MORPHOGENESIS

This differential gene expression is the cause of determination of different cell fates
and cell differentiation. In this unit we will look at early development processes
through which the embryo acquires its shape and structure. The differentiated cell
types are not placed in the embryo in a random manner but are arranged in organized
structures for example limbs, heart, lungs, eyes wings and other internal organs. This
formation of organized structures from simple epithelial sheets and mesenchymal
masses is termed morphogenesis. The early germ layers- ectoderm, endoderm and
mesoderm undergo extensive rearrangement, through regional specification and
directed movement of cells from one location to another in the embryo to form the
three dimensional animal body. These morphogenetic processes involve cell shape
changes, cell migrations and cell to cell interactions which will determine how the
embryo will get its shape. We will learn more about cell to
cell interaction and patterning with the example of the development of a 45
Block 3 Developmental Biology of Vertebrates-I
vertebrate eye and the influence of chemical signals called morphogens in the
process. Finally we will look at the different processes of cell death which are
important in morphogenesis for giving rise to the shape and contours of organs in the
embryo.

MORPHOGENETIC PROCESSES
Before we start discussing the morphogenetic processes, it is important to know that
all embryonic cells are basically of two types - epithelial and mesenchymal
(Fig.11.1).This categorisation of embryonic cells relates to cell shapes and cell
behaviour rather than to their embryonic origin. Epithelial cells can arise from all
three germ layers and mesenchyme arises from mesoderm and ectoderm. An
epithelium is a sheet of cells that rests on a basement membrane and each cell joins its
neighbour by specialised junctions; the cells have a distinct apical – basal polarity.
Epithelial cells form sheets, tubes and lining of organs. Mesenchyme is made up of
loose cells embedded in the extracellular matrix lying between the ectoderm and
endoderm of the developing embryo. It fills up much of the embryo and later forms
the fibroblasts, adipose tissue, smooth muscle and skeletal tissues.

Fig.11.1: Epithelial cells and mesenchymal cells are the two basic cell types in
the embryo.
46
Unit 11 Cell to Cell Communication
Remember that for the embryo to form its structure from a single cell the zygote, the
processes that take place broadly are cell division to make more cells; then these cells
have to be differentiated as per cell fate; the cells have to move, rearrange themselves,
change their shapes and aggregate to form different tissues and organs. Thus the
embryo acquires a recognisable shape and structure of the particular organism. All this
takes place because of communications that occur between the cells of the embryo.
Till a little more than two decades ago, not much was understood regarding the
manner in which cells communicate with each other to construct an organism from a
single cell, the fertilized egg or zygote, through genetically controlled events.
However, towards the late 20th century it became clear that molecules in or on cell
membranes were involved in the ability of cells to adhere, migrate and influence other
cells.

In this section we will discuss three basic processes that require cell to cell
communication through the cell surfaces - cell movement, cell adhesion and cell
signalling. These are the key properties of cells that are involved in changes in
embryonic form. Let us look at the property of cell motility first.

Cell Movements
Cell movements or motility is an active phenomenon that is essential for many
biological processes such as morphogenesis, wound healing, immune response and
even cancer metastasis. In this unit our focus is on morphogenetic processes where
cell movement is targeted to specific sites in the developing embryo to form tissues
and organs, A good example of these cell movement or cell migrations is seen in the
movement of neural crest cells (multipotent cells that arise from embryonic ectoderm
and give rise to different types of cells) and germ cells in vertebrates. Short range
movements are also important and cell motility is responsible for both movements of
individual cells as well as change of shape while remaining part of a tissue. For
example, the folding of epidermal sheets to make tubes is caused by changes in the
shape of the cells.

All cells move and change shape by rearranging their internal cellular skeleton
(cytoskeleton) or scaffolding by contraction of the cytoskeleton fibres made up of
microtubules and microfilaments that are actin – myosin complexes also termed as
actomyosin complexes. These actomyosin complexes are simpler version of those
seen in muscles. The energy required to produce the movement comes from adenosine
triphosphate (ATP). In non muscle cells these actomyosin complexes are concentrated
in the region just below the cell membrane. Moving cells also have a polarity that is, a
front and a back region. The mechanism of cell movement can best be seen in the
movement or crawling of fibroblasts (a type of connective tissue that secretes collagen
found in the extracellular matrix) on a substratum which is the extracellular matrix
inside the embryo or glass surface of petri-plates under in vitro conditions (Fig.11.2).
Fibroblasts extend a flat process called lamellipodium which is rich in microfilaments
made up of a crisscross of actin. From the lamellipodium extend focal contacts that
attach it to the substratum and these are connected to the microfilament bundles of the
lamellipodium. During movement the
microfilaments contract and the body of the cell is pulled forwards. Cells of the 47
Block 3 Developmental Biology of Vertebrates-I
embryo essentially move in a similar manner. Instead of the large lamellipodium they
may have multiple thin filopodia that make the contact with the extracellular matrix
as they move over it.

In the embryo the cell movement is directional towards a signal which is a

chemoattractant that is detected by the proteins on the cell membrane. These
chemoattractants are diffusible molecules and the cells move towards increasing
concentrations of the diffusible molecules. You will learn more about this
phenomenon in a later section.

Fig.11.2: Fibroblast moves by extending the large flat lamellipodium that makes
contact with the substratum.

