Download lecture - McLoon Lab

Regionalization of the
nervous system 1
September 19, 2016
Yasushi Nakagawa
Department of Neuroscience
Complex organization
of the adult brain
forebrain
midbrain
Hippocampus
ex
cort
Neo
Ce
reb
hindbrain
ell
um
Midbrain
Thalamus
Olfactory bulb
Striatum
Pons
Medulla
Hypothalamus
Allen Brain Atlas
http://www.brain-map.org/
How are all these regions established?
The neural plate
has a two-dimensional structure
anterior (rostral)
posterior (caudal)
lateral
medial
medial
lateral
The neural plate
has a two-dimensional structure
Medial-lateral axis becomes ventraldorsal axis after neural tube closure
anterior (rostral)
lateral
medial
medial
lateral
anterior
(rostral)
dorsal
ventral
posterior (caudal)
posterior
(caudal)
Fate map of the neural plate
10.2 NEURAL PLATE, CLONAL STRUCTURE, AND TOPOLOGIC SPATIAL DIMENSIONS
AHP
Sbpall
OP
Septal roof
ac Poa
Eye
Pall
FIGURE 10.2
Secondary
prosencephalon
Model of neural plate topology of
the fundamental AP and DV subdivisions. Some parts
are colored for contrast. The spinal cord is shortened
for simplicity.
THy
Forebrain choroidal
roof
hp2
PHy
hp
m
rm sth
forebrain
hp1
PTh
p3
Diencephalon
zli
p2
Alar–basal
boundary
RP
Isthmic
organizer
AP
BP
Th
Ep
p1 PT
FP
m1
Midbrain
m2
Cb
Prepontine
Pontine
midbrain
Hindbrain choroidal roof
Hindbrain
hindbrain
Pontomedullary
Medullary
Spinal cord
spinal cord
Telencephalon
Alar hypothalamus
Basal hypothalamus
191
Puelles (2013)
being ‘epichordal.’ This viewpoint, recently incorporated
into the prosomeric model (Puelles et al., 2012a), importantly implies that the entire forebrain including the
hypothalamus and the telencephalon is fundamentally
ing topologically from ventral (floor) to dorsal (roof).
diverse prospective subregions within the hypotha
mus therefore can be interpreted conveniently, even
paradoxically, as being all equally rostralmost, akin
Subdivisions at later stages
TG
SC
IC
ch
Tel (evaginated)
alon
eph
c
en
Th
p2
zli
PT
p1
Pal
10.2 NEURAL PLATE, CLONAL STRUCTURE, AND TOPOLOGIC SPATIAL DIMENSIONS
AHP
Sbpall
OP
Septal roof
ac Poa
Eye
Pall
FIGURE 10.2
Secondary
prosencephalon
Septal
roof plate
PHy
m
rm sth
hp1
zli
p2
RP
Isthmic
organizer
AP
BP
Th
Ep
hp2
ac
Eye
Midbrain
m1
m2
Cb
ntia nigra
isth
Cephalic
flexure
cbc
r1
r2
ch
r4
PHy
NH
Hindbrain
r3
Pons
r5
r6
r7
r8
r9 r10
r11
p1 PT
FP
m1
Midbrain
m2
Cb
POA
u
ta
bs
RM
PRM
M
hp1 RTu
PM
THy
Pa
SPa
Tu
PTh
p3
Diencephalon
Alar–basal
boundary
Dg
Forebrain choroidal
roof
hp
ZI
Model of neural plate topology of
the fundamental AP and DV subdivisions. Some parts
are colored for contrast. The spinal cord is shortened
for simplicity.
THy
hp2
PTh 191
p3
S
Di
PThE
PIsth
pc
Hb
St
BIC
E
hbc
Prepontine
Pontine
Hindbrain choroidal roof
Hindbrain
Pontomedullary
Where are the AP and DV axes?
Spinal cord
Medullary
Spinal cord
Telencephalon
Alar hypothalamus
Basal hypothalamus
FIGURE 10.3
Model of further subdivisions (including neuromeric ones) in the neural tube. Roof and floor Puelles
plate areas
are in gray (AC, ante
(2013)
commissure), and choroidal roof tissue in black. The colored areas are the same marked in Figure 10.2, adding the pontine hindbrain region in b
Regionalization generates diversity
in cell types in the nervous system
Cells in early neural tissue acquire identity that is appropriate for their location
(“positional identity”). This process is called regionalization.
Positional identity contributes to the generation of different types of neurons
and glial cells.
