Study Guide
Chapter 11 Early Development Cleavage, Gastrulation and Axis Formation Amphibians and Fish
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University:
Kentucky State University -
Course:
BIO 111 | Principles of Biology Academic year:
2019
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Views:
139
Pages:
53
Author:
Caroline Mcfarland
Chapter 11
Early development: Cleavage, Gastrulation and
Axis Formation
Amphibians and Fish Fertilization, cortical rotation, and cleavage
• The point of sperm entry marks the ventral side, while the site 180° opposite to it marks the
dorsal side.
• The sperm centriole organizes the microtubules of the egg into parallel tracks in the vegetal shear
zone Fertilization, cortical rotation, and cleavage
• The parallel microtubular tracks organized in the shear zone allow the cortical cytoplasm to rotate
about 30° with respect to the internal cytoplasm.
• In some amphibians, this rotation exposes the gray crescent: a region of gray colored inner
cytoplasm directly opposite the sperm entry point. Gastrulation starts in this region. Unequal radial holoblastic cleavage
• Cleavage in most frog and salamander embryos is radially symmetrical and holoblastic. Unequal radial holoblastic cleavage
• Although, the micromeres and macromeres are unequal in size, they continue to divide at about
the same rate until the twelfth division.
• An amphibian embryo containing 16 to 64 cells is commonly called morula.
• At 128 cell stage, the blastocoel becomes apparent, and the embryo is called a blastula. Unequal radial holoblastic cleavage
The amphibian blastocoel serves two major functions:
1. It can change its shape such that cell migration can occur during gastrulation.
2. It prevents the cells beneath it from interacting prematurely with the cells above it The mid-blastula transition: preparing for gastrulation
The mid blastula transition: The transition from the early rapid cell cycle (only M and S phases) of
the embryo to a stage characterized by:
(1) Cell cycle that includes the “gap” stages (G1 and G2) in addition to M and S phases.
(2) Loss of synchronicity of cell division
(3) Transcription of new (zygotic) mRNAs needed for gastrulation and cell specifications Amphibian Gastrulation
Vegetal rotation and the invagination of the bottle cells
• In amphibians, the cell movements of gastrulation are initiated on the future dorsal side of the
embryo, just below the equator. Amphibian Gastrulation
Vegetal rotation and the invagination of the bottle cells
• Initially, the cells invaginate to form the slitlike blastopore.
• Marginal zone: the region surrounding the equator of the blastula, where the animal and vegetal
hemispheres meet
Superficial endodermal
Cells of margin zone Amphibian Gastrulation
Vegetal rotation and the invagination of the bottle cells
• Vegetal rotation: during frog gastrulation, internal cell rearrangements place the prospective
pharyngeal endoderm cells adjacent to the dorsal blastocoel and immediately above the involuting
mesoderm Amphibian Gastrulation
Vegetal rotation and the invagination of the bottle cells
• The prospective pharyngeal endoderm cells then migrate along the basal surface of the blastocoel
roof, traveling toward the anterior of the embryo.
Superficial layer
of marginal cells Amphibian Gastrulation
Involution at the blastopore lip
• Notochord: the transient mesodermal rod that plays an important role in inducing and patterning
the nervous system.
• The cells constituting the dorsal blastopore lip are constantly changing as the original cells migrate
into the embryo and are replaced by cells migrating downward, inward, and upward.
Xenopus brachyury Amphibian Gastrulation
Involution at the blastopore lip
• Yolk plug: the large endodermal cells that remain exposed on the vegetal surface surrounded by
the blastopore of the amphibian gastrulating embryo. Amphibian Gastrulation
Involution at the blastopore lip
• The yolk plug, eventually internalized. Amphibian Gastrulation
Convergent extension of the dorsal mesoderm
• Shortly before their involution through the blastopore lip, several layers of deep IMZ cells
intercalate radially to form one thin broad layer. Amphibian Gastrulation
Convergent extension of the dorsal mesoderm
• When the deep cells of IMZ reach the blastopore lip, they involute into the embryo and initiate a
second type of intercalation called the mediolateral intercalation.
• This intercalation causes convergent extension along the mediolateral axis that integrates
several mesodermal streams to form a long, narrow band.
