Lecture 10 - Patterning from the Organizer

Question 1

What does the organizer comprise?

Answer 1

mesendodermal progenitor cells that during gastrulation give rise to 3 distinct embryonic tissues:
- anterior endoderm
- prochordal mesoderm
- notochord

Question 2

What is formed during gastrulation?

Answer 2

The A/P & D/V pattern of the embryo - under the direction of the Spemann-Mangold Organizer

Question 3

How was the Spemann-Mangold Organizer discovered?

Answer 3

Part of embryo put in the another location. This induces a secondary invagination. These induced secondary structures and the development of a double sided organism (e.g. 2 heads - 2 ventral parts touching each other.

Question 4

What else can induce the development of secondary structures?

Answer 4

The microinjection of Goosecoid mRNA which is expressed specifically in the organizer can have similar effects

Question 5

What can a transplantation of tissue from the dorsal blastopore lip lead to?

Answer 5

A small piece of tissue removed from the dorsal blastopore lip & transplanted to the central side of a host embryo can induce a secondary embryonic axis in the host embryo, comprising mesodermal & neural tissues & organs.

Question 6

How do the inductive properties of the Organizer change during gastrulation?

Answer 6

Early Organizer - complete second axis including head & trunk

Late Organizer - partial second axis comprising trunk tissue only

Question 7

What occurs as gastrulation proceeds?

Answer 7

The Early Organizer tissue migrates into the embryo & under the ectoderm that forms the blastocoel roof, creating the A/P axis of the embryo as it moves

The Organizer tissue in the early gastrula migrates anteriorly across the blastopore roof.

Question 8

Explain how organizer tissues from early, mid- and late gastrulae have different fates in the embryo.

Answer 8

Organizer at early, mid- and late gastrula-stages have DIFFERENT INDUCING POTENTIAL because it expresses DIFFERENT SIGNALLING MOLECULES over time

Early organizer - prechordal plate mesoderm & pharyngeal endoderm

Late organizer - notochordal mesoderm

Question 9

How are the primary inducing properties of the Organizer achieved?

Answer 9

by secreting molecules that INHIBIT Wnt & BMP signalling pathway activities

Question 10

Why do early, mid- & late gastrula-stage donors have different inducing potential?

Answer 10

because the Organizer tissues express different signalling molecules - e.g. BMP levels

Question 11

What does Early Organizer tissue express?

Answer 11

Wnt antagonists & BMP antagonists –> Head & Brain

Question 12

What does Mid- and Late Organizer tissue express?

Answer 12

only BMP antagonists –> Trunk & Spinal Cord

Question 13

What are examples of BMP antagonists?

Answer 13

• Chordin
• Noggin
• Follistatin
• Cereberus
• IGF

Question 14

What are examples of Wnt antagonists?

Answer 14

• Dickkopf
• Cereberus
• FrzB

Question 15

Where are BMP Antagonists expressed?

Answer 15

BMP Antagonists - Noggin, Chordin & Cereberus are expressed in the Spemann-Mangold Organzier & they act on ectoderm to suppress epidermal fate & induce neural fate in the neural plate.

Question 16

What does Cereberus do?

Answer 16

Cereberus from the organizer inhibits both Wnt8 and BMP4 activity, which allows the anteriorization & dorsalization of neural plate tissue, leading to head induction

Question 17

Describe the double gradient model of embryonic axis formation

Answer 17

Spemann-Mangold Organizer generates:

1. A D/V gradient of BMP signalling activity throughout the embryo - lowest in neural plate & dorsal axial mesoderm (where BMP antagonist activity is highest)
2. An A/P gradient of Wnt signalling activity in the developing neural plate - where Wnt antagonist activity is highest.

Question 18

What is induction an example of?

Answer 18

A change in fate mediated by an extrinsic/ non-cell-autonomous signal

Question 19

What promotes brain formation in the anterior neural plate?

Answer 19

Dual inhibition of Wnt & BMP signalling promotes spinal cord formation

Question 20

What promotes spinal cord formation?

Answer 20

BMP inhibition without Wnt inhibition

Question 21

How are the more detailed elements of A/P patterning in the brain & spinal cord created?

Answer 21

Asymmetric division.
Cell A divides asymmetrically. One daughter cell adopts the same fate as mother cell (A). Second daughter adopts a different fate X, because it may lack certain components present in the mother cell & then cell-autonomously differentiates to the alternate fate C.

Question 22

What occurs in the Early Organiser?

Answer 22

Anterior TFs are active in neural plate, leading to the formation of prechordal mesoderm/pharyngeal endoderm

Question 23

What occurs in the Mid-Organiser?

Answer 23

Notochord represses Anterior TFs in overlying neural plate and promotes posterior TFs.

Question 24

What occurs in the Late Organiser?

