Download Polarity and Segmentation

Polarity and Segmentation
Chapter Two
Polarization
• Entire body plan is polarized
– One end is different than the other
• Head vs. Tail
– Anterior vs. Posterior
• Front vs. Back
– Ventral vs. Dorsal
• Majority of neural tube = spinal cord
• Anterior end = brain
• Vertebrate body is
symmetrical but
polarized (tail vs.
head)
• How does
polarization of the
animal body
originate during
development?
• In vertebrates, the
rostral portion of
neural tube gives
origin to brain
structures
• Caudal (tail) areas
of neural tube give
origin to spinal
cord
CNS Subdivided
CNS = central nervous system
• Spinal cord
– Derived from neural tube directly
• Brain
– Specialized anterior section of neural tube
• Brain has three primary divisions:
– Prosencephalon – forebrain
– Mesencephalon – midbrain
– Rhombencephalon – hindbrain
3 primary brain vesicles are
further divided:
Prosencephalon
1. Telencephalon:
Cerebral hemispheres
2. Diencephalon:
Thalamus, hypothalamus,
and optic vesicles
Mesencephalon remains
Rhombencephalon
1. Metencephalon
Cerebellum
2. Myelencephalon
Medulla
•
In Drosophila, development of
the anterior portion of the
nervous system also undergo a
three-chamber pattern:
1) Protocerebrum
2) Deutocerebrum
3) Tritocerebrum
•
The main difference here is that
the ventral nerve cord is
generated by delamination of
epithelial cells that are fused
together into interconnected
ganglia
•
So how do regional differences
between rostral and caudal
structures originate in
vertebrates and insects?
Formation of the Anterior-Posterior Axis
In Drosophila
•
Anterior
Posterior
In Drosophila, the A-P
axis is set by two
molecules:
1) bicoid
2) nanos
•
Anterior
Posterior
•
•
These two molecules
create an opposing
gradient
Opposing gradient
separates Anterior
from Posterior
Triggers other genes to
be on or off
A/P axis in Drosophila
Levels of development:
1. Cytoplasm gradients of nanos and bicoid
1. Inherited from mother’s oocyte
2.
3.
4.
5.
Gap genes
“Pair rule” genes
“Segment polarity” genes
Homeotic genes
1. Trigger regional gene expression differences
A/P axis in Drosophila
Formation of the Anterior-Posterior Axis
In Drosophila
Anterior
Posterior
Normal development of
anterior region in nanos -/-
Lack of anterior region
in bicoid mutant
Development of
anterior structures
following injection of bicoid
protein in posterior region
Exact reverse for Nanos and posterior development
Once the A-P layout is defined, what factors
determine the differentiation of each body
segment?
Once A/P axis is laid out
Now each segment
must express unique
genes in order to be
different
In Drosophila,
differentiation of each
segment requires the
expression of Hox
homeobox genes
Homeobox genes are
all transcription
factors
Once the A-P layout is defined, what factors
determine the differentiation of each body
segment?
Homeobox genes are arranged in a linear array on the
chromosome
Homeobox genes at the 3’ end are expressed in more
anterior locations
Homeobox genes control regional identity of body
segment
Once the A-P layout is defined, what factors
determine the differentiation of each body
segment?
Homeobox genes are
conserved among animal
species
Vertebrates:
Have more hox genes
Complex interactions
Overlap of same function
Still exist in order 3’ to 5’
Homeobox genes
• Hox genes
• Always transcription factors
• Bind DNA directly:
– Through the homeobox domain
• Activate the genes that directly
cause specific regional identity
• Deactivate other genes
Distaless is NOT a homeobox gene
•
•
•
•
•
Insects have three pairs of legs
One pair on each thoracic segment
No legs on the abdominal segments
Distaless gene forms the legs
Distaless expression is suppressed in
abdominal segments of insect
• By BT-X – a hox gene
– BT-X binds the DNA and repressed distaless
Example of Hox mutations:
wild type
Antennapedia
Mutation in the antennapedia
complex result in the formation
of a leg where an antenna was
supposed to exist
Do Hox genes control vertebrate
development as well?
• XlHbox1 in
Xenopus
Regional expression pattern
of XlHbox1
Do Hox genes control vertebrate
development as well?
• Injection of an antibody against the
XlHbox1 protein result in the
enlargement of the hindbrain
Knock Out Mice
Specific genes
deletion studies
Study the role of
homeobox genes
in segmental
differentiation
Hox knockouts have allowed the study of
regional differences in the developing
mice hindbrain
Segmentation of the hindbrain
result in the formation of
rhombomeres – similar to
Drosophila’s segments
Rhombomere formation in the
mouse is encoded by different
combination of Hox homeobox
genes.