Key molecular mechanisms of regionalization in the nervous system are:
-shared with embryonic patterning in Drosophila
-used over and over throughout brain development as well as in the adult
brain.
Outline of lectures
Regionalization 1 (9/19/15)
neural induction and early regionalization along AP axis
AP patterning in Drosophila embryos as a model of regionalization
Regionalization 2 (9/21/15)
AP patterning by secondary organizers
DV patterning
Research talk (10/3/15)
patterning, cell division and cell fate regulation in the mouse thalamus
How does AP patterning occur in
early neural tissue?
wires.wiley.com/devbio
Advanced Review
(a)
(b)
Ozair, Kintner, Brivanlou (2013)
FIGURE 3 | Schematic of graded BMP activity in the gastrula and neurula ectoderm. (a) A schematic fate map of the early gastrula shows the
approximate positions of the future neural plate (NP), border region, and epidermis, viewed from the dorsal side. The cement gland (CG) and sensory
placodes form in the anterior border region mid-dorsally, whereas the neural crest arises more laterally. Diffusible antagonists produced in the
organizer region of the mesoderm, including noggin, chordin, and follistatin, result in a graded distribution of BMP signaling in the neighboring
ectoderm. The relative position of epidermis (EP), NP, organizer (O, in blue), CG, and neural crest (NC) is shown. Sensory placodes form at various
positions in the border region but are not shown here for simplicity. (b) Correlation with neurula fate map shown in Figure 1.
-BMP inhibitors (noggin, chordin, follistatin) secreted from the organizer tissue
play a central role in neural induction.
-Does BMP inhibitors also control AP patterning?
-Does
as well as segregation of the ectoderm into neuroEvolutionary Conservation of Molecular
genic and epidermal territories. Exposure of these
Circuitry
Underlying
Neural
Induction
organizer produce other molecules that control AP patterning?
-Are there
embryos to exogenous BMPs does not repress neural
Inhibition of ongoing TGFβ signaling to delineate
markers, and conversely, BMP knockdown does not
neural and non-neural ectoderm has been conserved
promote neuralization, even though it has a role in
evolutionarily.
theAP
fruit patterning
fly Drosophila for
example,
other
sourcesInof
molecules?
D–V patterning in these embryos.97 Taken together,
short gastrulation (sog) is a homolog of the organizerthese observations perhaps suggest that D–V patternspecific BMP inhibitor chordin. Sog was identified
ing by the BMP pathway is an ancient mechanism
Spemann organizer sequentially
gives rise to axial mesoderm
underneath the future neural tissue
-Dorsal lip cells involute and form the axial mesoderm that underlie
the presumptive neural plate.
•prechordal mesoderm: rostral, originated from early dorsal lip
•notochordal mesoderm (notochord): caudal, originated from late
dorsal lip
-Like the dorsal lip itself, axial mesoderm can also induce neural
tissue in amphibians, which led to an assumption that neural induction
occurs largely via vertical signaling between the mesoderm and the
overlying ectoderm.
Sanes, 2011
-Do rostral and caudal axial mesoderm cells have different activity of
inducing rostral vs caudal neural tissue?
Temporal specificity of induction
Young (early) dorsal lip
induces a secondary head.
Older (late) dorsal lip induces
a secondary trunk.
Regional
specificity
of
induction
gional specificity arises from the organizer
nterior-posterior
mesoderm
portions
that emerged
from
transplantation
of different
rostral-caudal
parts
of the archienteron roof into early gastrula
ves 4 different outcomes:
induced tissue
ancers
aratus
balancers and oral apparatus
head
head structures (forebrain and
midbrain)
idbrain)
hindbrain
spinal cord
spinal
Otto Mangold (1933)
This led Mangold to propose that there are distinct organizers that induce different regions
of the neural tissue separately (“head-trunk-tail organizer model”)
Activation-transformation model
ectoderm
irkhäuser Verlag, CH-4010 Basel/Switzerland
implants
323
Grafts of early ectodermal tissues were transplanted into different
parts of the future neural tissue.
The proximal part included neural tissue, whereas the distal part
included non-neural tissue
Within the induced neural tissue, the more distal part was always
rostral whereas the more proximal part was always caudal
The level of the graft in the host always determined the
regional character of the most caudal neural tissue in the graft.
