The notochord is transient & becomes part of spinal column.
With neuralation, ectoderm overlaying the
notochord forms the neural tube to give rise to brain & spinal cord.
Somites, mesoderm in blocks flanking notochord, become the ...
1) vertebrae and ribs
2) muscles of the trunk and limbs and
3) contribute to the dermis.
Neural crest cells
Neural crest cells migrate away from the neural tube to develop into...
1) skull (bone)
2) sensory and autonomic nervous systems
3) pigment cells
Somites are formed after gastrulation along the antero-posterior axis.
Notochord and somite development in chick
In the chick, mesoderm forms anterior to the
regressing node of the primitive streak.
Pre-somitic mesoderm is the region between the last formed somite and
the regressing node.
This region will become 4 or 5 somites which form simultaneously as
pairs on either side of the notochord.
The position of somites along A/P axis determines fate.
Anterior somites form cervical vertebrate.
Posterior ones, ribbed thoracic vertebrate.
Develop in a temporal and spatial order.
Rearranging pre-somitic mesoderm will not
change the pre-established timing.
The pattern is laid down earlier by an A/P
axis signal.
Mesoderm and homeobox genes
Homeobox genes are...
a large family of transcription factors.
Share a similar 60 amino acid DNA binding homeodomain which is encoded
by 180 basepair homeobox sequence.
Homeobox gene family (transcription factor proteins).
Homeotic transformation is often observed
in mutants of genes that have this domain.
Identified first in Drosophila (Bithorax and Antennapedia complexes)
as a split cluster.
There are four separate clusters of Hox genes (subset of the homeobox
genes) in vertebrates.
Hox gene clusters
Hox genes ( Hox gene clusters) are a subset
of the homeobox genes of transcription factor genes.
Might have arisen by rounds of duplication of an ancestral gene, followed
by a quaduplication of the cluster in mammals.
Paralogous groups are composed of the most
similar members of each cluster.
Partially overlapping zones of expression
which vary in the anterior extent of their expression define distinct regions.
Various genes respond to the combination of gene products expressed.
Most homeobox genes are not Hox genes (i.e. Pax genes)
Hox genes pattern the A/P axis
The differences between vertebrate (i.e. anterior -attach to
skull; cervical; thoracic have ribs; lumbar, sacral and caudal) clearly
demonstrate that identity of somites differ along the A/P axis.
Hox genes are expressed along the A/P axis in mouse.
First, anterior Hox genes expressed in early gastrulation as mesoderm
begins to leave the primitive streak.
More posterior Hox genes turn on as development continues.
Defined patterns of Hox gene expression are seen in...
1) mesoderm (after somite formation) &
2) neural tube (neuralation).
Hox genes pattern the somites
Hox genes show a sharp anterior border and a much less defined posterior
border.
Lot of overlap by every region (almost) has distinct set of Hox gene
expression.
Most anterior somites express Hoxa1 and Hoxb1 only.
Posterior regions express all Hox genes.
The anterior head, forebrain and midbrain do not express Hox genes
but have other homeobox genes (etx & otx.)
For example, the Hoxa complex has members that are expressed very differently.
Hox gene expression is co-linear
Hoxa1 has its most anterior expression in the posterior head.
Hoxa11 has its most anterior expression in the sacral (lower back)
region.
Hox gene expression is co-linear as order of genes on the chromosome
(per cluster) reflects the order of spatial and temporal expression along
the A/P axis.
Hox gene expression is conserved between mouse and chick.
Mesoderm becomes notochord and somites
The fate of somites depend upon adjacent tissues
signals.
In chick-quail trans-species grafts (with distinctive
nuclei), somite fate maps have been constructed.
Dermamyotome is dorsal & lateral somites (express Pax3) and becomes
the myotome (forms muscle cells) and the dermatome (an epithelial sheet
that forms dermis).
Medial somites (MyoD) forms axial & back muscles.
Lateral region of somites form abdominal & limb muscles.
Ventral medial region of the somite contain sclerotome cells (future
cartilage which express Pax1) and migrate to surround the notochord and
form the vertebrate.