Streams of
Narrow band
mesodermal cells Amphibian Gastrulation
Convergent extension of the dorsal mesoderm
• The radial and mediolateral intercalations of the deep layer of cells appear to be responsible for
the continual movement of mesoderm into the embryo.
Superficial IMZ cells Amphibian Gastrulation
Convergent extension of the dorsal mesoderm
• Those mesodermal cells entering through the dorsal lip of the blastopore give rise to the central
dorsal mesoderm (notochord and somite).
• Those mesodermal cells entering through the ventral and lateral blastopore lips create the
mesodermal mantle.
• Mesodermal mantle: the cells that involute through the ventral and lateral blastopore lips during
amphibian gastrulation and form the heart, kidneys, bones, and part of several other organs. Amphibian Gastrulation
Convergent extension of the dorsal mesoderm
• The endoderm is derived from the prospective pharyngeal endoderm,
superficial cells of IMZ and the subblastoporal vegetal cells.
• The superficial cells of the IMZ form the lining of the archenteron roof.
• The subblastoporal vegetal cells become the archenteron floor.
• The remnant of the blastopore- where the endoderm meets the
ectoderm- becomes the anus. Amphibian Gastrulation
Epiboly of the prospective ectoderm
The mechanisms that drive epiboly during Xenopus gastrulation:
1- An increase in cell number (through division) coupled with a concurrent integration of several
deep layers into one. Amphibian Gastrulation
Epiboly of the prospective ectoderm
The mechanisms that drive epiboly during Xenopus gastrulation:
2- The assembly of an extracellular fibronectin lattice secreted by the presumptive ectoderm cells of
the blastocoel roof.
• This fibronectin lattice allows the vegetal migration of the animal cap cells and enclosure of the
embryo The work of Han Spemann and Hilde Mangold
primary embryonic induction
• Spemann and Mangold showed that of all tissues in early gastrula, only one has its fate
autonomously determined.
• This self-determined tissue is the dorsal lip of the blastopore.
• When the tissue was transplanted into presumptive ventral epidermis of another gastrula, it not
only continued to be dorsal lip but also initiated gastrulation and embryogenesis in the
surrounding tissue. The work of Hans Spemann and Hilde Mangold
primary embryonic induction
• The dorsal blastopore lip of an early gastrula was removed and transplanted into a presumptive
ventral epidermis of another early gastrula.
• Eventually, a secondary embryo formed, conjugated face to face with the host embryo. The work of Hans Spemann and Hilde Mangold
primary embryonic induction
• Similar results were obtained in other amphibian species including Xenopus.
Twined larvae The work of Hans Spemann and Hilde Mangold
primary embryonic induction
• Spemann referred to the dorsal lip cells and their derivatives (notochord and head
endomesoderm) as the organizer because:
1. They induced the host’s ventral tissue to change their fates to form a neural tube and dorsal
mesodermal tissues (such as somites)
2. They organized host and donor tissues into a secondary embryo with clear anterior-posterior
and dorsal-ventral axes. The work of Hans Spemann and Hilde Mangold
primary embryonic induction
• In amphibian embryos, the primary embryonic induction refers to the process
whereby the dorsal axis and central nervous system form through interactions with
the underlying mesoderm, derived from the dorsal lip of the blastopore Molecular mechanisms of amphibian axis formation
The dorsal signal: β- catenin
• Experiments by Pieter Nieuwkoop and Osamu Nakamura showed that the organizer
receives its special properties from signals coming from the prospective endoderm
beneath it.
• Nieuwkoop and Nakamura laboratories demonstrated that the properties of this newly
formed mesoderm can be induced by the vegetal (presumptive endoderm) cells
underlying them. Molecular mechanisms of amphibian axis formation
The dorsal signal: β- catenin
• The polarity of this induction depended on whether the endodermal (vegetal) fragment
was taken from the dorsal or ventral side.