Cell shape also changes by the contraction of microfilaments and the associated
motor proteins actin and myosin. If the constriction happens in the apical region of
epithelial cells it will reduce the apical surface area and elongate the cell (Fig.11.3).
This happens initially during invagination during the process of gastrulation when the
cells leave the epithelium to move inside the gastrula (you will learn more about the
cell movements during gastrulation
in Unit 12).
48
Unit 11 Cell to Cell Communication

Fig. 11.3: Cell shape change in epithelial cells by apical constriction and result
in elongation of the cell.

SAQ 1
Fill in the blanks:

i) Cell movement takes place because of rearrangement of

…………………of the cell along with...................................molecules.

ii) Cells in the embryo move over ………………… ………………….

iii) Embryonic cell movement is a response to ………………. ……………..

from other cells.

Cell Adhesion
The other important property that is involved in changes in animal embryonic form is
cell adhesiveness. Animal cells stick to one another and to intercellular matrix
through interactions involving cell surface proteins. These cell surface proteins can
determine how specifically and tightly the cells adhere to one another. These proteins
can affect the cell surface tension and contribute to the arrangement of cells in the
three germ layers and later in different tissues.

Differences in cell adhesiveness also help to maintain the boundaries between

different cell types and tissues. Different cell types have both, different types and
different amounts of cell adhesion molecules on cell surfaces thereby, having
selective affinity for each other which is important for giving positional information
to embryonic cells. Because of cell adhesion embryonic cells do not sort out randomly
but can actively move to create tissue organisation. The differential adhesion
interactions of cells form a certain hierarchy. If cell type A is situated internal to cell
type B and the final position of cell type B is situated internal to C, then cell type A
will always be internal to C. There are different
49
Block 3 Developmental Biology of Vertebrates-I
classes of cell adhesion molecules, but the major cell adhesion molecules appears to
be cadherins (calcium dependent adhesion molecules). Cadherins are
transmembrane proteins that interact with other cadherins present on adjacent cells.
Cadherins are anchored to their cell by a complex of proteins called catenins
(Fig.11.4). This cadherin- catenin complex is the classic adherin junction seen in
epithelial cells. As the catenins bind to the cytoskeleton of the cell they integrate the
epithelial cells and keep them together. Cadherins perform several related functions.
(i) their external domains serve to adhere cells together. (ii) cadherins link to and help
assemble the actin cytoskeleton, thereby, providing the mechanical forces for forming
sheets and tubes of cells. (iii) cadherins can serve to initiate and convert signals that
can lead to changes in a cell’s gene expression
Fig.11.4: Diagram showing cadhedrin - cadherin cell adhesion. Inside the cell

the cadherin molecule associates with a catenin molecule which itself

is bound to the actin microfilament system of the cytoskeleton
(adapted from Takeichi 1991).
50
Unit 11 Cell to Cell Communication
Cadherins can be of different types (see Box 11.1) and cadherin of one cell binds to
the other cells by the same type of cadherin. For example,cells with E-cadherin will
stick to other cells with E-cadherin and sort out from cells with N- cadherin. This
type of binding is known as homophilic binding. The sorting of cells was first
demonstrated in the 1950s. Since it was known that amphibian tissue dissociate into
single cells when placed in alkaline solutions,
a single cell solution of the three germ layers of amphibian embryos was made and
when the pH was restored these cells adhered to one another in the petri dishes. Thus
the behaviour of recombined cells could be studied. The results of the experiment
were striking , the cells sort out into their cell type and into their own regions
(Fig.11.5).

Fig 11.5: Reaggregation and sorting of cells from two different amphibian
neurulae.Presemptive epidermal cells from a pigmented embryo and
neural plate cells from a unpigmented embryo were dissociated and
mixed together. The cells first aggregate together and then the cells
segregated according to their cell type position. The presumptive
epidermal cells cover the neural cells.

The sorting of cells is the combined effect of cell adhesiveness and cell movement.
Initially the cells move randomly but then they seek similar cells to aggregate because
they have stronger adhesive interactions. The ectoderm is the tissue with the
strongest cohesive interaction among cells so it forms the outermost layer of the
embryo. The less cohesive mesoderm and endoderm cells are arranged internal to the
ectoderm. Cell sorting hierarchy is, therefore, strictly dependent on the amount of
cadherin interactions amongst cells.

In vitro experiments with fibroblasts that are generally migratory cells and do not
express E- cadherin, showed that when activated E-cadherin genes were added to the
culture, the fibroblasts started expressing E- cadherin and they became tightly bound
to each other and started behaving like epithelial cells. Blocking the function of
cadherins by using antibodies that inactivate cadherin or by blocking its synthesis at
translation stage can prevent epithelial tissue
from forming and causes the cells to disaggregate. 51
 Block 3 Developmental Biology of Vertebrates-I
Cell adhesion, cell movement and formation of epithelial sheets and tubes depend on
the ability of cells to attach to extracellular matrix. When epithelia are to be made the
attachment has to be very strong but when cells move or migrate, they have to break
their attachment to other cells and reform them at another location. In some cases the
extracellular matrix has to serve as a pathway for the moving cells, providing direction
or the signal for a developmental event. Fibronectin a component of extracellular
matrix in a way paves the roads on which the cells move, for example, fibronectin
paths lead the germ cells to gonads and heart cells to the midline of the embryo. The
binding of the cell to the extracellular matrix takes place through another family of
adherin molecules called as integrins and the mechanism is not dependent on
calcium unlike that in cadherins. Integrins are large protein molecules that span
across the cell membrane. On the outside of the cell, it binds to the fibronectin of the
extracellular matrix; on the inside of the cell, it serves as an anchorage site for the
actin microfilaments that move the cell.

