L9 - Neural tube Induction and Pattering Flashcards
What aspects of NS formation is common to both vertebrates and invertebrates
Neurogenic region found next to the skin This migrates down then goes inside
What feature of NS formation is found only n invertebrates
Delamination forming the neruoblast cells
What feature of NS formation is found only in vertebrates
Inc cell cell adhestion contactEither side the neural plate is bound by ectoderm, this then fuses above the neural plate
What would happen if all of the cells took their default state
All would become skin
Define differentiation
Process by which cells are able to become different and acquire specialised properties
What is cell differentiation governed by
Changes in gene expression which influences the repertoire of proteins expressed in a cell
Over time what happens to specialisation and pluripotency
Spec inc P.P dec
Is BMP found is vertebrates of invertebrates Give an example of an organism it is found in
Vertebrtes Xenopus
Is Dpp found in vertebrates or invertebrates Give an example of an organism it is found in
Invertebrates Drosophila
BMP7 homologue in drosophila
Screw
What does BMP stand for
Bone morphogenic protein
What does dpp stand for
Decapentaplegic
BMP1 homologue is drosophila
Tolloid
BMP4 homologue is drosophila
Dpp
Chordin homologue is drosophila
Short gastrulation (SOG)
Screw homologue in xenopus
BMP7
Dpp homologue in xenopus
BMP4
Tolloid homologue in xenopus
BMP1
SOG homologue in xenopus
Chordin
Normal function of Chordin/SOG
Inhibition of BMP/dpp signalling
What is the normal function of BMP4 and dpp
The activation of non-neural homeobox genes
Describe the BMP signalling when BMP has bound to the receptor
Phosphorylation of the receptor Phosphorylation of R-SMADCombines with SMAD4 Enters the nucelus via the nuclear pore Trancriptional complex Dephosphorylation and it leaves the nucleus
What are the 3(4) components of the SMAD transcriptional complex
PHOSPHORYLATED R-SMADSMAD4 Co factor
What two antagonists inhibit BMP signalling
Noggin and chordin
What is the function of noggin
Causes the inactivation of SMAD4
What are BMP and chordin examples of
Genes that have been conserved throughout evolution
What are the two mechanisms by which chordin and sog act as antagonists of the BMP/dpp signalling pathwat
Act as a sponge moping up dpp/bmp (binding to and inactivating)Competitively bind at receptors preventing binding of dpp/bmp
What does Dpp and SOG broadly dictate
The dorsal and ventral axis
What genes are expressed on the dorsal side of the embryo (insect)
Dpp, tld, tsg and SV-2
Describe the activity of the dpp pathway on the dorsal side of the embryo (insect)
No inhibition of the dpp pathway
What cell fate is taken by cells on the dorsal side of the embryo (insect)
Epidermal
What genes are expressed on the ventral side of the insect embryo
Sog and screw
What is the action of SOG
Inhibition of dpp binding to its receptor
What fate is taken by cells on the ventral side of the cell
Neural cell fate - neurogenic region is formed
What side is the NS found in insects
Ventral
What side is the NS found in vertebrates
Dorsal
Why is there a difference between the sides which the NS is found in vertebrates and invertebrates
Possibly due to a rotation of the head
Genes expressed at the dorsal end of a vertebrate - what fate do these cells then take
Chordin and Admp cells take a nerual cell fate
Genes expressed at the ventral end of an invertebrate What fate do these cells take
Sog, Screw
Molecular pathway ECTODERM IN PRESSENCE OF BMP SIGNALLING
High SMAD1 and low SMAD1Msx1, GATA1, Vent (epidermalising transcription factors) LEF1Epidermal differentiation
Molecular pathway ECTODERM WITHOUT BMP SIGNALLING
Low SMAD1 and high SMAD7 Xlpou2, SOXd/B (neuralising transcription factors)NeurogeninNeuroDNeural differentiation
What process results in the formation of the neural tub
Neuralation Neural plate rolls up to form the neural tube
What occurs during spinobifia
Failure of the neural tube to close properly
What can reduce the chances of spinobifida occuring
Folic acid
How do we know there are 3 germ layers found in the embryo
Repitore of proteins expressed - use of an antibody stain mRNA being expressed - look at using an in-situ hybridisation