Notochord induces sclerotome cells.
From notochord transplantation experiments, an additional notochord
induces
unsegmented pre-somitic mesoderm
to produce greatly increased amount of cartilage.
In the mouse, FGF and retinoic acid gradients
help pattern the A/P axis.
Neural tube (ventral side: the floor plate) induces
cartilage.
Lateral plate mesoderm and the ectoderm induce the dermamyotome.
Signals that may pattern the somites are secreted signaling proteins
that may include...
Sonic hedgehog which may specify the ventral somites.
BMP-4 which may specify the lateral somites.
Wnt family proteins which may specify the dorsal somites.
Regulation of the Pax homeobox genes (transcription factors)
Pax genes are regulated by signals from the notochord and neural tube
to control the somitic cell fate.
Pax3 is expressed early in all cells that will form somites.
Pax3 is modulated by BMP-4 and Wnt to confine it to muscle precursors.
Pax3 is further down regulated in back muscle precursors but remains
active in future limb muscle cells.
In mice, Splotch (Pax3-minus) mutants lack limb muscles.
Technique: Insertional mutagenesis (gene knock-out)
A targeting vector is constructed that has the central (functional)
region of a gene replaced with a drug resistance gene.
This is transfected into ES cells and selected by drug exposure.
By homologous recombination, a fraction
of the transformants will have one copy of the original gene replaced with
the altered (non-functional) form.
These cells are injected into the inner cell mass of a blastocyst.
Resultant chimeric mice give rise to heterozygous mutants which can
be bred to generate mutant homozygotes.
Altering Hox gene expression alters axial patterning
In mice, gene knock-out experiments produce mutants.
There is redundancy, where a missing gene can be at least partially
compensated for the expression of related genes.
Paralogous genes from another Hox complex may compensate for gene loss.
Posterior prevalence: mutation affects the anterior extent of
gene expression.
Homeotic transformations (conversion of one body part to another) result
from Hox gene loss.
Loss leads to cells assuming a "more anterior value" i.e. Hoxc8 mutant
mice have extra ribs.
Abnormal expression of Hox genes in anterior regions lead to tissues
becoming more like posterior positioned tissues.
Retinoic acid can alter positional value
Retinoic acid is a derivative of vitamin A.
It has very important role in signaling vertebrate development.
In early development, retinoic acid can cause homeotic transformation
of the vertebrate.
It can diffuse across plasma membranes to bind protein receptors and
form an active transcription factor.
Retinoic acid interferes with the normal expression of Hox genes.
Later, it can alter positional development in limb development.
The Spemann Organizer has conserved functions
As gastrulation proceeds, the A/P axis become specified and as it progresses
the blastopore lip can only induce more posterior
structures.
Hensen's node (the chick Organizer), at the anterior of the primitive
streak contributes to notochord and somites & can
induce another axis (more difficult than frogs)
Xenopus and mouse share a number of genes
that are expressed in the organizer.
Brachyury
FGF (fibroblast growth factor)
Control of Hox genes is unknown.
Neural plate is induced by mesoderm
Dorsal lip transplantation experiments demonstrate that a nervous system
can be induced from ectoderm.
BMP-4, secreted growth factor, inhibits cells from forming neural tissue.
Inhibition of BMP-4 allows neural tissue.
noggin (secreted by the organizer) inhibits BMP-4 acts to dorsalize
the mesoderm.
noggin also induces neural tissue.
chordin, expressed by the future neural plate cells of the organizer.
Both noggin and chordin directly bind BMP-4 and inactive it to allow
the induction of neural tissue.
Zebrafish spinal cord is distant from organizer
Ectoderm that gives rise to spinal cord is far from the organizer in
zebrafish.
This is induced in a two-step manner:
1) FGF from ventral-vegetal region induce neurectoderm, then
2) BMP promote posterior neural tissue formation.
Neural plate is induced by signaling mechanism(s)
Nervous system can be patterned by signals from the mesoderm
In newt neurula, mesoderm transplantation
into younger newt embryos, anterior explants induce head and brain.
