molecular basis of morphogenesis

8/6/2019 Molecular Basis of Morphogenesis

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1.Presumptive areas of blastula are predestined for a

specific part in future development.

Transplantation experiments:

1) autoplastic- piece of an embryo transplanted to

another site in the same embryo

2) homoplastic- host (recipient) and donor individuals

are of the same species

3) heteroplastic- host and donor species belong to the

same genus

4) xenoplastic- host and donor species belong to

different genera.

Only autoplastic and homoplastic transplants are

successful in adult vertebrates.

Success in xenoplastic transplants have been achieved in

invertebrates, and between embryos of salamanders and

frogs, mammals and birds.


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2. Presumptive areas have definite normal fates or

prospective significance.

At the time of the stages used in theseexperiments, their fates have not been fixed

3. Ability of the parts of an early embryo to develop in

more than one way is called prospective potency.

Prospective potency of epidermis and neural

system areas of gastrula embryo are practicallyidentical (includes mesodermal, endodermal

tissues), in spite of their different prospective

significance. Grafted material develops in

conformity with its new surroundings. An epidermis area of an early gastrula was

heteroplastically transplanted into the neural

system area of another embryo in the same stage

of development, and vice versa.


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Transplanted epidermis became neural plate of

host; reverse transplantation showed an oppositeresult.

Presumptive ectoderm transplanted into marginal

zone of early gastrula: graft was drawn into

blastopore and found in host interior


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4. At the end gastrulation, prospective potencies ofpresumptive epidermis and neural system areas will

be narrowed down to their prospective significance.

The fixing of the fate of a part of the embryo is

called determination.


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5. The determination of parts of the ectoderm

depends on the surroundings in which theendodermal cells find themselves, and does not

come from causes inherent in the ectoderm itself.

Hilde Mangold- heteroplastic transplant of the

dorsal lip of the blastopore of an early gastrula

to the lateral lip of the blastopore of the host early gastrula resulted to a graft which gave rise

to a secondary embryo in the host.

Comparison of traits reveal that both donor and

host embryos participated in formation of parts

of secondary embryo.

Graft influenced development of certain parts of

embryo (induction); source of influence is called

inductor.


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6. Contact with archenteron roof (chordomesoderm)

determines development of mid-dorsal ectoderm into

the neural tube. Grafts taken from dorsal lip of blastopore and adjoining

marginal zone induce neurulation in early gastrula.

7. Reacting cells must be competent, a particular state

that enable them to differentiate under the influence of

the inductor. Competence of host cells to inductor is highest in the

early gastrula, declines in late gastrula, and fades away

at neurulation. Absence of inductor causes neural

ectoderm to differentiate into epidermis.

8. Due to its ability to induce a complete 2ndary embryowhen transplanted, the amphibian dorsal lip of the

blastopore (also the posterior edge of blastodisc in

fishes, or the anterior half of the primitive in birds) is

called the primary organizer.


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9. The capacity of a certain part of the egg to serve as

the organization center is not fixed at the start of

development, but introduced gradually. Appearance of a definite amphibian organization

center is dependent on interaction of cortical and

subcortical cytoplasm of gray crescent, and the

presence of yolky cytoplasm of vegetal

hemisphere.

The primitive streak in birds is dependent on

underlying hypoblast for its formation;

discrepancies in the orientation of the epiblast

and hypoblast result to primitive developing inaccord with hypoblast orientation.

10.Tissues of mammalian (rabbit) gastrula have

competence for neural induction with chick primitive

streak as an inductor, and vice-versa.


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1. Regional differentiations in the normal embryos are

controlled by a balance between neuralizing andmesodermalizing substances distributed in the form

of gradients.

a.Regional inductions:

Archencephalic (forebrain, eyes and nose

rudiments)- neuralizing (dorsalizing or activating)factor alone

Deuterencephalic (hindbrain and ear vesicles)- low

mesodermalizing + neuralizing factor

Mesodermal (notochord, muscle, kidney and limb-bud) mesodermalizing (caudalizing or

transforming) factor alone

Spinocaudal (trunk organs and tailbud)- high

mesodermalizing factor + low neuralizing factor


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The dorsal lip of blastopore of early gastrula contains head

inductor; same structure at late gastrula contains spinocaudalinductor. The gradient of the neuralizing substance is highest

mid-dorsally and declines towards the lateral and ventral parts

of the embryo. Mesodermalizing substance is concentrated at

the posterior end of embryo and forms a declining gradient both

in the anterior and lateral direction.