Box 11.1: Cadherin molecules.

There are many type of cadherin molecules and cadherin – cadherin attachments are the
strongest when they are of the same kind.
 E-cadherin: epithelial cadherin is expressed on all early mammalian embryonic cells
and later gets restricted to the epithelial cells of embryo and adults
 P –cadherin: this is placental cadherin expressed on the trophoblast of the
placenta and on the uterine wall epithelium. It possibly facilitates the attachment
of the placenta to the uterus
 N- cadherin: is neural cadherin, first seen on mesodermal cells in gastrula cells as they
lose their E-cadherin expression. It is also seen on cells of the developing central
nervous system.
 C-cadherin: is found to be critical for keeping the blastomeres together in
Xenopus and for normal development for gastrulation cell movements.
 Protocadherins: are calcium dependent adhesion molecules like cadherins but lack the
connection to cytoskeleton through catenins.

SAQ 2
a) How are boundaries between different tissues maintained?

b) What is the difference between cadherins and integrins?

Cell Signalling
Cells typically communicate with each other by use of chemical molecules or
signals. These molecular signals are highly diverse and are responsible for specific
protein-protein interactions, which can result in diverse cellular responses like
changes in gene transcription, cell metabolism, cell migration
and cell death. The signal molecules generally contact other proteins that may
52
Unit 11 Cell to Cell Communication
be housed in or on the plasma membrane of the target cells. The signal protein
molecules are called ligands (a general term for molecules that specifically bind to
other molecules). The proteins that are attached to or embedded in the cell membrane
of the target cells are known as receptors. A receptor in the membrane of one cell
can bind a similar receptor in another cell in homophilic binding. In contrast,
heterophilic binding occurs between different receptor types. Binding of a ligand to
a receptor generally sets up a signal transduction pathway. The major signal
transduction pathways all appear to have a common theme (see Fig.11.6). Each
receptor spans the cell membrane and has an extracellular region, a transmembrane
region, and an intracellular cytoplasmic region. When a ligand binds to its receptor’s
extracellular part, it induces a shape change in the receptor’s structure. This shape
change is transmitted through the membrane and alters the shape of the receptor’s
cytoplasmic part, giving that domain the ability to activate cytoplasmic proteins. Such
a conformational change often makes the cytoplasmic part become enzymatically
active, using ATP to phosphorylate specific tyrosine residues of particular proteins.
Thus, this type of receptor is often called a receptor tyrosine kinase (rTk). The
active receptor can now catalyze reactions that phosphorylate other proteins, and this
phosphorylation in turn activates their latent activities. Eventually, the cascade of
phosphorylation activates a dormant transcription factor or a set of cytoskeletal
proteins.

Fig. 11.6: Structure and function of a tyrosine kinase receptor.

Signal transduction pathways that end in expression of a gene in the target cell are
generally slower than those that enzymatically activate biochemical
pathways or regulate cytoskeletal elements for movements. 53
Block 3 Developmental Biology of Vertebrates-I
Cell signals can be categorised according to the distance travelled in the body as:
autocrine, paracrine, direct contact and endocrine (see Table 11.1).

Table 11.1: Types of cell signalling

Paracrine signalling Takes place over short distances between cells by diffusion

Juxtacrine signalling Takes place between cells in direct contact

Autocrine signalling Signals are received by the same cell that sent them or
from adjacent cells of the same kind
Endocrine signalling Signals are carried over long distances through the
blood stream to the target cells

Direct signalling Occurs through gap junctions between neighbouring

cells. Allows flow of small molecules between cells

In an embryo, communication between cells can occur (i) across short distances, such
as between two neighbouring cells through their cell membranes in direct contact, or
between a cell membrane and extracellular matrix secreted by another cell called
juxtacrine signalling (Fig.17.7 A&B), or
(ii) between neighbouring cells through the secretion of proteins into the extracellular
matrix, called paracrine signalling (Fig.11.7 C). In the next section we shall
examine cell to cell interaction using these two types of communications.

Fig.11.7: Modes of communication between cells of the embryo. A) Paracrine

signalling in which one cell secretes a signalling protein or ligand into

the environment and across some distance from many cells. Only
those cells can respond that have receptors for that particular ligand.
The receptor can respond, either rapidly through chemical reactions in
the cytosol, or more slowly through the process of gene and protein
expression; B) and C) Juxtacrine signalling is local cell signalling
carried out via membrane receptors that bind to proteins in the
extracellular matrix (ECM) or directly to receptors from a neighbouring
cell.