Posterior explants induce trunk & spinal cord.
Neural plate explants induce specific neural structures (depending
upon position) when transplanted beneath the ectoderm of a gastrula.
Hensen's node is the chick organizer
Hensen's node (chick) can induce neural gene expression in Xenopus
ectoderm.
This demonstrates the evolutionary conservation
of neural induction signals and confirm similarity of Hensen's node and
Spemann's Organizer.
Early nodes induce anterior structures.
Latter nodes induce posterior structures.
Stem cells that arise from the node can specify different A/P positional
values over time to produce the spinal column.
The capacity to produce signals that generate anterior structure is
lost.
Model: The two signal model of neural patterning
Signal 1 from the mesoderm induces ectoderm to become anterior neural
tissue. (chordin and noggin are good candidates)
Signal 2 turns part of this into posterior neural tissue in a graded
manner (FGF, Wnts & retinoic acid are candidates).
Mouse and chick grafts of the primitive streak (ie node or Hensen's
node) can also induce neural tissue.
This model differs from another model which suggests that there may
exist a number of region specific inducer molecules.
Neural plate signals travel within the neural plate
Mesoderm does not have to lie in contact with ectoderm to induce it.
In newt and Xenopus embryos under high salt, the mesoderm does not
enter the embryo but develops outside.
This physically separates the mesoderm from the ectoderm.
This abnormal embryo is called an exogastrula.
N-CAM, a neural cell-cell adhesion protein, neurogenic factors and
other neural specific proteins can be expressed in the ectoderm in the
correct A/P order in exogastrula which suggests that the inducing signal
can travel a relatively long distance through the tissues.
Hindbrain rhombomeres restrict cell lineage
Posterior head & hindbrain development requires segmentation
of the anterior neural tube.
Segmentation events in the 3 day chick embryo's posterior head include...
1) somite formation from mesoderm on either side of notochord,
2) the hindbrain (rhombocephalon) is divided into 8 rhombomeres, and
3) the lateral mesoderm forms the branchial arches.
(Note: spinal cord is segmented into dorsal root ganglia and
ventral motor nerves by the somites)
Development of posterior head involves interactions
Neural crest cells innervate the face & neck to form the segmental
cranial nerves.
Neural crest cells also give rise to peripheral nerves and bones including
jaw (from the first branchial arch) and the bony parts of the ear from
the second arch).
Eight rhombomeres form by constricting the freshly closed neural tube
into eight evenly spaced sections.
Lineage restriction occurs with cells and their descendants remaining
in their rhombomere.
Cell movement restriction depends on adhesive properties which depends
upon ephrins and their receptors.
Within each rhombomere, the cells are under the control of the same
genes & act as a developmental unit.
Neural crest cells have positional values
Chick neural crest cells can be labled & their fate mapped.
The cranial neural crest cells migrate out
from the rhombomeres of the dorsal region of the hindbrain.
Branchial arch 1 is populated by cells from rhombomere 2,
Branchial arch 2 by rhombomere 4 cells &
Branchial arch 3 by rhombomere 6 cells.
Neural crest cells from rhombomeres 3 & 5 die by apoptosis (programmed
cell death).
Transplantation of rhombomere 2 cells to where rhombomere 4 cells should
be, results in formation of a second jaw.
Hindbrain Hox gene expression
Mouse embryonic hindbrain Hox gene expression is well defined
and correlates well the segmental pattern.
Paralogous group 1 (Hoxa1, b1 etc.) is expressed anterior to paralog
2 (Hoxa2, b2 etc.) followed by paralog 3.
Control of gene expression is complex (i.e. Hoxb2
is controlled in rhombomere 3 & 5 by one enhancer which is activated
by Krox-20 and in r4 by another enhancer).
Hoxa2 knock-out mice show head skeletal defects that are consistent
with a homeotic transformation of the inner ear (2nd branchial arch) to
jaw (1st arch).
(By neurula stage, the embryo is divided
into regulating organ-forming regions.)
email me at bestave@mun.ca