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3. Any factor that affects the gradient affect the whole

morphogenetic system.

Defects spread from the front end backward: noserudiments and forebrain, eyes, posterior parts of

brain, gill clefts, ear vesicles.

Raise the level of the dorsoventral gradient by

treatment with sodium thiocyanate: increase in size of

neural system.

Depressing the level of the dorsoventral gradient

gradient at its highest level by exposing embryo to

magnesium or lithium chloride or removing part of the

archenteron roof beneath anterior end of nervoussystem results to injured embryos that develop

cyclopia. Cyclopia and similar defects may result from

toxicosis of mother during early gestation, e.g.

contact with German measles illness or Rubella.


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Babies who have too little of the Sonic hedgehog protein

will be born without noses, and in the most severe cases

cyclopia. An abundance of Shh produces eyes that are far

apart, an abnormally wide nose, or even duplicated facialfeatures, called diprosopus. This can range from only two

noses, to a complete full second face with two mouths, two

noses, and up to four eyes. Each nose has only a single

nostril leading into the sinuses.


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4. Gray crescent area is

the center of gradient

for archencephalicdevelopment:

UV irradiation of gray

crescent caused

microcephaly,

anencephaly. Injection of oocyte

nucleus material or

cytoplasm of

untreated fertilizedegg into the

blastocoele restore

embryo to normal

development.


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Induction is region specific. Otto Mangold

transplanted 4 successive regions of archenteron

roof of late gastrula newt embryos into the

blastocoeles of early gastrula embryos.


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Spemann and Mangold showed that the dorsal lip of the

blastopore constitutes an organizer that has the ability to:

initiate gastrulation movements; dorsalize ectoderm intoneural ectoderm; cause neural plate to become neural

tube; become dorsal mesoderm and notochord; dorsalize

surrounding mesoderm into lateral mesoderm.

The field of molecular biology was refined in the late

1980s, enough to provide a starting point to understand

what genes encoded the products of the organizer.


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In the unfertilized egg, VegT and Wnt 11 mRNA are concentrated in

the vegetal pole. Powered by microtubules and motors atfertilization, the cortical rotation displaces Wnt 11 mRNA opposite

to the sperm entry point. This is crucial for giving the dorsal side

regional identity, and leads to the formation of a signaling center

in the vegetal region called the Nieuwkoop center.

AMPHIBIAN INDUCTION


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Vegetally localized mRNA ofVegT and Vg1, are endoderm-

inducers. VegT directs synthesis of Xnr proteins which aremesoderm-inducing signals. Dorsalizing factors Wnt 11

and Xnr activate the Wnt pathway that leads to the

accumulation of-catenin in the dorsal side. -catenin

acts with VegT to produce higher levels of Xnr.


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Translocation of the Dishevelled protein to the dorsal side of the

egg during rotation stabilizes -catenin in the dorsal cells of the

embryo, which establishes the Nieuwkoop center.


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F-catenin complexes

with Tcf3 which

activates theexpression of the

siamois gene.

The siamois product

and a TGF- signal

((Vg1/ nodalsignaling pathway)

activate the

goosecoid gene

that specifyorganizer genes.

Siamois also

interacts with Xnr to

activate goosecoid.


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The organizer works by secreting inhibitors of

BMP4, e.g., Noggin, chordin, andfollistatin. They

dorsalize the adjacent mesoderm by inhibiting the

ventralizing signal. What is the ventralizing signal?


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BMP signals are powerful ventralizing factors that induce the

expression of epidermal-specific genes. Varying inductionsare created by interaction of BMP4 with BMP antagonists

from the organizer, e.g. low BMP doses activate muscle

formation; intermediate levels instruct cells to become

kidney; high levels activate blood cell and CT formation.


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In the absence ofBMP

signaling, neuralizing

transcription factors

(X1pou2, SoxD) are

produced.

These factors in turn

activate the neurogenin

gene for the transcription

factor NeuroD that causethe differentiation of the

cell into a neuron.