Epithelial-Mesemchyme Transition
All the cell processes that we have learnt in the previous sub-sections are seen to
be integrated in another morphogenetic process –the epithelial-
54 mesenchyme transition also known as EMT. During this process the
Unit 11 Cell to Cell Communication
stationary epithelial cells get detached from the basal lamina and change their identity
to become migratory mesencymal cells that can invade tissues and form organs in
new places in the embryo. EMT is usually initiated by a paracrine signal from
neighbouring cells that initiates gene expression in the target cells. This gene
expression instructs the target cells to downgrade their cadherins, release their
attachment from their basal laminin, rearrange their cytoskeleton components and
secrete the extracellular matrix molecules that are characteristic of mesenchymal
cells. EMT can take place involving individual cells or the collective epithelial cells
(Fig.11.8)

Fig.11.8: Epithelial –mesenchyme transition. A) individual epithelial cells can

detach and change into mesenchymal cells; B) a sheet of epithelial
cells that moves along the front end towards the direction of migration.

This EMT is very important in the developmental process. For example, it is seen in
the formation of neural crest cells from the dorsal most region of the neural tube in
fish, birds and mammals. It is also seen in the formation of chick and mouse
mesoderm where cells that were part of the ectoderm detach and convert into
mesodermal cells that migrate into the interior of the embryo; and in cells that detach
from the somites and migrate around the developing spinal
cord to ultimately form the vertebrae. In all these cases the epithelial cells lose 55
Block 3 Developmental Biology of Vertebrates-I
their “epithelialness” that is the breakdown of the basal cadherins-intercellular matrix
adhesion and cadherin–cadherin junction at the apical region of the cells, thus loss of
cell to cell adhesiveness before they begin to behave like mesenchymal cells and
migrate.

In adults the process of EMT is needed for wound healing, regeneration of tissue
and is the cause of metastasis of cancer cells, where the solid tumour cells detach
from the tumour and migrate to other parts to form more solid tumours.

SAQ 3
a) How would you define a ligand in cell- to cell signalling?

b) What is the difference between juxtacrine and paracrine signalling?

c) How is EMT used in the embryo and in the adult?

CELL–CELL INTERACTION
In the last section we discussed the role of cell adhesion, cell motility and cell
signalling as crucial processes in morphogenesis. We should also realise by now that
from the early stages of embryogenesis, cells do not function in isolation or in a
random manner. All cell behaviours like cell adhesion, cell migration, differentiation
and cell division are regulated by signals being passed from one set of cells to
another. These cell to cell interactions allow the embryo to get its form and shape.
But how do organs develop in their proper place in the embryo? And how do cells
“know” that they have to migrate and position themselves? Pattern formation is
the process by which the cells find their positional information. There are two
general modes of pattern formation: (i) through use of morphogen gradients and
(ii) by sequential induction. Let us first take a look at the role of morphogen
gradients.

Morphogen Gradients
Pattern formation in the embryo can involve gradient of chemical signals known as
morphogens. This term was coined by Allen Turing in 1952 for substances whose
distribution through diffusion would determine the development of cells which would
respond to different threshold concentrations of the morphogen. This is a type of
paracrine signalling in which the concentration of morphogen is high near the source
of release and becomes lower as the distance of the responding cells increases from
the source. Morphogens can consist of cytoplasmic proteins such as transcription
factors that can form a concentration gradient in a single cell or syncytium as seen in
early embryo of Drosophila, or secreted as signalling molecules that travel from cell
to cell. In most cases, the responding cells find their positional information because
of the different concentrations of morphogens along its gradient. Thus morphogens
guide the formation of different cell types in a

56
specific positional order by inducing transcription of genes in a dose
Unit 11 Cell to Cell Communication
dependent manner. Let us understand this with a hypothetical example. In Figure 11.9
we see a plane of unspecified cells in a region of the developing embryo. Out of these
cells type A cell have matured enough may be due to maternal cytoplasmic
determinants (refer to Unit-10) so that their function is specified. A type cells begin to
express a signalling protein that functions in this case as a morphogen. The other cells
seen in the figure (B, C, and D) are still unspecified in their cell fate but are
competent to respond to this particular morphogen as they have the receptors to
bind to the morphogen. As the morphogen is secreted the cells closest to the signalling
cells come in contact with the signal first. Over time the morphogen diffuses and a
concentration gradient is formed as the cells closest to the source experience more
ligand-receptor binding and for longer duration than the cells located farther away
from the source. Cells show a differential response to the morphogen in term of the
duration and the concentration of the morphogen they are experiencing. As a result
they differentiate and mature by different pathways.

Fig. 11.9: Theoretical morphogen gradient induces different cell types. A type
cells release the morphogen shown in red and depending on the

concentration gradient and duration of exposure different cell fates

are seen for cell type B, C, and D

In this way a morphogen gradient describes a mechanism by which cells of one part of
the embryo can determine the location, differentiation and fate of many surrounding
cells and provide a basis for understanding many patterning processes. The role of
these signals range from establishment of initial polarity of embryos to specification of
cell identity in particular tissues, notably in limb appendages and nervous system of
both vertebrates and Drosophila. The best studied example of morphogen action is in
Drosophila embryo which you have studied in the previous unit (Refer to subsection
10.6.1). The graded distribution of Bicoid a transcription factor from the anterior end
to posterior end acts as a morphogen that establishes anterio-posterior polarity in early
embryo. Two different genes are expressed one named orthodenticle by high levels of
Bicoid and the second named hunchback gene by low levels of Bicoid.