Niewkoop (1952)
Niewkoop and others proposed that the neural tissue is patterned by a gradient of a
transformer that travels within the plane of the neural plate and induces different neural
fate in a dose-dependent manner such that forebrain, midbrain, hindbrain and spinal cord
ure 2. Nieuwkoop implant system and basis for his two-signal
del of neural
induction
patterning. (A ) Heterotypic
grafts of this transformer (activation-transformation model).
form
atandincreasing
levels
e made by placing folds of neural-competent ectoderm perpenularly into late gastrula or early neurula hosts. (B ) A summary
Activation-transformation model
Stern et al. 2006
What is the molecular basis for this classic model?
-We now know that “activation” occurs via inhibition of BMP signaling.
-Dissociated animal cap cells will generate neural cells with anterior identity (in the
absence of exogenous BMP). Is this the default positional identity? If so, what
molecules are responsible for “transformation”? Where are they expressed?
-Are there endogenous inhibitors of transformer activities that counteract such
activities? If so, where are they expressed?
-Molecules responsible for caudalizing activity include Wnts, fibroblast growth factors
(FGFs) and retinoic acid (RA).
iments. It w
induces a fi
a
the tail. Bo
axial organ
wt
the tail dev
embryogen
trula stage
ment was th
and trunk i
trunk organ
hdl
for this mec
induces tail
the other ha
trunk and
the FGF pa
b
structures
Dkk1–/–
Moreover, t
in X. laevis
extensively
their conclu
stage as the
plate and a
showed tha
required for
As the c
distinct tai
c
Dkk1
wnt8
the TAILBUD S
the ventral
Krm
Lrp6
Fz
Membrane
ectopic tail
Cytoplasm
host embry
axin
Dsh
the location
were unexp
β-catenin
Posterior
not become
tcf3
the fish equ
Furthermor
Anterior neural genes
(for example, Six3, Hesx1)
affect ‘ectop
organizers
β-catenin signalling
Figure 3 | Genetics of the role of Wnt/β
induced tai
Over-expression
ofmouse
Cerberus
in
the head organizer. Zebrafish and
mutants, as
chord and a
well as antisense/antibody-treated Xenopus laevis embryos,
generates
an
ectopic
head
organizer in
helped to reveal the requirement for active inhibition of Wnt/
the dorsal m
β-catenin signalling in the head organizer. a | In zebrafish
headless (hdl), the transcriptional repressor tcf3 is mutated94
and the ven
(see scheme in c). The mutant embryos (lower embryo) show
acteristic of
ECTODERM, MESODERM
loss of forebrain and upper jaw. b | In Dkk1 knockout mice, the
nent. Indeed
anterior part of the head fails to develop111. c | Schematized
at the end o
AND ENDODERMWnt/β-catenin signalling pathway. Components in red are the
However, o
products of genes for which loss-of-function studies have
TAILBUD STAGE
that they di
provided direct evidence for a role in the head organizer:
The embryonic stage when
ventral mar
(REFS
92,111)
,
Krm
(REF.
101)
,
wnt8
(REFS
80,81)
,
Lrp6
Dkk1
neurulation is completed and
(REF.
115)
,
axin
(REFS
95,96)
,
β-catenin
(REF.
119)
,
tcf3
(REF.
94)
,
after ablatio
tail formation begins, visible by
Six3 (REF. 112). For clarity, some components of the pathway
an emerging tail primordium.
The Thi
have been omitted. Part a reproduced with permission from
and Nodal
REF. 94 © (2000) Macmillan Magazines Ltd. Part b reproduced
BLASTULA
activity. All
with permission from REF. 111 © (2001) Cell Press. Part c
An early-stage embryo that is
tail develo
composed of a hollow ball of
modified with permission from REF. 101 © (2002) The
(TABLE 1). Th
cells.
Company of Biologists Ltd.
Tissues underlying the anterior
brain express Wnt inhibitors
Dkk1 ko mouse
prechordal
mesoderm
(PME)
anterior
endoderm
(ae)
Dickkopf
Frzb-1
Chordin
Noggin
Cerberus
Dickkopf
Frzb-1
notochord
(cm)
M267, March 2003
Eddy De Robertis
Follistatin
Chordin
Noggin
Anterior
WS
c process that
ation of the
he body plan.
