L6

Question 1

Invertebrate neural induction:

Answer 1

Induction of CNS dependent on competing gradients do Dorsal and Decapentaplegic (DPP/BMP), Dorsal targets high+low affinity (HA+LA) sites in genome, in lateral region only HA activated -> neuroecotderm, LA (suppress HA) in ventral -> mesoderm; Leads to competing gradients of SOG and DPP- at low concentrations Dorsal binds Sog (direct activation), at high binds Twist, upregulates Snail and SNAIL represses Sog

Question 2

Which genes to Twist, Snail, Sog and DPP activate?

Answer 2

Twist- mesoderm (ventral)
Snail- non-mesodermal (lateral+dorsal)
Sog- neuroectoderm (lateral)
DPP- dorsal ectoderm (dorsal)

Question 3

Effects of modifying gradients?

Answer 3

Knocking out a gradient or master TF leads to loss of entire layer

Tweaking a gradient leads to shift in the boundary between lawyers

Question 4

Layer boundaries?

Answer 4

Learn the fucking table

Question 5

SoxN

Answer 5

One of the earliest genes expressed throughout SOG domain; is a homologous of vertebrate Sox2

Question 6

After invertebrate neural induction:

Answer 6

SoxN dependent transcription programme in neuroectoderm cells leads to expression of a proneural gene; proneural factor confers a neural fate to a cell; not all cells in neuroectoderm progress to neuroblasts, rest revert to ventral fate and form ventral epidermis due to lateral inhibition

Question 7

Neural induction in frogs

Answer 7

Generation of main tissue layers
Animal pole (future ectoderm), vegetal pole (future endoderm); endoderm signal to ectoderm to transform into mesoderm at equatorial ring; mesoderm signal to ectoderm via Sox2+Oct4 to induce neuroectoderm (at dorsal midline, NSCs now committed); lateral ectoderm signals to lateral neuroectoderm to specify neural crest (thus is NE that subsequently acquires potent mesodermal properties); anterior neuroectoderm signals to anterior ectoderm to specify panplacodal ectoderm

Question 8

Placodal ectoderm

Answer 8

Forebrain (olfactory placode = nasal epithelium)

Hindbrain (otic placode = auditory epithelium)

Question 9

Cranial neural crest

Answer 9

Facial bones

Question 10

Cranial neural crest

Answer 10

Spinal chord (DRG of PNS, ganglionic cells of ANS)

Question 11

What are the key pathways in vertebrate neural induction?

Answer 11

BMP4/ FGF8

N.B. Early events ouch cells towards primitive germ layers, mesoderm then helps to induce neuroectoderm by blocking weak ectoderm signal (BMP) that allows FGF to drive ectoderm to neuroectoderm (SOX2-OCT4) fate

Question 12

Gradients of maternal mRNA…

(Neural induction step 1)

Frog

Answer 12

…polarise the egg

Dominant gradient early on is ventral TGFβ, quickly drives vegetal cells to endoderm, anterior BMP is slower acting (takes longer to specific ectoderm fate in animal cells)
Geminin gradient also makes Sox2 gene more accessible in animal cells

Question 13

The dorsal organiser (nodal gradient) arises…

Vertebrate neural induction step 2

Frog

Answer 13

Signalling in the vegetal plug establishes the dorsal organiser, the resultant Nodal signal now induces animal cells at equator to become mesoderm

Question 14

The Spemann organiser (BMP inhibitor) arises…

Vertebrate neural induction step 3

Frog

Answer 14

Mesoderm expresses WNT and migrates around the equator to the posterior (i.e above DO), WNT signal combined with high Nodal from DO generates the Spemann organiser

Question 15

Gastrulation-early

Frog

Answer 15

Cells on dorsal lip of blastopore invaginate and pass under ectoderm (derived from animal cap), invaginating cells are mesoderm

Question 16

Late gastrulation

Frog

Answer 16

Spemann organiser induces overlying ectoderm to form neural tissue instead of epidermis

Question 17

Neural induction in culture

Frog

Answer 17

Isolate animal cap from late blastula and culture -> intact epidermis

If same cells isolated -> neural

Thus cell-cell contact prevents ectoderm from becoming neural, and default state for ectoderm is neural

If isolated cells have BMP4 added, become epidermis again

If cells not isolated but have Noggin or Follistatin added, become neural again (blocks signalling between intact ectodermal cells)

Question 18

Where are Noggin/Follistatin/Chordin expressed?

Answer 18

Dorsal lip of blastopore (Spemann organiser), all three bind BMP4

Question 19

More recent work has shown that FGF signalling inhibits BMP signalling at Smad1 level, and that FGF upregulates Sox2

Frog

Answer 19

In the animal cap, BMP>Smad pushes cells towards ectoderm, before they fully commit Spemann organiser is established and releases various BMP inhibitors; mesoderm carries these under animal cap to block BMP which allows FGF>MAPK to be upregulated; MAPK now cross inhibits Smad intracellularly (blocking BMP transduction); BMP inhibition allows more potent FGF activation; FGF increase Sox2 to autoregulation -> cells now committed neuroectoderm (neural plate)

Question 20

Neural induction

Chick

Answer 20

Activation of near primitive streak by FGF4/8, generates most of the neural tube

Expansion driven by Hensen’s node BMP+WNT inhibition, expands the anterior neural tube (forebrain)

Question 21

Frog VS Chick

Answer 21

Chick has separated roles of BMP inhibition and FGF4/8 activation (+WNT needs to be blocked to form anterior)

[in chick, BMP not present in primitive streak region and a novel mechanism has evolved to modulate chromatin remodelling at Sox2 locus (Bert and Erni, which act via Geminin-Brahma, so evolved from chromatin remodelling mechanism in frogs]