Embryonic Induction
The embryonic process during which the close range influence of one cell or group
of cells on adjacent cell/cells resulting in the change of cell behaviour like change in
shape, mitotic rate, cell fate is called embryonic induction. There are at least two
components to every inductive interaction-the inducer, which is the tissue that
produces a signal or signals that change the cell behaviour of the receiving tissue or
responder. Not all tissues are capable of responding to the signals produced by the
inducer, only the tissues that have the ability to respond can be induced. This ability
to respond is called
competence which is an acquired condition.
57
Block 3 Developmental Biology of Vertebrates-I
Let us understand this phenomenon by taking the example of the development of the
vertebrate eye. When the development of eye is initiated, two bulges are seen in the
brain that approach the surface ectoderm in the head region. The head ectoderm is
competent to respond to the paracrine factors released by the brain bulges that are the
optic vesicles. The head ectoderm cells are induced to form the lens tissue of the
eye that is, the genes for expressing the lens protein are activated. The prospective lens
cells in turn secrete another paracrine factor that instructs the optic vesicle to form the
retina. Note that the two cell types that co-construct the eye induce each other.
The important part is that the head ectoderm is the only region in the embryo that is
competent to respond to the instructions from the optic vesicle signals. Experiments
done with Xenopus laevis embryo show that if the optic vesicle is placed in another
part of the head ectoderm it still can induce lens tissue but if the optic vesicle is
implanted anywhere else say underneath the trunk ectoderm it will fail to induce the
lens tissue, showing that only the head ectoderm is competent to respond to the
inducing signal (see fig.11.10).

Fig. 11.10: Ectoderm’s ability to respond to the optic vesicle inducer in Xenopus
is due to competence of ectodermal cells in the head region. The
optic vesicle is able to induce lens formation in the anterior portion
of the head ectoderm a); b) but not in the presumptive trunk region;
c) If the optic vesicle is removed no lens is formed by the overlying
head ectoderm; d) most of the other tissues implanted under the
head ectoderm are not able to substitute for the optic vesicle.

Further study in amphibians has shown that often one induction will give a tissue the
ability to become competent to respond to another induction in a sequential manner.
Recall that we had said that the competence is an acquired condition and in
amphibians the head ectoderm becomes competent to be induced by the optic vesicle
only through sequential signals from the underlying foregut endoderm and heart
forming mesoderm in the early gastrula. The next signal is given by the anterior
neural plate which instructs the synthesis of a transcription factor called PAX6 which
is required for ectodermal competence to respond to the optic vesicle. Thus we see
that though the optic vesicle appears to be the inducer of lens formation the
ectodermal cells had been made competent by at least two other tissue.Their
effect on the ectoderm is synergistic and cumulative. As lens induction begins at the
gastrula stage the competence of the non-neuroal ectoderm becomes
restricted to the head ectoderm in the regions fated to form the lens.
58
Unit 11 Cell to Cell Communication
Another important feature of embryonic induction is that an inducer tissue is often
induced by the receiver tissue as well. For example once the lens is formed it induces
the optic vesicle to become the optic cup and the walls of the optic cup become
distinguished to form two layers one, the neural retina and second, the pigmented
retina. Such induction is called reciprocal induction. At the same time the lens also
induces the ectoderm above it to form the cornea. The cornea forming ectoderm also
acquires its competence to respond to the signals from the lens tissue. Under the
influence of the lens the corneal ectoderm becomes columnar and secretes multiple
layers of collagen. The mesenchyme cells from the neural crest use this collagen
matrix to enter the area and secrete proteins that further differentiate the cornea. A
third signal which is hormonal, dehydrates the tissue and the makes it transparent.
Thus we can see that there is a sequence of induction events as shown in the
development of eye in Fig.11.11.

Fig. 11.11: Induction of lens in mouse. a) the optic vesicle extends toward the
surface ectoderm from the forebrain. The lens placode (the
prospective lens) appears as a local thickening of the surface
ectoderm near the optic vesicle b) the lens placode enlarges and the
optic vesicle has formed an optic cup; c) the central portion of the
lens-forming ectoderm invaginates, while the two layers of the retina
become distinguished; d) the lens vesicle has formed; e) the lens
consists of anterior cuboidal epithelial cells and elongating posterior
fiber cells. The cornea develops in front of the lens. The whole
process takes about 4 days starting from day 9 in the embryo.

Induction interactions are called instructive if the signal from the inducing cell
causes a new gene expression in the responding cell. In this case without the
inducer cell the responding cell is not capable of differentiation into a particular 59
Block 3 Developmental Biology of Vertebrates-I
cell type. Example of instructive interaction is when the optic vesicle of Xenopus is
placed in another part of the body’s ectoderm and it fails to induce the lens.
Permissive interactions during induction are seen when the responding tissue is
already specified but needs the environment to express its tissue characteristics. A
good example of permissive interaction is the extracellular matrix that is often
required by tissues to develop. The matrix itself does not alter the tissue type but
allows the already specified tissue to express the traits of the tissue they are supposed
to be. Thus, permissive interactions lend to regulate the degree of expression of the
remaining developmental potential of the already specified cells.

SAQ 4
Match the following statements with the given terms:

i) Permissive Interaction

ii) Morphogens

iii) Competence

iv) Instructive interaction

Statements

1) Compound or molecule that is diffusible and influences the movement and

organisation of cells as per a gradient.

2) High levels of transcription factor produces cell type B and low levels of
transcription factor A produces tissue type C

3) A signal from the inducing cell that is required to initiate a new gene
transcription

4) The responding cell is already specified but its degree of expression is

regulated by the inducer signal.