GENETIC
BMP inhibitor
Wnt inhibitor
a
PME
an
dc
ae
cm
— anterior–posterior (AP), dorsal–ventral (DV) and
left–right (LR) — in all germ layers (
). The most prominent feature of the AP
axis is the pattern of the CNS, forebrain, midbrain, hind
brain and spinal cord. The DV mesodermal axis is pat
terned to form axial (notochord), paraxial (somite)
intermediate and lateral plate mesoderm. The first evi
dence for organizer subdivision came from Spemann
Wnt signaling pathway
-evolutionarily conserved secreted proteins
-19 Wnt genes in human genome
456 Developmental mechanisms, patterning and evolution
-multiple signaling pathways depending on the cellular context and receptors
->15 Wnt receptors or co-receptors (including Frizzled (Fz) and LRP5/6 for β-catenindependent pathway)
Figure 1
Unstimulated
Stimulated
Wnt
Wnt
Wn
t
Wnt
Extracellular space
Dkk1
Lrp5/6
Lrp5/6
Fz
Gpr177
Dvl
Cytoplasm
CK1
β-catenin
Fz
U
U
U
U
β
β
β
GSK3
Axin
β
GSK3
APC
β-catenin
degraded
APC
Gpr177
CK1
SCF
complex
β-TrCP
Axin P P
Wn
t
Dvl
Gpr177
Wn
t
β-catenin
β-catenin
available for
nuclear import
Wn
t
Golgi
β-catenin
Po
rc
β-catenin
Ctnnbp1
n
W
nt
β-catenin
ER
β-catenin
Nucleus
Repressing TF
Activating TF
TLE
Tcf7l1
P
Tcf7l1
β-catenin
TARGET
Lef1/Tcf7/Tcf7l2
TARGET
TARGET
Targets,
e.g. Wnt3
Gpr177
s
nduces
e in
: at the
injected
A and
h
Animal
age and
mented
red
and
by in
jugates
entative
of the
ected or
0.1 (+)
NA were
2 and
n
h the
cap
d
ected
t8 were
En2 by
used as
ypically
of the
mpletely
Fig. 2B,
ction of
uble in
at the
blastomeres (2.5 nl/blastomere),
immunostained halves were selected from posterior to
cultured until tailbud stage and
anterior positions, according to the gastrula fate map.
scored for the induction of
Corresponding windows are shown in B for one
secondary body axes.
representative embryo. Note prominent nuclear staining in
Comparable results were
obtained in four independent
windows 1 and 2, and absence of nuclear staining in 7 and 8.
experiments. (B) RT-PCR
The AP axis is indicated (P→A) and the approximate
analysis of expression of the
position of the Gbx2 expression domain is given in relation to
indicated marker genes at (a)
early gastrula stage (stage 10+)
the windows. Data in C represent mean normalized densities
and (b) neural plate stage (stage
of the nuclear regions of all cells within a window.
15) in whole embryos (we) and
animal caps cut from embryos
injected at the eight-cell stage
XWnt3A and XWnt8 are both able to posteriorize neural fates
gradient proposed by Nieuwkoop and colleagues. In the late
into the four animal blastomeres
with no (co), 0.25 ng/blastomere Xwnt3A or 2.5
ng/blastomere
X
β
cat*
mRNAs.
H4,
histone4
for
normalization;
–RT,
negative
control
without
in these conjugates at long range, suggesting that a variety of
gastrula, the gradient appears to specify rather crude AP levels,
reverse transcriptase. (C) Pigmented donors uninjected (co) or injected with 0.25 ng/blastomere Xwnt3A or 2.5 ng/ blastomere Xβcat* mRNA
type
I
Wnts
signalling
via
β-catenin
(Wodarz
and
Nusse,
1998)
were grafted as depicted in Fig. 5A into the presumptive host anterior neural plate. Embryos were analysed at neural plate stage (stage 15) for because Gbx2 and HoxD1 are co-repressed at high XDkk1
expression of Bf1, En2 and Krox20. Frontal views
shown
in b′′,c′′).
Between 21 and 63 embryos were analysed for every typedoses. This crude pattern is refined at neurula stage when two
are are
able
to (dorsofrontal
elicit these
effects.
of transplant in six independent experiments. Note that Xβcat*-expressing grafts do not induce changes in host marker gene expression unlike
In vivo, we show that the presumptive neural plate of the
Wnt thresholds for Krox20 expression in rhombomeres 3 and
Xwnt3A (arrows), even at 5 ng/blastomere (not shown).
Graded Wnt/β-catenin signaling acts
as transforming activity
late gastrula embryo
is under
control of
a Wnt signalling
can be-Wnt
detected.
A similar refinement
of an initially crude
-In vivo and in vitro Wnt over-expression
-Late
Xenopus
gastrula
shows
an5response
antagonists
are gives
expressed
gradient
between
future
forebrain
and
spinal
cord.