What sets up the polarity of the early embryo
Established early on due to gravity and interaction of the placenta (in mammals) different cytoplasmic determinants have sunk to the one part of the egg. Leads to cells of this region becomming different and having a specific set of factors being activated
Which part of the embryo is fated to become the furture CNS
Cells immediatly adjacent to the organiser
What signal does the organiser secrete … what does this lead to
GSC is expressed this acts to upregulate Chordin , Noggin and Follistatin These secreted molecules act to inhibit BMP signalling
What the result of BMP signalling inhibiton
Neural plate induction
What are some antagonists of the BMP signalling pathways
Chordin NogginFollistatin
Outline the stages of mesoderm induction and patterning
Low level Nodal gives the ventral mesoderm High level Nodal gives the organiser Signals from the organiser acts to inhibit BMPs to dorsalise and pattern adjacent mesoderm Antagonism of BMPs –> Gives a neral identity
What receptors do BMPs bind to
TGFb-R
How to BMPs tend to act
Locally and diffuse to neigbouring cells
What fate do cells take if BMP signalling is active
Cells take an ectodermal cell fate
How do chordin noggin and follistatin act as inhibitors of the BMP signalling pathway
Antagonists compete for BMP binding - no longer able to activate receptors and cause activation of the BMP pathway
What is different in the development of chicks and humans
flattened into three sheets not a hollow sphere
What do cells in the organiser differentiate into
Axail mesendoderm and anteiror endoderm
What makes up the axial mesendoderm
Anteior endoderm, precordal mesoderm, notochord
What then happens to the axial mesendoderm
Involute, intercalates and undergoes convergent extension so it now lies as a rod underneath the neural plate
Describe invagination
Epithelial sheet bends inwards
Describe ingression
Where individual cells leave an epithelial sheet and become freely migrating mesenchyme
Describe involution
An epithelial sheet bends inward forming an underlying layer
Describe epiboly
Sheet of cells which streches by thinning
Describe intercalation
Row of cells which moves between one another creating an array of cells that is longer and thinner
Describe convergent extension
When rows of cells intercalate but this intercalation is highly directional
What is the anterior endoderm and prechordal mesoderm in close contact with
The pahrynx and ventral forebrain
Describe the process of the Organiser Graft exptWho performed it and when
Speeman and Mangold - 1920Organiser grafted from newt into a second newt - found that a twinned secondary axis was formed
What are the conclusions from the Organiser Graft expt
2nd neural tube was host derived shows that the neural tissue was induced from the ectoderm in response to signalling from the organsier The axial mesoderm (precordal mesoderm and notochord) and anterior endoderm was donor derived and therefore differentiates from the organiser
How were BMP antagonists discovered?
Extracted all the mRNA from the organiser cells and reverse transcribe to cDNA Each then tested for a gene/protein which mimiced the action of the organisers ability to induce a secondary neural plate
What occurs at the Speeman organiser
Gsc upregulated Leads to upregulation of chordin, noggin and follistatin Leads to inhibition of BMP signalling Leads to induction of a nerual cell fate
What are features of neural inducers in the organiser
Molecules must be expressed in the organiser Must be secreted and act of adjacent cellsOverexpression in an ectopic site should lead to induction of a secondary axis Inhibition should prevent the formation of any neural tissue - and any formation of the axis
Describe how the neural tube is formed
Cells of the nerual plate have apical-basal polarity On the apical side - band of F-actin, when this contracts, the cells are forced to change their shape (constriction at the apical edge of the cell) MANY MOLECULES ARE DIFFERENTIALLY DISTRIBUTED AT THE APICAL AND BASAL EDGES - AND MAY INTERACT WITH F ACTION CAUSING THE CONTRACTION AND SUBSEQUENT SHAPE CHANGE
Describe the role of follate receptors in spinofiida