In the presence ofBMP

signaling, epidermalizing

transcription factors(Msx1, GATA1, Vent) are

generated, leading to the

activation of the pathway

enabling the cell to

become a keratinocyte.


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1. In chick embryos, activation of various secreted

factors (Vg1, Nodal, Wnt8C, FGF8 and Chordin);

and transcription factors: (brachyury and goosecoid)

adjacent to the site of streak formation, is required for

the coordinated function of the organizer.

Removal of the hypoblast in the chick results in

correctly patterned ectopic streaks, suggesting that

the hypoblast may inhibit formation of the primitive

streak by secreting an antagonist to Nodal.

2. BMPs do not inhibit neural induction, nor does ectopic

expression ofchordin in the non-neural epiblast causeneural induction.

3. Recent evidence suggests that the anterior visceral

endoderm (AVE) is providing these signals for anterior

neuron production.

AVIAN INDUCTION


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4. FGFs produced in Hensen's node and primitive

streak, appear to induce trunk and hindbrain

neuronal expression in the epiblast cells. Other posteriorizing signals (Wnt3a, retinoic acid,

eFGF) can influence the anterior-posterior

specification of the neural tube.

In the head region, an additional set of proteins

(Cerberus, Frzb, Dickkopf) block these Wnt signal

from the ventral and lateral mesoderm.


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5.Ciliary beating around the

primitive node displaces

FGF8 to the left, causing the

expression of nodal andlefty-1 on the left side of the

streak, thereby activating

the Pitx2 gene responsible

for rotation of the gut and

stomach, spleen and lunglobation.

6.The D-V axis is defined by

the embryonic-abembryonic

(trophoblast opposite ICM)

poles. In birds, gravity iscritical in determining the A-

P axis, while pH differences

appear crucial for

distinguishing dorsal from

ventral.


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In mammals the

primitive node is anembryonic organizer.

(remove HN: no neural

tissue development;

transplant HN to host

embryo: 2nd neuralaxis is induced). The

mammalian node is

not a classic organizer

because it can't act

alone to induce theembryonic axis but

requires other anterior

germ tissues to be

fully effective.

Progressive cellular interactions

characterizes the organizer

activity of the Hensen's node

region in lower vertebrates.


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In mammals, there are 2 signaling centers: one in

the node, and one in the AVE (hypoblast), which

becomes part of the head organizer.

Prechordal plate (head organizer) pass through the

node and become associated with the endoderm

caudal to the oropharyngeal membrane.

They are followed by the chordamesoderm(precursor of the notochord), which is the major

axial signaling center of the trunk.

In birds, the prechordal plate alone can induce head

parts, but in mammals,

the prechordal platemesoderm sustains the

induction of the anterior

visceral endoderm.


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The expression of Nodalfrom the primitive streak drives

the expression of Shh and Goosecoid.

Goosecoid- activates genes of organizer. With SHH,

goosecoid activates expression of HNF-3B, which also

results in the expression of Noggin and Chordin.

Noggin, chordin, follistatin- BMP antagonists from the

organizer, they dorsalize mesoderm. They induce only

forebrain and midbrain types of tissues.


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BMP4- secreted throughout the bilaminar disc, acts

with FGF to ventralize mesoderm into intermediate and

lateral plate structures. The fate of the entire ectodermis dependent upon BMP concentrations: high levels

result in epidermis formation, lower levels at the border

of the neural plate and nonneural ectoderm induce the

neural crest. BMP inhibition leads to neural plate

induction. HNF-3F - from the organizer, this is required for node

formation, initiation of notochord function, and

establishment of midline structures cranial to the node.

T /brachyury - induced by products of HNF-3F and

goosecoid genes, they are necessary for normalmovements of mesodermal cells through the primitive

streak (T gene mutants have gross caudal body defects

in humans). Brachyury also antagonizes BMP4 to

dorsalize mesoderm in caudal regions of the embryo.


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To produce a head, the

BMP4 and Wntsignals are

blocked by transcriptionfactors Otx-2 and Lim-1 (a

homeobox gene); and the

signaling molecule

Cerberus-related 1 HeSXL

(mutants produceheadless phenotype).

WNT3a and FGF - induce

caudal neural plate

structures: hindbrain and

spinal cord. Retinoic acid - organize

the cranial-to-caudal axis

by regulating expression

of homeobox genes.