This
to
a
morphogen,
which
rise to distinct
increasing dose (Itoh and Sokol, 1997). Furthermore, we show
(Baker et al., 1999; Harland, 2000; Streit et al., 2000; Wilson
in Xenopus,byzebrafish,
chick
and
mouse
gradient
in at
thelater
anterior
partbeen
of the
gastrula
gradient
is detected
(1)
the Wilson
patternetof
with
which
thresholds
stages, has
described
for Activin
titrations with Wnt
antagonists
that different
doses
of Wnt AP
et al.,by
2000;
al., Wnt/β-catenin
2001).
Yet,marker
similar togene
mammalian
signalling are required for AP neural
patterning
during
stem
cells,
dissociated
animal
cap
cells
may
provide
a
powerful
expression reacts after
injection of Wnt antagonists and XWnt8
(Green et al., 1994; Wilson and Melton, 1994) and may depend
results in caudalizatoin
of the neural
signaling
gastrulation.
system to reconstitute neural differentiation in vitro by
(Fig. 3),
(2) and
by theapplying
graded
activation
a Wnt-responsive
on secondary cell-cell communication.
Soluble XWnt8 protein specifies forebrain,
midbrain
recombinant
Wntof
protein
in combination with other
tissue.
reporter
(Fig.
7),
and
(3)
by
the
graded
nuclear
accumulation
hindbrain fates with increasing concentrations in animal cap
growth factors.
cells neuralized by dissociation (Fig. of
1). β-catenin
These inductions
Thresholds
to different
dosesgradient,
of Wnt/β-catenin
signalling are
(Fig. 8). The
orientation
of the
with peak
occur at 2-20 nM concentrations, which levels
is consistent
with
the
also
suggested
by
animal
cap
conjugate
experiments
in the posterior
and lowest levels in the anterior, is(Fig. 2),
binding constant (~9 nM) of Wnt8 for Frizzled receptors
where an organized AP pattern is induced as a function of the
consistent
with theXwnt3A
predicted
properties
a transformer
(Bhanot et al., 1996; Hsieh et al., 1999b).
To our knowledge,
mRNA dose
and of theof
distance
from the Wnt source.
this is the first demonstration of dose-dependent Wnt signalling
effects with any Wnt protein. It should be noted, however, that
neural induction of animal cap cells by Fig.
dissociation
may not of various Wnts and Wnt antagonists establishes a
9. Expression
be comparable with neural induction in the
embryo,
which
may in the gastrulating Xenopus embryo.
Wnt activity gradient
involve instructive FGF and early Wnt/β-catenin signalling
(A) Expression of Xanf1, Xwnt8 (a) in whole-mount and Xwnt3A (b)
in sagittal section at late gastrula stage by in situ hybridization. Black
Fig. 7. An endogenous AP gradient of Wnt/β-catenin
signalling
in
arrowheads,
dorsal
blastopore lip; red arrowheads, leading edge.
the late Xenopus gastrula. (A) Eight-cell stage embryos were injected
Note expression
into the four animal blastomeres with 25 pg/blastomere
TOP-Flash of Xwnt8 in proximity to presumptive posterior
neuroectoderm
or FOP-Flash (firefly luciferase) and 5 pg/blastomere
pRL (Renilla(compare with fate map in Fig. 8A). Xwnt3A is
luciferase) plasmids, cultured until early neurula
stage (stage
and
expressed
in13)
chordamesoderm
and in a ring surrounding the
cut into four AP slices. Slices from three embryos per measurement
(not visible). (B) Expression domains in the late Xenopus
were pooled, extracted and a double luciferaseblastopore
assay was performed.
gastrula
of Wnts (a) and Wnt antagonists (b). Only ectodermal
Firefly luciferase activity was normalized to Renilla
luciferase
activity. Relative TOP/FOP-Flash reporter activation
is shownare
on the
expressions
shown in a. The circumblastoporal expression of
right. Note increasing AP activation for TOP- but not for FOP-Flash.
Xwnt3A
is
not
depicted
for simplicity. Note expression of Wnt
(B) Eight-cell stage embryos were injected into the four animal
antagonists
in
the
anterior
of the gastrulating embryo. (C) Simplified
blastomeres with 25 pg/blastomere TOP-Flash and 25 pg/blastomere
pRL plasmids, cultured until early neurula stage
(stagefor
13) AP
and four
model
patterning of neuroectoderm by a Wnt activity gradient
AP slices of neuroectoderm were explanted from
each embryo.
in Xenopus.