Answer 24

Notochord elongates & further posteriorizes the overlying neural plate which becomes the neural tube

Question 25

Describe the Activation-Transformation Model of Neural Plate/Tube Anterior-Posterior Patterning

Answer 25

Activation: 1. Organizer in the early gastrula secretes Wnt Antagonists & BMP Antagonists 2. Neuroectoderm exposed to these Organizer signals adopts forebrain fate. Transformation: 3. From mid-gastrula stages onwards, secretion of Wnt Antagonists by Organizer is attenuated (reduced). 4. Posterior neural plate is exposed to multiple gradients of posteriorizing factors - e.g. Wnts, Retinoic Acid & FGFs. 5. Forebrain fate is suppressed and over time, increasingly posteriorized neural fates are induced by posteriozing factors 6. Neural plate elongates and transitions to the neural tube

Question 26

What does activation-transformation involve?

Answer 26

Activation-Transformation involves simultaneous integration of multiple signalling pathways: e.g. Retinoic acid & FGFs Retinoic Acid: a diffusible ligand for nuclear Retinoic Acid Receptor transcription factors Fibroblast Growth Factors (FGFs): signal to ETS transcription factors in the nucleus via a cascade of Serine-Threonine kinases

Question 27

How is A/P regionalization of the neural plate achieved?

Answer 27

By 2 opposing gradients of antagonistic signals - "anteriorizing" & "posteriorizing" factors Bmp & Wnt antagonists are maintained anteriorly Fgfs, Wnts, RA are produced posteriorly

Question 28

How do transcription factors expressed in the neural plate interpret the anterior & posterior signal levels?

Answer 28

By activating different HOX genes in different positions, leading to regional specification.

Question 29

What do specific combinations of Hox transcription factor expression in the neural tube do?

Answer 29

Specify the distinct domains of the spinal cord.

Question 30

Describe how Retinoic acid influences Hox gene expression

Answer 30

Retinoic acid gradient induces specific patterns of HOX gene transcription at different positions along the A/P axis Low RA = top of body (neck - cervical) High RA = bottom of spine (tail)

Question 31

Describe the steps in neural tube formation

Answer 31

1. Flat neural plate 2. Formation of neural furrow 3. Elevation of neural folds 4. Neural folds bend inwards 5. Neural folds fuse 6. Neural crest cells migrate

Question 32

How is the vertebrate neural tube polarized along its D/V axis?

Answer 32

By 2 opposing signalling systems: 1. TGF-B ligands (mainly BMPs) secreted by the dorsal Roof plate - induce dorsal neuronal fates 2. Sonic Hedgehog (Shh) secreted by the ventral Floor Plate (& notochord) - induces ventral neuronal fates. - BMP signalling from the epidermis specifies the roof plate - BMPs from the roof plate pattern the dorsal half of the neural tube - Shh signalling the notochord specifies floor plate - Shh from the floor plate patterns the ventral half of the neural tube

Question 33

What occurs in the dorsal neural tube?

Answer 33

Different members of the BMP family are expressed in different domains, and induce different dorsal neuronal fates

Question 34

What occurs in the ventral neural tube?

Answer 34

Different concentrations of Shh induce different ventral neuronal fates A gradient of Shh protein is established in the ventral neural neural tube which determines neural progenitor fate by regulating expression of Homeodomain Transcription Factors such as Pax7, Pax6 & Nkx6.1 in a Shh-concentration-dependent manner

Question 35

Explain how neurons develop?

Answer 35

Bmp & Shh induce distinct domains of transcription factor expression in neural progenitor domains along the D/V axis of the early neural tubes. 3 days later These transcription factors define distinct neural identities that are realized when progenitors terminally differentiate into neurons a few days later.

Question 36

Summarize patterning from the organizer

Answer 36

1. The A/P axis of the embryo is formed during gastrulation, under the direction of the Organizer - a dynamic tissue comprising axial mesendodermal progenitor cells that contribute to endoderm, prechordal mesoderm & notochord. 2. In amphibian embryos, the Organzier first appears as the dorsal blastopore lip of the early gastrula. Transplantation of the dorsal blastopore lip from an early gastrula-stage embryos into the ventral side of the second embryo induces a secondary embryonic axis in the host that induces a second brain & spinal cord. 3. The inducing & patterning functions of the Organizer are achieved by secreting molecules that inhibit Wnt and/or BMP signalling pathway activities. 4. Induction of head and brain structures requires dual inhibition of both Wnt & BMP signalling, whereas induction of spinal cord & trunk requires inhibition of BMP signalling only. 5. The Organizer generates an A/P gradient of Wnt activity, and a D/V gradient of BMP activity - a double gradient that simultaneously drives A/P & D/V axis formation. 6. Patterning of the A/P axis of the neural tube requires Activation by Wnt/BMP antagonists to generate anterior-dorsal structures (e.g. forebrain) and Transformation by Wnts, Retinoic Acid & FGFs to progressively posteriorize neural tissues that is induced as gastrulation proceeds. 7. D/V patterning of the vertebrate neural tube is achieved by BMP, secreted by cells in the dorsal roof plate, and Shh, secreted by ventral notochord and floor plate, which establish a transcription factor code defining neuronal subtype identity. 8. General principle: axial patterning is achieved by gradients or stepwise differences in competing or antagonistic intercellular signals that create a range of different cell identities in between their localised sources.