Expressed on the apical domains of cells of the neural plate - when folate (derived from folic acid) When folic acid defficient not enough contraction of the F-actin leads to the neural tube not closing properly
What three systems must be developed
ENS PNS CNS
What regions make up the forbrain
Lateral ventricleTelencephalon Neural retina Lens3rd ventricle
What regions make up the midbrain
Mesencephalon and the cerebral aqueduct
What regions make up the hind brain
Metencephalon Myclencephalon 4th ventricle
What layers lie under the posterior end of the neural plate
Notochord
What layers lie under the anterior end of the neural plate
Anterior endoderm Prechordal mesoderm
What can be said about the signals from the anterior endoderm and prechordal mesoderm compared to the notochord, what does this cause
Slightly different signals Signals from the anterior endoderm and prechordal mesoderm cause the induction of the anterior neural identity
What experiemental evidence can be obtained if developement is stopped at the instnant where the prechordal mesoderm has involuted
Look at the molecular markers can see that the neural plate expresses makers which are later confined to the forebrain i.e. the early neural plate is anterior in character
Describe the activation-transformation model and its relevance to the determination of the posterior and anterior identities
Signals from the notochord cause cells at the back end of the neural plate to proliferateAt the same time other signals from the notochord changes these cells from an anterior to posterior identity Anterior TFs are turned OFFPosterior TFs turned ON
What is the activation-transformation model the basis for
The formation of the anterior forebrain and the posterior hind brain and spinal chord
What signals are involved at the anterior end
BMP antagonists and WNT antagonists
What signals are involved at the posterior end
FGFs (in a gdt highest most posteriorly)WNTS (in a gdt highest most posteriorly)RA (in a gdt highest most posteriorly)
Describe how RA (what does it stand for) signalling works
Retanoic acid Small so able to diffuse into cells where it binds to a cytosolic receptor - complex RA-R able to enter the nucleus where it directly influences gene expression
Segmentation occurs as a result of
Step wise difference in the gradients of the signalling moleucles
What are the two models for segemetation by a morphogen
Alan Turing - reaction diffusion Lewis- wolpert Postional infromation (french flag)
What does HOX stand for
Homeobox containing
Describe what the HOX genes are responsible for
Selection of a certain body area fate along the AP axis e.g. abdomen and the thorax
Describe the different genes that are involved in the segmentation of the drosophila
Gap Pair ruleHomeotic genes Segment polarity genes
What are the different segments of the drosophila
T1-T3 and A1-A8
T1-T3 controlled by
UBX
A1-A4 controlled by
AbdA
A5-A8 controlled by
AbdB
What causes activation of the HOX genes
Very specific threshold concentrations of posteriorising factors
How many HOX genes in humans Why is this the case
4Safety measure incase of mutation
What is homeobox DNA
Encodes a DNA binding protein of 60 amino acids called the homeodomain
What are the different regions of hind brain and spinal cord defined by
HOX signature
What does the RA gradient induce
Different patters of HOX transcript
What occurs in a knock out of HOXa1 and HOXb1
These genes are required to specify rhombomeres 4 and 5 and nerves that are normally made in this region So double knockout for these genes results in a mouse that will develop without the abducens nerve
Where are the midbrain strucutres induced
At the interaction between the forebrain and hindbrain boundary
What are the three primary brain vesicles
Prosencephalon Mesencephalon Rhombencephalon
What does the prosencephalon give rise to
Telencephalon Diencephalon
What does the rhombencephalon give rise to
Metencephalon Mycelnecphalon
What does the mesencephalon give rise to
Midbrain
What does the telencephalon give rise to
Olfactory bulbs Hippocampus Cerebrum `
What does the diencephalon give rise to
Retina Epithalamus Thalamus Hypothalamus
What does the metencephalon give rise to
Cerebellum Pons
What does the myclencephalon give rise to
Medulla
The early ground plan for the mature NS is established by?..