Wnts (red) and Wnt antagonists (green) are expressed in
Explants were extracted and a double luciferase assay was
theformesendoderm
(ME, ochre) underlying the neuroectoderm (NE,
performed. Mean reporter activations are shown
three
blue).90The
expression of Wnts and Wnt antagonist in the
independent experiments (every column represents
ectodermal
explants).
neuroectoderm is not shown for simplicity (see B). Their combined
activities result in the formation of a Wnt signalling gradient (dark
orange) that patterns the AP neuraxis. The AP axis is indicated
(A→P). Formation of the posterior nervous system also requires
other factors (FGFs, RA) which are not shown here. (D) The AP and
DV axes in the gastrulating Xenopus embryo are patterned by
gradients of Wnts and BMPs, respectively (a). The Drosophila wing
imaginal disc is patterned by gradients of Wingless (Wg) and
Transcription
Decapentaplegic
(Dpp), which factors
are secretedare
alongdifferentially
the DV and AP
compartment
boundaries,
respectively
(b).
expressed in rostral and caudal neural
Kiecker and Niehrs (2001)
tissue.
Early signals come from outside of
the nervous system and establish
gross AP patterns
Caudalizing factors (Wnts, RA, FGFs) are generally
Conclusions
produced by paraxial mesoderm (somites), not
Growth-factor antagonists that are secreted by the
axial mesoderm
(descendants
Spemann–Mangold
organizer
are at the heart of
of athe organizer),
three-dimensional
system
of positional
and they
do coordinate
not induce
neural
tissue on their own.
Retinoids and Spinal Cord Development
729
information that functions during axial patterning in
gin, and follistatin, the whole neural plate does the
not vertebrate gastrula. The Nodal signalling gradient(s)
form, only forebrain tissue as reflected in the exexpression
of RA-synthesizing
is crucial to set this process in motion by inducing
both
pression of Otx2 and often En2 as well (Lamb and
Harland, 1995; Hemmati-Brivanlou et al., 1994; Cox
Wnt and BMP growth factors as well as their enzyme,
antago- Radlh2 in chick somites
and Hemmati-Brivanlou, 1995). The transformation
nists at different doses (FIG. 5). A modernized doublestep is required next to generate the remaining domains
from forebrain tissue and there are three molecules
gradient model can therefore be proposed78, in which
involved in this process, Wnts, RA, and FGFs, which
orthogonal BMP and Wnt gradients pattern the DV and
are produced by the newly generated mesoderm after
gastrulation. Each of these factors can induce posteAP axis (FIG. 6). In the classical models that were put forrior neural gene expression from this activated, anteward for amphibian embryos, the AP-graded factor was
rior neural tissue. Thus, FGFs induce posterior neural
Figure 2 (A) Expression of Raldh2 in the stage 4/5 chick
gene expression (Krox20 and Hoxb8) in a concentraproposed
to be both mesoderm-inducing as well as
embryo revealed by in situ hybridization showing exprestion-dependent fashion (Lamb and Harland, 1995;
2
sion in the mesoderm
posterior
to the node (n),
eitherindistinguishside
. The two
processes
were
posteriorizing
Kengaku and Okamoto, 1995; Cox and Hemmati-Briof the primitive streak. (B) Transverse section through a
vanlou, 1995; Pownall et al., 1996) and are required
able because
mesoderm
by
stage 6 chick
embryo stained induction
with an antibodyis
to accompanied
RALDH2
Wnt throughout the BMP
neural plate for posterior tissue specishowing
in the mesoderm
below the Today,
open neuralmesoderm
induction
ofexpression
posteriorizing
factors.
fication. Wnts induce posterior genes and suppress
plate where RA signaling occurs (arrows). (C) Expression
Figure 6 | Double-gradientanterior
model
of embryonic
axis
Different
caudalizing
factors
induction
and
posteriorization
can
uncoupled,
genes
leading to posteriorization
of the neuof Raldh2
in the
stage 10 chick embryo revealed
bybe
in situ
hybridization
showing
expression
in
the
somites
and
not
in
ral
tissue
(Kiecker
and
Niehrs,
2001).
The
specific
formation. The model shows how perpendicular activity
with
Nodals
inducing
mesoderm,
and
Wnts,
FGF and
the
neural
tube
(central
clear
area).
