References

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References Gilbert (7th edition) Chapter 10,
320-330 Gilbert (6th edition) Chapter 10,
316-329 Wolpert (2nd edition) Chapter 3, 89-104
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Gastrulation in Xenopus (Gilbert fig 10.7)
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Xenopus animal/vegetal axis Nieuwkoop Centre

• In Xenopus, before fertilization, the egg has a
  radial symmetry.
• Sperm entry sets up the D/V axis with the dorsal
  side of the embryo at a place opposite of the
  sperm entry point.
• Within the first 90 minutes after fertilization,
  the cortex rotates approximately 30 degrees.
• The cortex is a gel-like layer of actin
  filaments about 5 micrometres thick just under
  the membrane.
• The vegetal cortex opposite the sperm entry point
  moves towards the animal pole.
• This leads to the formation of a 'signaling
  centre' opposite the site of sperm entry known as
  the Nieuwkoop Centre.

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Wnt signalling
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Nieuwkoop Centre specifies the Spemann Organizer

• The Nieuwkoop Centre's main role is to specify
  another signaling centre
• the Spemann Organizer which in turn leads to the
  induction and patterning of the mesoderm and
  patterning of the ectoderm

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Spemann Organiser
Organization of a secondary axis by dorsal
blastopore lip tissue. (A) Dorsal lip tissue
from an early gastrula is transplanted into
another early gastrula in the region that
normally becomes ventral epidermis. (B) The donor
tissue invaginates and forms a second
archenteron, and then a second embryonic axis.
Both donor and host tissues are seen in the new
neural tube, notochord, and somites. (C)
Eventually, a second embryo forms that is joined
to the host. (D) Structure of the dorsal
blastopore lip region in an early Xenopus
gastrula.
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• Nieuwkoop centre formed in the blastula.
• Cells of the Nieuwkoop Centre contribute to
  endoderm
• The Spemanns Organiser forms just above the
  Nieuwkoop Centre during late blasula, early
  gastrula
• Cells of the Organiser contribute to dorsal
  mesodermal derivatives
• Blastula transplantation experiments explained
  by induction of Organiser

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Induction of mesoderm in Xenopus

• Animal pole explants form ectoderm.
• Vegetal pole explants form endoderm.
• Animal pole explants cultured in contact with
  vegetal tissue produces some mesodermal tissue.
• Therefore, mesoderm depends upon signals from the
  vegetal region to turn animals pole cells from
  ectodermal fate to a mesodermal fate.
• If animal and vegetal tissues are separated by a
  filter, then mesoderm develops.

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The four signal model of mesoderm induction

• Class 1) General mesoderm inducer from the
  vegetal region makes default ventral mesoderm.
• Class 2) Dorsal mesoderm inducer from the vegetal
  region generates Spemann Organizer and
  notochord.
• Class 3) Spemann Organizer signal from dorsal
  region modifies the patterning of ventral
  mesoderm.
• Class 4) Ventral mesoderm patterning signal from
  the ventral region subdivides muscle, kidney and
  blood.

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Induction of mesoderm by endoderm

• At least 2 signals must come from the endoderm
• ventral mesoderm
• dorsal mesoderm (including the Organiser)
• The dorsal mesoderm signal comes from the
  Nieuwkoop Centre
• 1 component of the Nieuwkoop Centre is
  beta-catenin (but this is not the signal!)
• What is the ventralising signal?

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From Wolpert, chapter 3
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An inhibitor of Nodal-related (Xnr) proteins
prevents the formation of the dorsal blastopore
lip
It also inhibits the production of Goosecoid, a
protein that is synthesised just in the Spemanns
Organiser.
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Nodal-related (Xnr proteins)

• Are members of the TGF-b superfamily
• Produced in the vegetal hemisphere
• Diffuse and induce mesoderm in overlying animal
  cells
• Regions with little Xnr proteins develop as
  ventral mesoderm
• Regions with medium Xnr proteins develop as
  lateral mesoderm
• Regions with high Xnr develop as dorsal mesoderm
  (Organiser)

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Spemann Organiser
Organization of a secondary axis by dorsal
blastopore lip tissue. (A) Dorsal lip tissue
from an early gastrula is transplanted into
another early gastrula in the region that
normally becomes ventral epidermis. (B) The donor
tissue invaginates and forms a second
archenteron, and then a second embryonic axis.
Both donor and host tissues are seen in the new
neural tube, notochord, and somites. (C)
Eventually, a second embryo forms that is joined
to the host. (D) Structure of the dorsal
blastopore lip region in an early Xenopus
gastrula.
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Mesoderm induction

• Several secreted proteins are synthesised in the
  Organiser
• These include Noggin, chordin and frizbee
• They are good candidates because
• they cant induce mesoderm in animal cap
  experiments
• But
• they can dorsalise ventral mesoderm

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Rescue of dorsal structures by Noggin protein.
When Xenopus eggs are exposed to ultraviolet
radiation, cortical rotation fails to occur, and
the embryos lack dorsal structures (top). If such
an embryo is injected with noggin mRNA, it
develops dorsal structures in a dosage-related
fashion (top to bottom). If too much noggin
message is injected, the embryo produces dorsal
anterior tissue at the expense of ventral and
posterior tissue, becoming little more than a
head (bottom).
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Ventralising signals

• Candidates are
• BMP4 (TGF-b family). Knock out BMP4 function,
  get dorsalisation of mesoderm.
• Xwnt8. Xwnt8 ventralises mesodermal explants

• The big surprise was that the signals from the
  Organiser (Noggin, Frizbee etc.) interact with
  the ventralising signals
• Noggin interacts with BMP4 and prevent it
  binding to its receptor
• Frizbee interacts with Xwnt8 and prevents it
  binding to its receptor

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Xwnt8 is capable of ventralizing the mesoderm
and preventing anterior head formation in the
ectoderm. (A) Frzb is a protein secreted by the
anterior region of the organizer. It binds to
Xwnt8 before that inducer can bind to its
receptor. Frzb resembles the Wnt-binding domain
of the Wnt receptor (Frizzled protein), but is a
soluble molecule. (B) Xwnt8 is made throughout
the marginal zone.
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The three main stages of vertebrate development

• 1) setting up the main body axes (A/P and D/V)
• 2) specification of three germ layers (endoderm,
  mesoderm and ectoderm)
• 3) germ layer patterning (mesoderm and early
  nervous system)
• Maternal genes provide factor (RNA and proteins)
  to the egg during oogenesis (including
  sub-cellular localization to specific regions.
• Zygotic genes are expressed by the embryo's
  genes.
• Both maternal and zygotic genes may have long
  term effects upon the embryo's development.