• Those dorsalmost vegetal cells of the blastula, which are capable of inducing the
organizer, have been called the Nieuwkoop center. Molecular mechanisms of amphibian axis formation
The dorsal signal: β- catenin
• The Nieuwkoop center was demonstrated in the Xenopus
embryo by transplantation experiments. Molecular mechanisms of amphibian axis formation
The dorsal signal: β- catenin
• The Nieuwkoop center was demonstrated
in the Xenopus embryo by recombination
experiments. Molecular mechanisms of amphibian axis formation
The dorsal signal: β- catenin
• Thus, both the transplantation and recombination experiments demonstrated that the
dorsalmost vegetal cells can induce animal cells to become dorsal mesodermal tissues.
• These dorsalmost vegetal cells constitute the Nieuwkoop center. Molecular mechanisms of amphibian axis formation
The dorsal signal: β- catenin
What gives the dorsalmost vegetal cells their special properties?
• β- catenin is the major candidate for the factor that forms the Nieuwkoop center in the vegetal
cells.
• The accumulation of β- catenin is seen in the nuclei of dorsal cells and it appears to cover
both the organizer and the Nieuwkoop center Molecular mechanisms of amphibian axis formation
The dorsal signal: β- catenin
If β- catenin is originally found throughout the embryo, how does
it become localized specifically to the side opposite the point of
sperm entry?
Due to localization of two proteins GSK3 binding protein (GBP) and
Disheveled (Dsh) as well as Wnt11 mRNA from the vegetal pole of the
egg to the future dorsal side of the embryo during fertilization. Molecular mechanisms of amphibian axis formation
The dorsal signal: β- catenin
• But the mere translocation of GBP and Dsh does not appear to be
sufficient to protect β- catenin from degradation in the dorsal side.
• Wnt11 causes the production of new Dsh proteins.
• Wnt11 also stabilizes the reaction in which GBP and Dsh bind to and
block GSK3, thereby preventing the degradation of β- catenin Molecular mechanisms of amphibian axis formation
The dorsal signal: β- catenin
• During cleavage, β- catenin enters the nucleus and
interacts with Tcf3 transcription factor converting it
from a repressor into an activator of transcription Molecular mechanisms of amphibian axis formation
The vegetal nodal related signal
• In the organizer, Siamois and Twin interact with the phosphorylated Smad2 and then activates
the organizer genes.
But how does Smad2 itself become activated?
Xenopus Nodal-related (Xnr)
paracrine factors Molecular mechanisms of amphibian axis formation
The vegetal nodal related signal
But how is the gradient of Nodal-related proteins generated?
• In the most dorsal endodermal cells (Nieuwkoop center) β- catenin acts synergistically with Vg1
and VegT to activate the Xenopus nodal-related genes (Xnr1, 5 and 6).
• This creates a gradient of Xnr proteins across the endoderm Molecular mechanisms of amphibian axis formation
The vegetal nodal related signal
• In summary, the formation of the dorsal mesoderm and the organizer originates through the
action of two pathways:
1. The Wnt/ β- catenin pathway that activates genes encoding the Siamois and Twin transcription
factors.
2. The vegetal pathway that activates the expression of Xnr factors, which in turns activate Smad2
transcription factor in the mesodermal cells above them.
•
The high levels of Smad2 and Siamois/ Twin transcription factors work within the dorsal
mesoderm cells and activate the genes that give these cells their organizer properties. Molecular mechanisms of amphibian axis formation
Functions of the organizer
The cells of the organizer ultimately contribute to four cell types:
1. Pharyngeal endoderm.
2. Head mesoderm (prechordal plate).
3. Dorsal mesoderm (primarily the notochord).
4. The dorsal blastopore lip.
•
The dorsal blastopore lip eventually becomes the chordaneural
hinge that induces the tip of the tail. Molecular mechanisms of amphibian axis formation
Functions of the organizer
The properties of the organizer tissue can be divided into four major functions:
1. The ability to self differentiate into dorsal mesoderm (prechordal plate, chordamesoderm,
etc.).
2. The ability to dorsalize the surrounding mesoderm into paraxial (somite forming)
mesoderm.
3. The ability to dorsalize the ectoderm and induce the formation of neural tube.
4. The ability to initiate the movement of gastrulation.
Paraxial mesoderm
Organizer derivatives Molecular mechanisms of amphibian axis formation
Induction of neural ectoderm and dorsal mesoderm: BMP inhibitors
• It is the epidermis that is induced to form and not the neural tissue.
