Notch Signalling in Cortical Development Flashcards
(41 cards)
How variable is the notch signalling pathway across species?
Notch signaling pathway is highly conserved in evolution
Describe the Delta-Notch signalling pathway
Interaction between the Delta cell surface ligands and their Notch receptors on neural progenitor cells in close proximity to one another regulates transcription factors necessary for the generation of differentiated neurons.
Delta binding cleaves a protein fragment of the receptor (the Notch intracellular domain, or NICD). When the NICD is transported into the nucleus, it binds to a transcriptional complex including the recombining binding protein J (RBP-J), inhibiting RBP-J repression and resulting in the transcription of (among others) the Hes family of transcription factors, including the bHLH neurogenic factors responsible for neuronal differentiation.
Describe the relationship between Delta ligands and differentiation
Delta, Notch, and bHLH proteins are expressed at similar levels in a cluster of progenitor cells and neuroblasts. A stochastic increase in Delta ligands on a particular cell leads to downregulation of Delta in neighboring cells, while in the Delta-upregulated cell, bHLH gene expression is also upregulated, and the cell becomes primed for neuronal differentiation.
How many Notch receptors do Mammals have?
Mammals possess 4 Notch receptors (Notch1,2,3,4)
What kind of receptors are notch receptors? What happens when notch ligands bind to them?
Notch receptor is a hetero-oligomer transmembrane protein. Notch ligands bind to the extracellular domain and cause proteolytic cleavage and release of the intracellular domain
What are most Notch ligands? What does this mean for function?
Most Notch ligands are transmembrane proteins, making Notch-signaling a typical cell-cell interaction pathway
Is Notch more in development or in mature brains?
Notch function is highly versatile, ranging from neural development to mammary gland regulation and cancer development
What is Notch signalling most known function
Cell fate decisions
In what part of corticogenesis is notch signalling particularly important for?
Notch signaling is involved in corticogenesis especially during neurogenesis
Describe the normal pathway for notch in stem cell maintenance by the diagram used in the slides
It is involved in corticoexpansion: Mash1 and NGN2 activate Delta1 (Dll1) which with Mindbomb1 create the DII1 transmembrane protein.
When this interacts with the Notch receptor the NICD protein is detached and travels to the nucleus. Numb, however can inhibit it at this time.
NCID then forms a complex with Rbpj and MAML (mastermind-like) to activate the Hes1/Hes5 gene. These proteins inhibit Mash1 and Ng2.
What is the relevance of this inhibitory effect?
It carries out lateral inhibition. Lateral inhibition (in neural development) is a cell-cell interaction whereby
a cell adopting a certain fate, inhibits its immediately neighbours from doing likewise
Expression of Delta is important for differentiating cells, cells which are differentiating express Delta. There is lateral inhibition, it regulates its own expression. The bHLH genes, Mash1/ngn2 are downregulated by Hes1 and Hes5 and therefore do not activate delta to incur notch signalling in surrounding cells.
What pattern does this signalling dynamic incur in RGCs
Expression of Notch effectors (Hes1/Hes5) is highly dynamic in neuronal progenitors and Notch activation levels in RG exhibit an oscillatory pattern:
Notch activates the Hes1 gene, resulting in transcription of Hes1 RNA and translation of the Hes1 protein. This protein then represses further transcription of the gene and is degraded through the upiquitin-proteosome pathway.
What does the Smidt lab also think shows this oscillatory pattern?
They also think that Foxo6 may be oscillating, however this is very hard to show in the lab.
What downstream effects does this oscillitory pattern have?
There is dynamic expression of Notch effectors in neural progenitors. Ngn2 and DII1 expression oscillates in a pattern directly opposing a pattern of Hes1.
How does this pattern differ in a stem cell to a neuron?
In neural stem cells/ progenitors this expression pattern is oscillating. The decision to remain a stem cell is this balance and when this balance is disturbed then differentiation occurs. Differentiation occurs when Ngn2 and DII1 remain high, when notch signalling occurs, and Hes1 remains low.
Relate Notch and delta levels to the aforementioned migratory patterns
Notch and Delta levels seem interconnected with cell cycle and interkinetic nucleus migration.
Notch is high at the apical site and delta is high at the basal site.
Notch is therefore high at G1, when the NPC is closer to the apical site.
Delta is then high in the S phase, when the NPC is at the basal site.
Notch becomes higher as the cell migrates to the apical site again while going through G2 and is the highest as it undergoes mitosis.
Describe a way in which notch aids differentiation
It is involved in asymmetrically localised cell fate determination. mPar3 shows a dynamic distribution in RG during cell cycle, resulting in asymmetric cell division during the neurogenic phase. mPar3 and Notch signaling are interconnected in a regulatory feedback-loop
In the paper which described this mPar3 Notch feedback loop, what was the main question?
how is asymmetric cell division regulated in radial glia?
What were the methods of the mPar3 Notch paper?
Study was performed in WT and Par3 deficient mice and it utilised IHC, neocortical slice culturing, Cre-mediated ectopic gene-expression.
They did a clonal cell pair assay in which embryos at E.13 were removed from the womb and coronal slices were obtained. At E.14 cortical tissue was plated at clonal density and cultured for 24hrs before fixation and IHC.
Similarly for neocortical slice culturing they cut a section of the dorsal cortex and underwent organotypic slice culture before doing time-lapse imaging for 24 hours. They then fixated the tissue and underwent IHC.
What is confocal microscopy?
Confocal microscopy focuses the light at what you are looking at and filters the light being reflected. This allows you to look at different layers of cells rather than just looking at the tissue.
What did the mPar3 Notch paper find?
They found that the mpar3, as well as ZO-1 (junctional protein) expression was enriched at the luminal surface of the VZ. It found they largely colocalised with each other at the lateral membrane domain in the ventricular endfeet. Through further staining they found that mPar3 and ZO-1 were expressed together at the apical ends of the end feet (towards ventrical) which B cantenin and N-cadherin (junctional proteins) were expressed independently higher up on the end feet.
What is meant by these end feet?
Cell body of radioglia is attached to the ventrical via an endfoot; the cell body is not attached to the ventricle. The nucleus is also in the endfoot; it is not always in a rounded structure
What does it mean to say that RGs are polarised?
Cortical progenitors (RGs) are highly polarized, facing the lumen of the ventricular zone, and reaching to the pial surface via radial processes
How is apical basal polarity maintained?
Apical-basal polarity is maintained by adherens junctions (cell- cell adhesion)