Antagonistic signalling centres
What is secreted from the node/organiser … give an example of one
BMP antagonists e.g. chordin
What does the secretion of BMP antagonists cause the induction of
Induction of neural plate formation
Where there is an INTERMEDIATE level of BMP signalling what is this zone known as
Neural plate border Found between the neural plate and the non-neural ectoderm
3 crucial points of the neural plate border
Crucial for neural crest formation - and the formation of the PNS Crucial for the formation of the root plate cells and the patterning of the dorsal neural tub Crucial for complete closure of the neural tube
What does neural crest formation and neural tube patterning happen before
The expression of Shh in the notochord
STAGE 1] An early _______ established due to the ________ between BMP signalling and antagonism. This _____ ______ _______ then begins to express specific transcritpion factors such as _____
Border Balance Neural plate border Msx1
At the neural plate border BMP signalling level is
Intermediate
STAGE 2] Other signals (____ and ____) act together with _____ to turn _____________
WNT and FGFs Msx1 Turn on other transcription factors
What signals are from the preaxial mesoderms
WNTs and FGFs
What signalis from the non neural ectoderm
WNTs
Msx1 acts in a ____________________ and WNTS and FGFS act in a _______________ to cause the activation of ________ and ________
Intrinsic - cell autonomousExtrinsic - cell non-autonomous PAX3 and ZIC1
STAGE 3] ____ signals acts with the transcription factors of the ________________ to upregulate further TFs - (_____,______,_____) which are characteristic of neural crest cells
WNT Neural plate border c-myc, Snail, ID
PAX3 and ZIC1 act in a ____________WNTS act in a __________To cause the upregulation of ______ and ______
Extrinsic - cell non-autonomous Intrinsic - cell autonomous Snail and FoxD3
Snail is responsible for
Promoting cell survival Promotion of cells to undergo and epithelial mesenchymal transition
c-myc is an Upregualtion will causes
OncogeneProliferation of cells
ID will cause
Stem like behaviour of cells (multipotency)
STAGE 4a] In response to _______ and ______ genes that control _______ , _______ and ________ are activated
c-myc and SOX9Survival, multipotency and survival
Cascade of expression and effect of c-myc
c-myc –> ID –> Multipotency and proliferation
Cascade of expression and effect of SOX9
Sox9 –> Snail –/ Cell death (promotes cell survival)
Stage 4b] At the same time there is an upregulation of genes which promote ____________. This then causes the _________ of the _______ _______ cells and ____________
Epithelial –> Mesenchymal transition Delamination Neural crest Migration away
Neural crest cells are often descirbed as
The fourth germ layer
What sort of cells do the neural crest cells differentiate into
PNS Adrenal medulla Melanocytes Facial cartilage Dentin of the teeth
What are the different cell types of the neural crest cell formed by
Position of origin (due to HOX gene)Time of generation by the cells Migration pathway and the signals they encounter en route or at the target
Describe how the dorsal route ganglion is formed
NCCs usually at the posterior of the somite and force to move into the anterior of the somite by pissoffin
Describe how the sympathetic and parasympathetic NS are formed
NCCs reach are beneath the notochord and the aorta
Do all NCCs delaminate
No, some stay and form the roof plate clls
What are the roof plate cells crucial in
Through to play a part in the fusion of the two sides of the neural tube
What was previously thought to act as a morphogen
BMP from the roof plate
What is now thought to be the way in which the roof plate signals
Different BMPS secreted from the roof plate ==> each BMP induces a different dorsal cell type
What are the different progenitors on the dorsal side of the neural tube
P1 –> P6
What are the cell types which differentiate dorsally
Roof plate cells Neural crest cells Different classes of dorsal sensory relay interneurones
What is the single cell thick neural plate also referred to as
Neuroepithelium
When the neural tube forms what occurs to the medial lateral axis
Becomes the DV axis
Notochord is _______ to the neural tube
Ventral
Skin ectoderm is _______ to the neural tube
Dorsal
Neurones develop with ______________ throughout the entire DV axis
Bilateral symmetry
How can it be tested that the notochord acts as a DV axis organiser
Graft the notochord into an ectopic site
What occured when the notochord was grafted into an ectopic location
Induction of a secondary floor plate with bilateral symmetry of secondary motor neurones
What is the Shh homologue in drosophila
Hh
What does an insitu hybridisation against Shh show
That initially expressed in the notochord Induced expression of Shh in the floor plate cells
What does an antibody stain show about Shh expression
Forms a gradient - highest ventrally and lowest dorsally
What does Shh confer
Ventral neural tube identities so progenitors will ultimately give rise to cells that differentiate into ventral motor neurones
What occurs when a bead soaked in Shh is grafted into an ectopic site
Mimics the effect of the notochord and floor plate Induces the secondary floor plate and ventral motor neurones
What does Shh cause
Set of transcription factors which are a read outs of the Shh gradient
Where is the Shh gradient highest
Ventrally
What occurs as the ventral cells differentiate
The move laterally and differentiate into a neurone
Shh acts at an ___________ to confer a _______ pattern of TFs on progenitor cells
Early stage DV
Oncee the single cell neuroepithelium has been formed what is then induced
The neural tube
What does the mediallateral axis become
DV
Where does the notochord form
On the ventral surface
Whaere does the skin ectoderm form
On the dorsal surface
What is the floor plate closest to?