(D)
Transverse
section
are
responsible
for
patterning
parts of the nervous system that Wnts (most likely
gradients of Wnts and bone morphogenetic proteins (BMPs)
through
10 chick
stained with an agents.
antibody Gastruacida stage
acting
as embryo
posteriorizing
Wnt8c and Wnt11) are responsible for specifying retinoic
are
to
RALDH2
showing
expression
in
the
somites
adjacent
regulate head-to-tail and dorsal–ventral
patterning.
The
colour
at different
AP
levels
ingradient
theto into two
the caudal forebrain, midbrain, and anterior hindbrain
lation
elaborates
the
DV–AV
Nodal
the
neural tube where
RA
signaling occurs
(arrows).
et al., 2002).
RA induces
posterior genes
scales of the arrows indicate(Nordstrom
the signalling
gradients;
arrows
orthogonal
gradients
of BMP and Wnt. BMP antagonists
nervous
system
genes) and
down-regulates
anterior
indicate the spreading of the(Krox20,
signals.Hox
Patterning
begins
at
genes (Otx2) in noggin-induced ectoderm (Papalo(but
not
9-cis-RA)
are
found throughout
the cordparticularly
with
are
expressed
in
all
organizer
derivatives,
in
gastrula stages, but for clarity,
it
is
depicted
in
an
early
pulu and Kintner, 1996) and in the chick neural plate
peak levels (four to five times higher) in the brachial
the chordamesoderm,
which
convergent
amphibian neurula. The formation
head,
trunk
and tail with the well(Muhr etofal.,
1999),
in accordance
and lumbar regions (Ulven
et al.,undergoes
2001). These reestablished
the anterior induction of
requires increasing Wnt activity.
Note observations
that Nodal of
signals
gions correspond to the ‘‘hot spots’’ of RA synthetic
AND ENDODERM). The most prominent feature of
Mechanisms for early regionalization
are similar in different vertebrate
b
species
dc
ae
an
Xenopus
PME
sh
cm
a
cm
PME
an
c
ep
dc ps
hn
an
ae
hb
b
d
ae
PME cm
Mouse
an
sh
AVE
an
n
ps
PME
PME
cm
an
ps
n
c
ep
hn
ps
an
cm
axis is the pattern of the CNS, forebrain, midbrain
brain and spinal cord. The DV mesodermal axis
terned to form axial (notochord), paraxial (so
intermediate and lateral plate mesoderm. The fir
dence for organizer subdivision came from Spe
himself, who found that transplantation of the ea
TRULA lip (PRESUMPTIVE PRECHORDAL MESENDODERM,
into the BLASTOCOEL of host embryos resulted in th
mation of secondary heads. By contrast, late gastru
Cylinder-shaped
mouse embryo
becomes
(—
PRESUMPTIVE
CHORDAMESODERM
) induced
secondary
tr
anterior–posterior
(AP),
dorsal–ventral
(DV
asymmetric by the formation of anterior
Tissues
that—
correspond
the
Spemann–Ma
left–right
(LR)
in all germ
layers
(ECTODERM, MES
visceral
endoderm
(AVE).
Liketo
the
amphibian
organizer
have).been
identified
in the
chick
andof
mo
anterior
endoderm,
AVE
produces
Cerberus.
AND ENDODERM
The
most
prominent
feature
t
HENSEN’S
NODE
and in
as theforebrain,
shield. Asmidbrain
in the fro
axis is the
pattern
of fish
the CNS,
Formation
oftrunk
AVE and
triggers
the formation
tinct
head,
tailThe
organizers
haveofbeenaxis
reco
brain
and
spinal
cord.
DV
mesodermal
primitive streak (ps) 9–18
on the opposite (caudal)
(FIG.
.
in
other
vertebrates
terned
form axial
(notochord),
paraxial
side.
Theto
primitive
streak
is 1)
equivalent
to the (so
intermediate
and lateral
mesoderm.
The fir
amphibian
blastopore,
andplate
produces
Fgf, Wnt
and
Nodal.
Regionally
specificsubdivision
induction came from Spe
dence
for organizer
The
Spemann–Mangold
organizer is the
himself,
who found that transplantation
ofregion
the ear
Node (or Hansen’s node) is formed at the
gastrulation
originate.
TRULA lip (PRESUMPTIVE
PRECHORDAL
,
anterior
end ofmovements
primitive streak.