5) In the presence of tissue A tissue B develops in a certain way but in the

absence of tissue A it does not develop in that way.

6) Signals from Tissue A induce the formation of a cell type B in one region of
the embryo but when tissue A is transplanted to another region of the embryo it
fails to induce the cell type B.

CELL DEATH
Death is the ultimate event in the life of all organisms. Death and degradation of cells
takes place at particular times. Death of cells is also induced in cases of acute injury
and infection, DNA damage, toxicity and trauma or when cells turn cancerous. Cell
death may occur through ‘cellular suicide’ with the help of the intracellular organelles
called lysosomes (autophagy); or when induced by acute injury or infection
(necrosis); or through programmed cell death
(apoptosis) Apoptosis is genetically controlled through a pre-designated
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 Unit 11 Cell to Cell Communication
pathway. Autophagy and Necrosis cause cell death through alternate
pathways.

poptosis
Apoptosis (both p are pronounced) was a term coined in 1972 for the process of
active cell death resulting in the orderly breakdown of cellular
structures. This is also known as programmed cell death since apoptosis is
triggered by the expression of specific genes and blockage of mRNA or protein
synthesis. It was best understood in the nematode C elegans where out of a total of
1090 somatic cells 131 undergo programmed death during development. This number
of dying cells in invariant in the species. Since then the significance of programmed
cell death was established in developmental biology of other species for genetically
determined elimination of cells.
Apoptosis occurs normally during development and aging as a homeostatic
mechanism to maintain cell populations in tissues and as a defence mechanism in
immune reactions and diseased conditions. Apoptosis is often initiated by removal of
growth factors from cells and sometimes as an active response to a signal during the
developmental process. For example, it is the process responsible for elimination of
tissues during finger and toe formation in limbs of vertebrates, and loss of the tail in
tadpoles during metamorphosis.

Some of the major events of apoptosis include chromatin condensation and nuclear
fragmentation. As apoptosis proceeds, individual cells or clusters of cells become
increasingly round and undergo pyknosis (the reduction of cell volume), the
membrane bulges out in small portions known as blebs and organelles undergo
modifications. However the cell does not lose its membrane integrity and the cells
contents are not released in to the intercellular spaces and no inflammatory responses
occur. The dying cells are usually phagocytosed typically by macrophages to prevent
secondary necrocis. Apoptosis is an energy-dependent process. Three major
biochemical events occur during apoptosis:

i) DNA and protein breakdown: Early apoptosis is characterized by the

breakdown of DNA into 50-300kb fragments that are then cut into
oligonucleotides by endonucleases. This results in DNA fragments of a range
of lengths.

ii) Activation of caspases: Late apoptosis is characterized by the activation of

cysteine proteases known as caspases that break down the protein and
cytoskeletal components of the cell and activates the enzyme DNase, which
continues the degradation of the cell’s DNA. Caspases breakdown about 100
cellular proteins which bring about changes in the cell which are characteristic
of programmed cell death or apoptosis, for example out of a family of a dozen
caspases, one is inhibitor of DNA and causes DNA fragmentation; another
called nuclear lamin brings about nuclear fragmentation. A cascade of caspases
breaks the cell into small membrane-covered vesicles by folding the cell
inwards. The cell membrane remains intact during the process despite vesicle
or blebs budding on the cell surface. As apoptosis progresses, these cells will
lose the cell-to-cell adhesions and will separate from the
neighboring cells (Fig 11.12).
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Block 3 Developmental Biology of Vertebrates-I
iii) Membrane modifications: In which an intracellular transmembrane
protein, is exposed on the surface by which phagocytes recognise the dying
cell.

The process of apoptosis is divided into two pathways: the intrinsic pathway if
the apoptosis is initiated in the mitochondria and
the extrinsic pathway which is started by the binding of signalling
molecules or ligands to transmembrane receptors that involve death receptors
domains on the intracellular side and these lead to a cascade of reactions that
start the apoptotic process.

Necrosis
Necrosis another cell death process is different from apoptosis. It can happen
simultaneously or independently of programmed cell death. Necrosis is accidental
death induced by an acute injury or a deadly disease or DNA damage as opposed to
programmed cell death. Necrosis, therefore, is not a naturally occurring phenomenon
but is induced by serious injury. In Greek necrosis means death.

Necrosis is considered to be a toxic process which is uncontrolled and passive that is,
it does not require an energy source to take place. It affects a large field of cells and is
mediated by two main mechanisms- injury to the cell membrane and interference with
the supply of the energy to the cells. Some of the major morphological changes that
occur with necrosis include cell swelling; formation of cytoplasmic vacuoles;
disruption of ribosomes and organelle membranes; swollen and ruptured mitochondria
and lysosomes; and eventually disruption of the cell membrane. This loss of cell
membrane structure results in the release of the cytoplasmic contents into the
surrounding tissue (Fig.11.12).

62 Fig. 11.12: Cell death by necrosis and apoptosis.

 Unit 11 Cell to Cell Communication
The unregulated release of products of cell death to the outside, initiates
inflammatory response in the surrounding tissue. Leucocytes and nearby
phagocytes eliminate the dead cells through phagocytosis. Damaging substance
from leucocytes can cause damage to surrounding tissues and if untreated, necrosis
results in building up decomposing dead tissue and cell debris at or near site of
necrotic cell death (e.g. gangrene). Often necrotic tissue needs surgical removal.

utophagy
Eukaryotic cells have their own method of removal of nonfunctional or damaged
organelles or other cell components or even cells. Lysosomes are the organelles in the
animal cell that carry out this function and hence they are also called “suicidal bags”.
Autophagy is another form of cell death process by which lysosomes help to
maintain homeostasis through recycling of the cells’ own organic material (Fig.
11.13).