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Hox genes pattern the

anterior-posterior axis

and help tospecifypositions along that axis.

IfHox genes are knocked

out, segment-specific

malformations can

arise.o Causing the ectopic

expressions of the

same genes can alter

the body axis.o Homology of gene

structure and expressions between Drosophila and

mammalian Hox genes suggest that this patterning

mechanism is evolutionarily ancient.


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Expression patterns of

genes that regulate somitedifferentiation. SHH and

noggin, secreted by thenotochord and floor plate of

the neural tube, cause the

ventral part of the somite to

form sclerotome and toexpress PAX1, which in turn

controls chondrogenesisand vertebrae formation.

WNT proteins from the

dorsal neural tube activate

PAX3, which demarcates thedermomyotome. WNT

proteins also direct thedorsomedial portion of the

somite to form epaxial

(back) muscles and to

express the muscle-specificgene MYF5.

The middorsal portion of the somite is

directed to become dermis by

neurotrophin 3 (NT-3) expressed by thedorsal neural tube. Hypaxial (limb and

body wall) musculature is derived from the

dorsolateral portion of the somite underthe combined influence of activating WNT

proteins and inhibitory BMP4 protein,

which together activate MYOD expression.


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VEGF induced by Shh, this induces

hemangioblasts to form hematopoietic stem

cells, the precursors of all blood cells. Peripheralhemangioblasts differentiate into angioblasts,

the precursors to blood vessels.

PDGF and TGF directs maturation and

modeling of the vasculature until the adultpattern is established.

Notch pathway - induced by VEGF expression,

this specifies arterial development and

suppression of venous cell fate through

expression of ephrinB2.

PROX1 - induces lymphatic vessel differentiation.

Blood vessel and nerve outgrowth appear to

involve guidance factors (e.g., NGFs).


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Blood vessels form in two

ways: vasculogenesis(top), in which vessels

arise from blood islands,

and angiogenesis

(bottom), in which new

vessels sprout from

existing ones. Duringvasculogenesis, FGF2

binds to its receptor on

subpopulations ofmesoderm cells and

induces them to form

hemangioblasts. Then,under the influence of

vascular endothelial

growth factor (VEGF)acting through two

different receptors, these

cells become endothelial

and coalesce to form

vessels.

Angiogenesis is also regulated by VEGF,

which stimulates proliferation of endothelialcells at points where new vessels will

sprout from existing ones. Final modeling

and stabilization of the vasculature areaccomplished by PDGF and TGF-.


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Chordin, activin, nodal, sonic

hedgehog are also involved

in the L-R asymmetry of

certain body parts. The

protein activin, a member of

the TGF- signaling family,

directs development of parts

in the right axis. There are over 24 genes

currently known for L-R

assymetry, and gene

mutations downstream in the

cascade are responsible forright-left switching for

various organs (e.g., heart as

seen in situs inversus,

Kartageners syndrome).


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The Vg1 protein

activates a Nodal

protein solely on the

left side of the body. Asin other vertebrates,

the Nodal protein

activates expression of

Pitx2, which is critical

in distinguishing left-

sidedness from right-sidedness in the heart

and gut tubes.

Activation of transcription factor Pitx2 regulates genes involved in

lateral plate mesoderm differentiation. Other factors such as Shh,

FGF8, Lrd and Kifare also involved. The nodal flow model suggeststhat morphogens and signaling molecules involved in defining the L-

R axes is driven by the action of cilia that cause molecules to flow

from right to left across the primitive node. Cells in the node with

mutations that affect ciliary action (e.g. null mutations for Lrd which

encodes ciliary dynein) alter L-R symmetry.


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ERRORS OF GASTRULATION


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Spinal cordAbnormalities


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Little Samuel Armas was diagnosed with spina bifida and would not survive if

removed from his mother's womb. His mother knew of Dr. Bruner's remarkable

surgical procedure: he performs these special operations while the baby is still in

the womb. 'The tiny hand of the 21-week-old fetus emerges from the mother's uterus

to grasp the finger of Dr. Joseph Bruner as if thanking the doctor for the gift of life.

Our God is an awesome God!


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molecular basis of morphogenesis

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