NodeMESENDODERM
isThe first org
cells
endofuphost
anteriorly
whereas
BLASTOCOEL
embryos
resulted
inlas
th
into to
themigrate
equivalent
to the amphibian
organizer,
and the
produces
chordin.
of contrast,
theend
node
(cm
will
localize
to Derivatives
theheads.
posterior
oflate
the
em
mation
of
secondary
By
gastru
and
PME) also
produce
BMPis)inhibitors
like
Therefore,
the
organizer
not
a homogenous
(PRESUMPTIVE
CHORDAMESODERM
induced
secondary
tr
chordin.
butTissues
a dynamic
while
cellsSpemann–Ma
migrate durin
thatstructure;
correspond
to the
trulation,
they
acquire
different
fates,
inducing
p
organizer
been
identified
the
chick
and mo
AVE
cannothave
induce
neural
tissueinby
itself,
but
16,19
.As
Prospectiv
ties
and
gene-expression
inhibits
caudalization
neural
tissue
by
HENSEN’S
NODE
and in of
fishthe
asprofiles
the shield.
in the fro
blocking
Wnt,
BMP
and
Nodal
pathways.
tinctare
head,
trunk
and
tail
organizers
have
been
reco
cells
among
the
first
to gastrulate
and
they
ar
9–18
AP patterning of Drosophila
embryos as a prototype
The fly consists of a head (with mouth,
eyes, antennae), three thoracic segments
(T1-3) and 8-9 abdominal segments (A1-9)
The segmentation starts to develop in early
embryos
AP patterning of Drosophila
embryos as a prototype
-Genetic screens pioneered by NüssleinVolhard and Wiechaus in the 1980s
identified a hierarchy of genes that establish
anterior-posterior polarity of Drosophila
embryos and divide the embryo into a
specific number of segments with different
identities.
-Basic ideas of the identified gene
regulatory cascade apply to many other
aspects of animal development, including
the regionalization of the vertebrate nervous
system.
-Many genes identified in this screen have
vertebrate homologs that are important in
the patterning of the neural tissue.
Polarization starts in unfertilized
oocytes
bicoid and nanos mRNAs are near the
anterior and posterior end of the oocyte,
respectively (egg-polarity genes)
Bicoid protein diffuses and forms a
concentration gradient, regulating the
graded expression of Hunchback
Hunchback, Krüppel and Giant are products
of the gap genes, which mark out coarse
subdivisions of the embryo
Pair-rule genes are required for
alternative body segments
Expression of even-skipped (eve) and fushi
tarazu (ftz) are under the combinatorial
regulation of gap genes
eve expression
Segment polarity genes organize the
AP pattern of individual segment
Segment polarity genes stabilize boundary
between segments
Genes encoding two secreted proteins,
Wingless and Hedgehog, are segment
polarity genes. They promote each other’s
expression as well as a transcription factor
Engrailed
Homeotic selector genes are required
for the identity of each segment
extra pair of
wings
Ultrabithorax
legs sprout from the
head instead of
antennae
Antennapedia
Mutations that transform parts of the body
into structures appropriate for other
positions are called homeotic mutations
Homeotic selector genes code for
DNA-binding proteins
These proteins contain 60 amino acids of a conserved DNA-binding domain called the
homeodomain
These genes are located in two clusters (Antennapedia complex and Bithorax complex) on
chromosome 3
The order of genes on the chromosome corresponds almost entirely to the order in which
they are expressed along the AP axis of the body (co-lineality)
A-P axis in vertebrates is also
controlled by Hox genes
In the mouse, there are four complexes, HoxA,
HoxB, HoxC and HoxD complexes, each on
different chromosomes
Each of the four complexes is the equivalent of
the Drosophila set
Members of each complex are expressed in a
head-to-tail series along the AP axis, just as in
Drosophila (the pattern is most clearly seen in
the neural tube, from the hindbrain to the spinal
cord, but is visible in other tissues such as the
mesoderm)
Regulation and functions of the Hox genes will
be discussed later
Hox genes ≠ Homeobox genes
Homeobox: 180 nucleotide DNA sequence
(encoding 60 amino acid of the conserved
DNA-binding domain called the homeodomain
Homeobox genes: genes containing a
homeobox
Hox genes: genes on the Hox cluster on
Drosophila chromosome or the Hox A-D
clusters in the vertebrates (some vertebrates
have fewer than four clusters). They only
comprise a small portion of homeobox genes.
In the vertebrate brain, Hox genes are not
expressed rostral to the hindbrain. Many
homeobox genes that are not Hox genes are
expressed in the midbrain and forebrain.
“Homeotic”: functional term that describes the
homeotic transformation (not the same as
homeobox)