Fig.11.13: Electron micrograph of lysosome of a cell breaking down a damaged

mitochondrion for its disposal.

The process of autophagy is evolutionary conserved, meaning whereby that the

process of autophagy is the same in single celled organisms like yeast to humans.
Similar to apoptosis, autophagy has an important role in development processes,
human disease and cellular response to nutrient deprivation. In some circumstances, a
correlation has been documented between reduced autophagy and cancer suggesting
that autophagy is a safeguard mechanism against unrestricted cell growth. This self-
degrading process is used for three key reasons and they are: (i) To recycle proteins
and organelles that do not pass quality control or are in excess in an effort to maintain
homestasis, (ii) To sustain the cell in times of need by degrading less-essential cellular
components as an alternative fuel source, and (iii) To defend itself against pathogens.

There are three main forms of autophagy known as macroautophagy (usually

referred to as autophagy), microautophagy that involves the transfer of the cytosolic
cargo directly into the lysosomes through membrane invagination and chaperone
mediated autophagy in which proteins known as chaperones accompany the cargo
inside the lysosome (Fig. 11.14).
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Block 3 Developmental Biology of Vertebrates-I
The role of autophagy in cell death can be:

i) Autophagy – associated cell death where induction of autophagy

accompanies apoptosis or some other cell death pathway.
ii) Autophagy - mediated cell death where autopahgy pathways activate
apoptosis pathways.
iii) Autophagy – dependent cell death which occurs independent of both
apoptosis and necrosis.
Autophagy is influenced by several genes, which if lost can result in some
cancers.

The characteristic morphological features of autophagy are: enclosing cytoplasmic

organelles in double or multi-membrane vesicles and delivering them to the cells own
lysosomes for degradation. In one sense the cell eats itself! The process of autophagy
is dependent on constant protein synthesis and ATP. In general there are 4 stages in
the process – (i) Induction which may be due to intracellular stimuli or signals to
stimulate the protein kinase, a regulatory protein. (ii) formation of autosomes in
which the cellular material to be disposed off is broken down and is called ‘cargo’
which is then packaged into an autophagosome (iii) fusion with lysosomes or
vacuoles, (iv) autophagy body breakdown and recycling.

Fig 11.14: Various forms of autophagy and how they take place.
Macroautophagy in which the organelles and cytoplasmic parts are
enclosed in a autosome that merges with the lysosome.
Microautophagy in which the cytoplasmic cargo is drawn in to the
lysosome by involution of the membrane and chaperone mediated
autophagy in which the cargo is delivered by the assistance of a
chaperone protein into the lysosome.
Unit 11 Cell to Cell Communication

SAQ 5
In the table given below, fill in at least four distinguishing characteristics of
apoptosis, necrosis and autophagy.
Apoptosis Necosis Autophagy

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Block 3 Developmental Biology of Vertebrates-I
SUMMARY
In this unit you have learned that:

 All embryonic cells are either epithelial cells which are tightly connected to
one another in sheets or tubes or mesenchymal cells which are unconnected or
loosely connected. The process by which these cell types in the embryo get
organised to give form and structure to the embryo is known as
morphogenesis.

 Morphogenetic processes that give structure to the embryo involve key cell
properties like cell movement, cell adhesion and cell to cell communications.
Cell movements and cell shape changes take place by the rearrangement of the
internal cytoskeleton made up of microfilaments and microtubules. Cells
typically move by a crawling motion which is best seen and studied in
fibroblast cells both in vivo and in vitro conditions. Moving cells in the embryo
use the intercellular matrix as a substratum forming pathways for migration.

 Animal embryonic cells stick to one another and to the extracellular matrix due
to cell surface proteins known adherins. These belong to cadherin family or
integrins molecules. Cadherins are transmembrane proteins and interact with
other cell cadherins of the same kind for the cell to cell adhesions. Their
intracellular domains consist of a protein known as catenin which also binds to
the cytoskeleton filaments in the cell. Integrins bind the cell to the extracellular
matrix. Adherin molecules are responsible for cell sorting and maintaining
tissue boundaries.

 Cell to cell communication occurs by use of chemical signalling. The signalling

proteins called ligands bind to receptor or proteins on target cells to set up a
signal transduction pathway that often controls the gene expression of the target
cell. Cell signalling can be categorised according to distance travelled in the
embryo as paracrine, autocrine, juxtacrine and endocrine.

 An example of cell to cell interaction is seen in the epithelial- mesencymal

transition that can involve individual cells or sheets of epithelia in the
embryo. EMT is also seen in the adult in the process of wound healing,
regeneration and is crucial in cancer metastasis.

 Pattern formation is the result of cell to cell interaction. It is the process by

which the cells find their positional information in the embryo.The two general
modes of pattern formation are by morphogen gradients and through sequential
embryonic induction. As the cells are exposed to different concentrations of the
morphogens in a gradient they express different genes at different times and
locations. Induction involves the influencing of a set of cells by another set of
cells or a cell so that the behaviour of the influenced cells/cell changes.
However only those target cells can be influenced by induction that are
competent to respond to the influencer, signals.

