Lecture 7 - Conservation of neural development in vertebrate

Question 1

How does neural induction show conservation in the vertebrates?

Answer 1

-secretion of BMP inhibitors from the dorsal mesoderm is involved in neural induction in all vertebrates

Question 2

What is the purpose of neural induction?

Answer 2

-specifies the region of the ectoderm that will give rise to the CNS

Question 3

What experiment showed that the organiser (dorsal mesoderm) is involved in neural induction? And what conclusions were reached from this experiment?

Answer 3

The Organiser Graft experiment

• a secondary axis forms from the ectopic grafting of the organser (spemanns)
• the nervous system of the secondary axis is host derived
• the cells of the organiser give rise to the mesoderm of the head and the notochord
• the source of neural inducing signals is the dorsal mesoderm (head mesoderm and notochord)

Question 4

What do the cells of the organiser give rise to?

Answer 4

The mesoderm of the head and notochord

Question 5

Where is the source of neural inducing signals?

Answer 5

the dorsal mesoderm (the head mesoderm and the notochord)

Question 6

What are the signalling molecules involved in neural induction?

Answer 6

Noggin

Chordin

Question 7

What is noggin?

Answer 7

• a neural inducing molecule
• expressed at the right place and the right time to be involved in neural induction
• in early gastrula it is expressed in the organiser
• in the neurula it is expressed in the notochord and connective tissue of the head (shown through in situ hybridisation)

Question 8

What is chordin?

Answer 8

• a novel neural inducing moleule
• induces neural tissue in ectoderm explants
• expressed in the right place at the right time to be involved in neural induction
• in the gastrula it is expressed at high levels in spemanns organiser
• in the neurula it is expressed in the underlysing mesoderm and the notochord

Question 9

What is the mode of action of noggin and chordin?

Answer 9

• Noggin and chordin are secreted peptides in the EC environment
• they do not bind to cell surface receptors
• in the developing embryo, noggin and chordin binds to and inhibit the function of BMP (bone morphogenicproteins)

Question 10

What is BMPs?

Answer 10

-members of the TGFβ super family of signalling moleulces

-

Question 11

What is the process of BMP signalling?

Answer 11

• TGFβs bind to the cell surface receptors (complex of type I and type II) [ligand initially binds to type II then is directed to type I]
• Both receptors are serine/threoine kinases and through binding and the formation of the receptor complex, their activty is activated
• this leads to auto and trans phosphorylation events on the serine/threoine residues within the kinase domains
• the activated receptor complec phoshporylated the SMAD group of receptor proteins
• SMAD then associateds with coSMAD (SMAD4) and this complex is translocated to the nucleus where it acts as a transcription factor

Question 12

What are the two types of ligands that activate SMAD receptor proteins?

Answer 12

Nodal-lie ligands (activate SMAD 2 and 3)
BMP-like ligands (acivate SMAD 1 and 5 - then associate with SMAD4 which activates/inhibits different sets of target genes involved in mesoderm initiation)

Question 13

What is the action of Noggin and Chordin in ectoderm explants?

Answer 13

induce neural tissue

Question 14

What experiment showed the action of Noggin and Chordin in ectoderm explants?

Answer 14

• Take an animal hemisphere explant which would normally form the ecoderm
• expose it to factors and observe how it affects development
• if cultured with simple salt solution after 3 days, it forms cilliated epidermal tissue
• if cultured with the Noggin protein after three days, forms forebrain-like tissue

Question 15

How is BMP involved in the model for neural induction?

Answer 15

In the gastrula embryo, there is a BMP gradient from the ventral mesoderm to the dorsal mesoderm.

• BMP inhibitor underlies the dorsal ectoderm, producing a region of high levels of BMP agonist leading to low levels of BMP in the neural plate which will form the CNS
• BMP in the epidermis will activate genes necessary for the formation of the skin

Question 16

What criteria must be fulfilled to assign function to a molecule?

Answer 16

Must have:

1) Relevnat biological activity (ectopic expression)
2) Present at the right time (in situ hybridisations)
3) Present at the right place (insitu hybrisations)
4) Must be necessary (does process fail when inhibited - backward/forward screen (or in amphibians use anti-sense technolgy)

Question 17

What methods are there to inhibit gene function in non-genetic model organisms? E.g. amphibians

Answer 17

Anti-sense technology
-knowedge of the gene sequence allows the design of antisense molecule which wil hybridise to the taget sequence. Target molecules complimentary to mRNA to inhibit their function
Nucleic acids
-expression of DNA or RNA antisense molecules can either destroy or inhibit translation of target mRNA
Morpholino oligo mediated gene knockdown (best)

Question 18

What is the process of Morpholino oligo mediated gene knockdown?

Answer 18

• polymers contain A, C, G, and T bases and are able to hybridise with RNA and DNA (e.g. target mRNAs)
• modified chemistry in the polymer backbone in which a morpholine ring replace ribose or deoxyribose sugar (means that enzymes are not able to target the RNA for destruction)
• highly stable and non-toxic
• morpholinos can be injected into embryos
• binding of the morpholino oligo blocks the ability of the ribosome to begin translating the target mRNA

Question 19

What results from a triple morpholino knockdown of chordin, follistatin and noggin?

Answer 19

Knocked out together and separately
-expressed in overlapping domains in the dorsal mesoderm
-shows redundancy of funtion amongst the BMP antagonists:
Single knockdowns show nop effect on neural development
Dounle knockdowns: still show significant expression and redundant function
Triple knockdown: results in a loss of neural tissue in neurula stage embryos

Question 20

What is involved in neuronal differentiation?

Answer 20

• neural induction defines a neural terratory (the neural plate/neuro-epithelium)
• not all cells in the neural plate will differentiate as neurons, only a subset of cells in the neural plate
• only individual cells in the neural plate are picked out to be glia/neurons

Question 21

What are the two types of neural cells?

Answer 21

Neurons

Glia

Question 22

Where do neurons and glia arise from?

Answer 22

Neruons and glia arise from neural projenitors in the neuro-epithelium

Question 23

What are neural progenitors?

Answer 23

stem cells which proliferate producing daughter cells - some of which will remain as neural progenitors and some of which will become commited neural precursors and ultimately differentiate into neurons or glia

Question 24

What are neural precursors?

Answer 24

Cells that have commited to differentiating into neurons or glia

Question 25

What is neurogenesis?

Answer 25

the process of generating neurons

-highly conserved developmental process

Question 26

What is involved in neurogenesis in flies?

Answer 26

• sensory bristles of the peripheral nervous system form from Sense organ precursor (SOP) cells derived from the lateral ectoderm
• neurons of the CNS form from the neuroblasts derived from the ventral neuro-ectoderm

Question 27

What is involved in neurogenesis in the Amphibian primary nervous system?

Answer 27

-primary nervous system of aquatic vertebrates forms very early in development to allow swimming as escape from predation
either side of the midline there are three regions of the neurons that differentiate:
-sensory neurons (lateral group)
-intermediate neurons (in between )
-motor neurons (medial group)

Question 28

What can be used as a marker of differentiating neurons?

Answer 28

N-tubulin