans, despite the obvious differences in the later
phenotype.
Gastrulation is the point in develop at which pluripotent stem cells become the multipotent
germ layers, differentiating into the cell types that make up the body; for example, the
mesoderm forms the muscle, blood, bone, and cartilage. However, if not done co
ectly, there
are consequences for the rest of development and hence, it is often deemed the most
important point in ones’ life. Across a variety of organisms, the process is similar with cells
moving and invaginating, forming the multipotent layers; this similarity has given rise to the
use of various animal models with the process similar, but since they develop quicker and can
e done outside the body, there are many advantages. During gastrulation, a tube is formed at
the back end which eventually goes to the mouth with this process causing cells to proliferate,
differentiate, migrate, and communicate.
COMMON CELLULAR AND MOLECULAR FEATURES OF DEVELOPMENT
To understand this, many animal models are used. The four essential cellular processes across
multicellular organisms are:
1. Cell Proliferation: This is done as the organism must get larger
2. Cell Specialisation: We begin as a single cell, but must go to 220 cell types.
3. Cell Interaction: These interactions can be physical, or through signalling with the
appropriate signals at the right time triggering migration or differentiation.
4. Cell Movement: Without cell movement, development cannot proceed appropriately.
Underpinning the cellular processes are various molecular mechanisms, with cells beginning
with an identity that must either be renewed or changed. These include gene expression
(selective expression change in proteins produced change in effector functions),
autocrine and paracrine signalling, epigenetics (determine what genes are expressed by
emodelling chromatin), and microRNA.
Part of the genome codes for proteins that are responsible for multicellularity, where more
genes dedicated to this in organisms with more cells. This is seen as there are more
transmem
ane molecules used for cell adhesion, signalling, and ion transport in worms then
unicellular yeast, whilst there are more gene-regulatory proteins in humans when compared
to yeast.
The development of the cerebellum requires Engrailed-1, with the lack of this protein
preventing cerebellum formation. However, it was noted that by using the Engrailed protein
of the fruit fly (Drosophila) helped cerebellum growth, showing the similarity across
organisms in the production of the same protein.
COMMON MECHANISMS OF MAINTAINING CELL IDENTITY
CELL MEMORY
When a new cell is produced, it retains the memory of
its cell type (i.e. a muscle cell will divide into two
muscle cells), with neurons maintaining their identity
to prevent them from becoming another cell type.
One of the key methods of doing this is the use of
positive feedback loops, with the expression of a
protein turned on, with this protein then moving to the
promoter region (transcription factor) to produce more
of this protein with this loop ensuring that the cell
identity is retained, even after cell division. However, there are other mechanisms of cell
memory such as DNA methylation and chromatin remodelling.
MASTER GENES AND GENE CIRCUITRY
Generally, this is where there is a signal that causes the expression of a master gene, which is
esponsible for a circuit, changing the expression of other proteins that then cause the
development of various organs.
In the development of the eye in the Drosophila, it was found that there was a pocket of cells
that expressed ey (the master gene) with this then placed nea
y, forcing the expression of
this gene in the wrong area and hence, caused the production of an eye. However, the lack of
innervation in this area meant that this eye could not see.
Another case is in muscle development, with the master gene being MyoD with forced
expression in chicken skin fi
oblasts then creating muscle cells; this was observed as it was
oth long and multinucleated, as well as contractile.
MORE CELLS AND SPECIALISATION – INCREASING SIZE AND
COMPLEXITY OF THE EMBRYO
There are many mechanisms that control the specialisation including:
- Combinatorial Gene/Protein Control: Since there are
limited numbers of genes (20,000), combinations of these
genes are needed to control development. In each cell
division, the induction of a regulatory protein/s cause
differences in the gene combinations of each cell, increasing
the complexity of the organism.
- Symmetric and Assymetric Division: In assymmetric division (as is observed in the
case of the P granules in the C. elegans germ line), the cells are born differently with
the granules moving to one side of the cell, with the cell cleaved to ensure all granules
are on one side and hence, in one cell. Specifically in the case of the C. elegans, it is
done until the 32 cell stage, with the cell that contains the P granules producing the
germ cell lines. Symmetric division is seen when the sister cells produced are the
same, but from here, the provision of signals means that it forms differently whilst the
other is maintained due to the lack of a signal.
