Neurogenesis and DV Patterning

interkinetic nuclear migration

a cell cycle-dependent oscillation of nuclei between apical and basal positions in pseudostratified epithelia, notably in the developing vertebrate nervous system

undergo neurogenic division

How do stem cells exit the cell cycle?

• Proliferative = symmetric and vertical, expands neural tube

• Neurogenic = asymmetric and horizontal

Neurogenic vs proliferative cell division

the ventricular zone (VZ)

Where do cells divide?

the mantle

New born neurons exit the VZ and migrate to?

1. Ventricular zone

2. Intermediate zone (grey matter/mantle = cell bodies)

3. Marginal zone (white matter = axons)

Three zone spinal cord

neurogenesis

a pool of stem cells must be preserved for later born neurons and glia

proneural genes

expressed and promote differentiation into neurons and inhibit glia fate

neural progenitors

immature, dividing multipotent cells that differentiate into specialized cells like neurons and glia

class B bHLH (basic helix loop helix) proteins that dimerize with class A bHLH proteins

Proneural protein structure

Neural stem cells and neurons

Present in neurula embryo neural plates

neurogenin and neuroD

Types of proneural proteins

1. Delta (ligand) binds to notch (receptor)

2. Cleave of NICD (Notch intracellular domain)

3. Into membrane/nucleus

4. Activates Hes TFs

5. Represses proneural proteins

Notch pathway

lateral inhibition

induces neuron formation in salt and pepper pattern

• delta positive cells = neurons

• notch cells = stem cells/glia, since activated by adjacent neurons with delta ligands

How does lateral inhibition work?

• gain of function = overexpression of glia

• loss of function = overexpression of neurons

Notch experiment cell fates

1. Alar plate (dorsal)

2. Basal plate (ventral)

3. root plate

4. floor plate

root and floor plate act as signaling centers (non-neuronal)

Neural tube plates

ependymal cells

ciliated cells lining neural cavity

bipolar neurons in the DRG

sensory neurons

• Roof plate and floor plate = non-neuronal

• roof plate specified by BMP4 and Wnt from epidermis (dorsal side) —> roof plate makes BMP4 and Wnt after specification

• floor plate specified by Shh from notochord (ventral side) —> floor plate makes Shh after specification

• Wnt/BMP and Shh act as morphogens to create a gradient and pattern cells depending on concentration of signals received

• RA helps with further differentiation, secreted by somites

DV patterning

• Patched usually inhibits Smoothened (both transmembrane proteins)

• Shh prevents inhibition of Smoothened (moves away laterally) —> activates Gli TF —> turns on gene expression

Shh pathway

• Shh: Gli = not phosphorylated and gene expression activator

• No shh: Gli = phosphorylated and gene expression repressor

Gli duality

• gain-of-function: transplanted notochord is SUFFICIENT to induce ectopic FP and MNs

• loss-of-function (NECESSARY/required): notochord removed —> no MNs or floorplate

How did they determine that the notochord was secreting the ventralizing signal?

Shh expression is expressed in the right place and time to play in role

How did they determine that Shh is the signal emitted from the notochord

• loss-of-function: block Shh with antibody —> no MNs

• intermediate tissue + notochord in dish —> MN formation (in-vitro)

Shh induces MN formation

How did they show that Shh was responsible for generation of ventral cell types?

• several genes encoding TFs expressed in restricted domains, so…

• intermediate NT region in dish —> add defined amounts of Shh —> immunofluorescence for which TFs expressed

• different TFs expressed at different time points —> Shh = temporal morphogen

How did they demonstrate that Shh functioned as a morphogen?

bHLH proteins

required for differentiation of different populations of spinal cord neurons and glia

combinations of Olig2 and Neuorg2

leads to differentiation of motor neurons

1. Neurulation- TGF-beta, BMP

2. Caudalization- RA

3. Ventralization- Shh

4. Maturation- NTF (neurotrophic factors)

Making motor neurons