 The development of the vertebrate eye is a good example of embryonic

induction event. Often one induction event is necessary for the target cell to
induce another target tissue leading to a cascade of induction events as seen in
the development of lens of eye.
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Unit 11 Cell to Cell Communication
 Programmed cell death or apoptosis is also important in development.
Apoptosis is a means of cell death without the spilling of cell contents into the
surrounding. It involves molecular pathways where proteases like caspases
cause the nucleus to condense and cell to shrink, displaying signals on the
surface to attract phagocytes. Other cell death processes in organisms are
necrosis and autophagy which have different pathways from apoptosis.

TERMINAL QUESTIONS
1. In what ways can inducing signals be transmitted in the developing
embryo? Explain using examples.
2. What are cadherins? What is their role in cell to cell interactions?
3. Mark the following statements as true or false.
i) During development cells move by extending their cell processes in all
directions without any polarity
ii) The cytoskeleton of the cell is actively involved in cell movement and
cell shape change.
iii) Cadherin on one cell binds strongly to similar cadherins molecules on
other cells
iv) Integrins are cell adhesion molecules that attach cells to other cells at the
apical region
v) Permissive interaction involves gene expression in the induced cell
vi) Morphogens are paracrine signalling molecules
vii) In apoptosis the cell organelles are delivered to lysosomes and the cell
contents are spilled out
viii) Cell death by necrosis indicates that cell has undergone a toxic
reaction
ix) Optic cup is induced to form the retinal layers by reciprocal
induction from the lens
x) Pattern formation is the result of morphogenesis
4. Define morphogenesis and list the cell behaviours responsible for it.
5. Why is cell death a necessary event in embryonic development?
6. What is a signal transduction pathway? Write out its step in the order they
would occur in a target cell.

ANSWERS
Self-Assessment Questions
1. i) cytoskeleton; adhesion

ii) extracellular matrix.

66 iii) chemical signals

Block 3 Developmental Biology of Vertebrates-I
2. i) Differential adhesiveness of cadherins interactions. Similar
cadherins attach to each other on cell surfaces while dissimilar
cadherins do not attach or have very weak attachments.

ii) Cadherins depend on calcium and integrins are not calcium dependent.
Cadherins bind to other cells and to the cytoskeleton while integrins
bind the cells to the extracellular matrix and to the cytoskeleton.

3. i) A small molecule that is released by one cell and binds to a complementary

receptor on the membrane of the target cell or in the cytosol.

ii) Juxtacrine signalling happens when there is direct contact between the
signalling molecule on a cell and receptor molecule of another cell while
paracrine signal happens when a signal molecule is released from a cell
and received by another cell which may or may not be a neighbouring
cell.

iii) EMT is responsible for the formation of neural crest cells that are
migratory and mesenchymal in behaviour from the dorsal most
ectodermal cells of neural tube. These cells are epithelial in their
behaviour.

In adults the process of EMT is needed for wound healing,

regeneration of tissue and is the cause of metastasis of cancer cells,
where the solid tumour cells detach from the tumour and migrate to
other parts to form more solid tumours.

4. Statement i)-morphogen; statement ii)-morphogen gradient; statements

iii) and v)- instructional induction; statement iv) -permissive interaction;
statement vi)-competence

5. Distinguishing characteristics

Apoptosis Necosis Autophagy

1. Programmed cell Independent cell Signal comes from
death death not genes
programmed, does
Requires energy
not need energy
2 Membrane of cell not Injury to cell Lysosome mediated
disrupted contents of membrane, recycling of cell
cell do not spill out, unregulated cell contents, to maintain
blebs formed death, contents spill homeostasis
out
3 No inflammation in inflammatory Autosomes formed
surrounding tissue damage to that merge with
removed by surrounding cells, lysosomes that
phagocytes forms scars often breakdown the
has to be removed cellular components
surgically
4 Important in Reaction to toxic For homeostasis and
development of digits event in the body recycling of cellular
by elimination of cell components
populations, 67
Unit 11 Cell to Cell Communication

Terminal Questions
1. Refer to Section 11.3

2. Refer to Subsection 11.2.2

3. i) F; ii) T; iii) T; iv) F; v) F; vi) T; (vii) F; viii) T; ix) T; x) F

4. Refer to Section 11.2

5. To maintain cell numbers in tissue and organs, for homeostasis, to eliminate

extra cells and tissues during morphogenesis and organogenesis e.g., formation
of digits in vertebrates for recycling of cell content in times of need.

6. A signal transduction pathway is the series of events that take place in a target
cell when a signal or ligand is received by a competent receptor on the cell. It is
generally culminated in gene expression or in biochemical reactions inside the
cell or regulate cytoskeleton elements.

Steps in the transduction pathway:

1) When a ligand binds to a transmembrane receptor it changes the shape

of the receptor

2) The receptor activates a protein in the membrane on the

intracellular side

3) Binding to phosphate groups and other enzymatic reactions are

enabled in several steps

4) Ultimately the cell response is effected. If the signal is passed on to the

nucleus, it will activate or repress a gene, or other biochemical reactions
are elicited in the cytoplasm or cytoskeletal components are regulated.

68