The notochord
The roof plate is ______ wherase the floor plate is ______
Dorsal Ventral - next to the notochord
What occurs at the same time as the neuralation
The axial mesoderm (notochord and prechordal mesoderm) forms and comes just beneath the ventral midline of the neural tube
What do neurones develop with around the midline throughout the whole DV axis
Bilateral symmetry
What is secreted by the notochord and the floor plate cells What does this act as
Shh Morphogen
How can it be tested that the notochord and floorplate secrete a morphogenWhat were the results of this experiement?
Grafting the notochord into an ectopic site into a host embryo Induces a secondary floor plate and second set of motor neurones around - these exhibit bilateral symmetry
What does an in-situ hybiridisation over time for Shh show
Initially expressed in the notochord This is then induced in the floor plate cells
What experiment could be used to show that Shh is acting as a morphogen
Antibody stain
What does an antibody stain for Shh show
Forms a concentration gradient (highest ventrally - lowest dorsally)
What does Shh cause
Expression of a certain set of TFs in the cells that it act on
What dose the TFs induced by Shh confer
Ventral neural tube identities so progenitors will give rise to cells that differentiate into ventral neurons
What is the experimental evidence that it is SHH as a signalling molecule
Bead soaked in purified Shh protein is able to mimic the effect of the notochord and floor plate - induces a secondary floor plate and motor neurones
In real life the set of TFs are readouts of the
Shh gradient
Shh is highest
Ventrally
What occurs to the cells as they differentitate
As they divide one cell retained as the mother cell and one moves laterally and differentiates into a neurone
Shh acts at an __________ to confer ________ of TF on progenitor cells
Shh acts an early stage to confer a DV pattern
What is the Shh gradient
Highest at the notochord and then the floor plate
What is the BMP graient
From neural plate border cells and then the roof plate cells
BMPs are members of the
TGF beta superfamily
Progenitor domains made up of bands of proliferating cells are established by …
The opposing action of BMPS/WNTS (dorsally) and Shh (ventrally)
Which way do cells move as they differentiate
Laterally
Descibe how hedgehog signalling occurs
Hh ligand binds to PtcPtc then unable to inhibit Smo Smo then causes an increase in the ammount of Gli activator
Hh signalling creates a balance of
Gli activator and repressor
What is the Hh signalling in the inactive form
No Hh ligand bound to PtcNormal function of Ptc is to inhibit Smo Smo unable to cause increase in Gli activator Smo unable to cause increase in Gli activator ]More gli repressor than gli activators
Describe the phenotype of the Shh -/-
Multiple dysmorphology No ventralisation of neural tube Holoprosencephaly CyclopiaAbnormal limbs/digits Lack of pituitary
What is holoprosencephaly
Where there is no separation of the two parts of the brain
What does Shh govern
Governs ventralisation along the whole rosto-caudal axis
How come different neurones are formed at different levels along the A-P axis of the neural tube
The DV shh gradient intersects the AP hox gradient - forms a Cartesian grid of information - leads to different types of neurones being formed that are in the same DV position but differ on the AP axis
What type of neurones form in the forebrain
Hypothalamic neurones
What type of neurones form in the midbrain
Dopaminegric neurones
What type of neurones form in the hindbrain
Serotonergic neurones
What neurones form elsewhere along the DV axis
Motor neurones