• The “default fate” of the ectoderm is to become neural tissue.
• Certain parts of the embryo induce the ectoderm to become epidermal tissue by secreting
bone morphogenetic proteins (BMPs).
• The organizer tissue acts by secreting molecules that
block BMPs, therefore, allowing the ectoderm “protected” by
the BMP inhibitors to become neural tissue.
Norrin The regional specificity of neural induction along the anterior- posterior axis
• The specifications of the anterior- posterior axis is predicted based on the gradient of Wnt
protein, with head being the area of the lowest concentrations of Wnts. The regional specificity of neural induction along the anterior- posterior axis
The head inducer: Wnt antagonist
• The most anterior regions of the head and brain is underlain by pharyngeal endoderm and
head (prechordal) mesoderm. The regional specificity of neural induction along the anterior- posterior axis
The head inducer: Wnt antagonist
• The functions of paracrine antagonists in the ectoderm The regional specificity of neural induction along the anterior- posterior axis
Trunk patterning: Wnt signals and retinoic acid
• Recent studies showed that the Wnt proteins especially
Wnt8 act as posteriorizing molecules that posteriorize the
neural tube.
• The Wnt proteins form a gradient that is highest in the
posterior of the embryo and weakened anteriorly. The regional specificity of neural induction along the anterior- posterior axis
Trunk patterning: Wnt signals and retinoic acid
XWnt8 overexpression
Forebrain
Spinal cord neurons
Spinal cord neurons
Spinal cord neurons
XWnt8 suppression
Forebrain
Spinal cord neurons
Forebrain
Most anterior marker of the forebrain The regional specificity of neural induction along the anterior- posterior axis
Trunk patterning: Wnt signals and retinoic acid
There are two major gradients in the amphibian gastrula:
• BMP gradient that specifies the dorsal- ventral axis
• Wnt gradient that specifies the anterior- posterior axis The regional specificity of neural induction along the anterior- posterior axis
Trunk patterning: Wnt signals and retinoic acid
• The dorsal- ventral and posterior- anterior axes, are established by the initial axes of Xenopus
nodal related (Xnr) factors and β- catenin in the vegetal cells.
• Nodal related factors activated by β- catenin and VegT/Vg1 causes the expression of high levels
of activated Smad2 in the dorsal mesoderm (organizer).
• Smad2 interacts with Siamois/Twin to induce the expression of the genes that encode for Wnt and
BMP blockers The regional specificity of neural induction along the anterior- posterior axis
Trunk patterning: Wnt signals and retinoic acid
• A model of organizer function and axis specification in the Xenopus gastrula. The regional specificity of neural induction along the anterior- posterior axis
Trunk patterning: Wnt signals and retinoic acid
(Fibroblast growth factors)
(Retinoic acid) The regional specificity of neural induction along the anterior- posterior axis
Trunk patterning: Wnt signals and retinoic acid
Functions of retinoic acid (RA):
1. Pattern the hindbrain, as it appears to interact with FGF signals to activate the posterior Hox
genes.
2. RA is critical in allowing the growth of the most posterior part of the tadpole, the tail
• Together, the Wnt, FGF and RA gradients function to determine the boundaries of Hox genes
along the anterior- posterior axis, which specify the regions of the neural tube
Tail bud of developing tadpole
Undifferentiated caudal mesoderm cells
RA synthesis extends posteriorly Specifying the left- right axis
• In all vertebrates, the crucial event in the left- right
axis formation is the expression of a Nodal gene in
the lateral plate mesoderm on the left side of the
embryo.
Xnr1 is expressed
in the left
Left
Right
The expression of Xnr1
is reversed
Left
Right
• In Xenopus, this gene is Xenopus nodal- related 1
(Xnr1), and it is expressed on the left side of the
tadpole.
• The heart in the tadpole is found on the left side and
the heart loops to the right.
• The gut coiling is counterclock wise.
Counter clockwise
clockwise Specifying the left- right axis
• Xnr1 protein activates the expression of a gene that is normally expressed on the left side
only and encodes the transcription factor Pitx2.
• Pitx2 determines the direction of heart looping and gut coiling.
Chapter 11 Early Development Cleavage, Gastrulation and Axis Formation Amphibians and Fish
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