MORPHOGENS – CRITICAL INDUCTIVE SIGNALS IN DEVELOPMENT
Morphogen: A secreted molecule that goes into the environment, acting as a signal.
These are integral in forming gradients that alter the development of the organism. In cases
where a morphogen is released, the concentrations are higher at the source whilst the
diffusion of the morphogen means that the levels decrease further away from the source,
meaning it has less of an effect on development (less likely to bind to a cell further away).
However, morphogen inhibitors can also act, with the source of the inhibitor at a point on the
opposite side thereby having the opposite effect and if in conjunction with the morphogen
itself, the final signal is uniform distribution of the inducer. Different morphogens lead to
different cell types, as it provides short range instruction in the em
yo regarding
development in terms of proliferation, shape changes, migration, differentiation, and death.
This information must be transduced from the morphogen through binding onto a specific
eceptor and hence, cells without this receptor will not be able to respond to the morphogen.
Ligand classes can also have inhibitors that prevent ligand binding at the receptor.
- Sonic Hedgehog Gradients: In the wing bud of the chick, there is a polarising region
which is responsible for the development of the limb. If removed and placed in a
different location on a new chick (meaning there are now two regions but in different
locations), it is noted that the morphogen gradients cause a double-up of digits. A
similar concept is observed with Sonic Hedgehog and FGF10 gradients in lung
anching, as FGF10 binds onto its receptor but then Sonic Hedgehog is secreted at
the bud, inhibiting it directly in front, creating two new centres on the sides, which
then helps to keep the cycle perpetuating.
CELL-CELL ADHESION
There are a class of cadherin genes that code for cadherin proteins. The proteins are dimeric
with calcium binding causing a conformational change, activating them. If two active dimers
on each cell interact, they bind and lock in with respect to each other.
E-Cadherin and N-Cadherin differ and only bind onto the same type of cadherin (i.e. E-
Cadherin binds to E-Cadherin and so on). When sorted out (through calcium addition), it is
noted that they bind based on their expression, as well as level of expression (cells expressing
higher levels of E-Cadherin will bind to other cells expressing more E-Cadherin).
These are critical in neural tube formation as is observed in the Xenopus with it forming it
from the surface ectoderm. This is because the surface ectoderm has E-cadherin whilst N-
Cadherin is only in the Neural Tube. As such there is a sorting out process as shown above.
The neural plate invaginates into the neural tube whilst the neural crest is above it, separating
it from the surface ectoderm.
They are also seen in the Neural Crest movement, with Cadherin 7 found in the PNS. N-
Cadherin is in the neural crest, with the downregulation of this coupled with the upregulation
of Cadherin 7 causing the migration away from the cluster and at the right locations,
Cadherin 7 is then downregulated with N-Cadherin upregulated again (once differentiated).
CELL-ECM ADHESION
To help stay put, Integrins bind to ECM proteins at the basement mem
ane where they
adhere. These are seen in the linings of blood vessels, skin, etc. to prevent the cell from
moving out. Integrins are large proteins that stick out of the mem
ane (and therefore cell),
inding to ECM proteins like collagen, preventing the cell from moving excessively.
LECTURE 9 – MODELLING DEVELOPMENT WITH EMBRYONIC STEM CELLS
ICM TO PRIMITIVE ECTODERM – A CRITICAL STEP IN DEVELOPMENT
The transition from the ICM to the Primitive
Ectoderm happens between days 3.5 and 5.5 in
mouse development, with it like humans despite
the developmental period differences (20 days
in mice when compared with the 9 months in
humans). Further similarities include the
process of hatching from the zona pellucida,
and the subsequent binding to the uterine walls.
The ICM is a small pocket of cells XXXXXXXXXXwhich are pluripotent, forming all adult cells seen
in the full organism. The primitive ectoderm forms all germ layers with it needing
differentiation, as well as maintenance of its pluripotency. However, this process is very
complex as it goes from 30 cells to 6500 cells within hours; this rate is much faster than
cancer cells.
For this process to occur, the key requirements are:
- Maintenance of Pluripotence: This is because both the ICM and Primitive Ectoderm
are pluripotent
- Differentiation Signals: The ICM must convert to the Primitive ectoderm, which
equires various signals for differentiation.
- Proliferative Burst: As explained earlier