Hox mutations affect the
development of specific
rhombomeres
Hox knockouts have allowed the study of
regional differences in the developing
mice hindbrain
Deletion of
Hoxa1 gene
results in fusion
of rhombomeres
(R) 5 & 6 and
reduction of R4
Deletion of
Hoxb1 gene
results the loss of
motoneurons R5
Double mutants
of Hoxb1 &
Hoxa1 genes
show a combined
effect
Segmental Specification is Encoded by
Combination of Factors
Segmental arrangement of
the chick hindbrain
rhombomeres (r1-r7) result in
a steroptypic pattern of
motoneuron localization in
hindbrain
Early transcription factors,
Eph family receptors, and
homeobox genes establish
the segmental specification of
the hindbrain rhombomeres
Important to Note
• Vertebrate hindbrain segmentation
• Occurs by exact same mechanism as
insect body segmentation
• Express different hox genes
– Produce different regional identities
• In insects get body segments
• In vertebrates get different motor neurons
– In exactly to the correct brain region
Removing Hox genes:
Entire hindbrain looks like R1 segment
What controls Hox genes?
•
•
•
•
Step backwards one
If Hox genes regulate all other genes
What regulates Hox genes?
In flies:
– Cytoplasmic gradients
– Control Gap genes
– Control Pair rule genes Æcontrol Hox genes
• Same in vertebrates?
What signal molecules pattern
the Hox expression?
• Retinoic acid (RA) has been
show to regulate Hox gene
expression.
• RA crosses cell membrane and
bind to cytoplasmic receptors.
• The RA-receptor complex can
translocate to the nucleus and
regulate gene expression after
binding to RA response
element (RARE)
There is a gradient of RA expression in the
developing embryo
• RA levels are significantly
higher in posterior regions of
Xenopus embryos
• RA normally activates
posterior identity
• Suppresses anterior identity
• Exposure of the developing
embryo to RA results in
malformations of anterior
structures (head structures fail
to develop)
Retinoid Acid Controls Formation of A-P
Axis and Hox expression
in vitro and in vivo
• Low RA – results in Hox genes
normally expressed in the
anterior portion
• High RA – results in Hox genes
normally expressed in the
posterior portion of the embryo
• Target deletion of RA receptors
results in head structure
formation
• RA normally regulates in more
posterior regions
Heads vs. Tails?
• Spemann and others proved there were
both head and tail organizers
• This meant that if you transplanted a small
piece of tissue from head to anywhere it
would still form into a head
• Same with piece of tail tissue
• What are in these regions?
– Transcription factors that induce other genes
Heads vs. Tails?
• Nieuwkoop transplanted small piece of
head tissue to different places along axis
• All pieces transformed into head tissue
• However, if transplanted into caudal
regions – actually formed two tissue types:
– Anterior and Posterior both developed
• New theory:
– Activator = 1st signal
– Transformer = 2nd signal
Activator-Transformer Hypothesis
Activator = gene that turns ectodermal cells
into neural tissue
• Anterior is default state of neural tissue
Transformer = gene that turns neural tissue
into posterior types
• Posterior requires two signals:
– Neural positive
– Posterior positive
Activator Genes
•
•
•
•
Genes that induce neural tissue
Noggin, Chordin, Follistatin
All produce anterior structures
Therefore anterior must be default for
neural tissue
• Remove these activator signals and get
NO neural tissue
– Rather than posterior-like tissue
Transformer Genes
• Retinoic acid
– Posteriorize embryos
– Regulates hindbrain hox genes
• Wnt and beta-catenin
– When wnt is inhibited a second head can form
– Adding wnt posteriorizes neural tissue
• FGF
– Induce posterior gene expression
The Activator-Transformer Hypothesis
• Blocking of BMP signal enables
formation of neural tissue. In
this case noggin, chordin,
follistatin play the role of neural
inducers that allow formation of
anterior structures
• Formation of a retinoid acidgradient enables polarization of
the neural tissue. In this case
RA is the transformer that
promotes Hox gene expression
and formation of caudal
structures
Complexity
• Although anterior type seems to be
“default”
– Suppressing BMPs is necessary to form
anterior tissue
• Suppressing BMPs is NOT sufficient to
form functional anterior tissue
• If all you do is suppress BMPs you will
form anterior structures
– Will not be fully functional or normal
Anterior Tissues
Anterior tissues require two signals:
1. Inhibition of BMPs
1. Forms neural tissue
2. Inhibition of wnt pathway
1. Anteriorizes the neural tissue completely
•
•
Inhibition of BMPs is necessary, but not
sufficient to form anterior structures
Inhibition of wnt pathway is necessary
but not sufficient
Role of wnt in Xenopus embryos
G = Suppressing BMPs and wnt pathways
I = Suppressing BMPs alone
– Small head and cyclopia
Wnt inhibitors
• All expressed within organizer:
• Cerberus
– Injection of Cerberus will cause ectopic head
formation
• frzB
– Injection forms larger than normal heads
• dkk1
– Ectopic head formation
Wnt
Remove wnt
= extra head
Inhibit wnt
= extra head
Remove wnt inhibitor
= messed up head
Cooperation
• BMP inhibitors work together with wnt
inhibitors to form head and brain
Wild Type
Double mutant:
dkk and Noggin
FGF
• Third transformer is FGF
• Both FGF and wnt signals suppress
expression of enzyme cyp26
• cyp26 is an enzyme that breaks down
retinoic acid
• Without cyp26 – RA builds up:
– Posteriorization of tissues
• Exact mechanism/interaction of FGF and
wnt is unknown
Any Questions?
Read Chapter Two