Quiz 4 short answer Flashcards
The segmentation pattern that develops in the drosophila embryo through the expression of various sets of genes is locked in by the expression of Hox genes.
a) How many Hox genes are present in Drosophila? Which chromosome are they found on? (1 mark)
b) Which axis do they pattern? (1 mark)
c) Regulation of the Hox gene expression occurs through what main mechanism? (1 mark) What are the 2 complexes that facilitate this mechanism? (1 mark) briefly explain their function (1 mark)
a) 8 genes, on chromosome 3
b) Anterior posterior (AP)
c) -Hox genes are controlled by WNT and HH
-function thru bithorax complex (abdominal and thoracic segments) and antennapedia complex (thoracic and head)
-regulation via epigenetic mechanisms- chromatin remodelling
Trithorax: keeps chromatin open once HOX genes are on (most posterior gene would be dominant) - blocks polycomb action
Polycomb: keeps chromatin closed in regions where HOX genes are not yet expressed
Gamma secretase cleaves notch after delta binding in the notch signaling pathway. What would be the effect of a gamma-secretase inhibitor on differentiation of neural progenitors to neurons (2 Marks)?
The cleavage of notch after delta binding causes notch activation.
-since gamma secretase cleaves notch, it activates it - if inhibited notch activation will not occur.
-active notch causes progenitor cells to stay progenitors and inactive notch causes differentiation into a more specialized cell type
- this means that a gamma secretase inhibitor would stop notch signaling, causing progenitor cells to differentiate - mother cell or progenitor cell population will not be maintained
A Competition between secreted signaling proteins patterns the vertebrate embryo along the animal vegetal axis and dorsal ventral axis. What signaling pathway is involved in this initial patterning along these axes? Explain how the activators and repressors of this pathway lead to the formation of endoderm and ectoderm and how the ectoderm is further subdivided into neural vs dermal tissue. (5 marks)
TGFb (Nodal and Lefty) signaling patterns the animal vegetal (AV) axis
-Nodal: TGFR activator, acts locally (because it diffuses slowly)
-Lefty: SMAD inhibitor that acts DISTAL (animal pole) to site of secretion (because it diffuses rapidly)
-*Both nodal and lefty secreted at vegetal pole
-More nodal is active at the vegetal pole (since it slowly diffuses), activates TGFB signalling, causes formation of endoderm on vegetal side
-More lefty is active at the animal pole (since it quickly diffuses), inhibits TBFB signaling and causes formation of ectoderm at animal pole
Sub pattern:
-a gradient of inhibition of TBFb family members sub-patterns DV axis while patterning AV
-Noggin and Chordin: SMAD inhibitors/TGF antagonists ; released at dorsal side
-generate diffusion gradient of inhibition concentration on dorsal side
-cause dorsal = neural tissue
-ventral (high TGFB activity, no noggin or chordin) = dermis
Sub-pattern:
-BMPs: activator
-Noggin and chordin block BMP on dorsal side (but not ventral) in a gradient manner
-High BMP results in epidermal tissue (ventral), low BMP results in neural (dorsal)
And then when you put these gradients together you get ectoderm and
endoderm but within the ectoderm you have the division of dermal and neural
tissues
Explain how a shift in the Notch Delta signaling pathway leads to both the differentiation of the progenitor cells into functional cells AND how it maintains the progenitor or mother cell population in the formation of the Drosophila bristle. (4 marks)
(4 marks)
Active notch = stay progenitor
Inactive notch = differentiate
-Notch signaling turns off when time to differentiate
Two cells have notch and delta - neither active
-Activation of notch by delta on neighboring cells through lateral contact signals inhibition of specialization - allows them to stay mother cells
-shift in signaling - do not know how this happens exactly (transient bias creates slight asymmetry)
-ends with one cell with active notch, one with delta = notch sell stays as mother cell, delta differentiates into one of the more specialized cells (socket, shaft, sheath, neuron)
-sequential rounds of notch-mediated lateral inhibition occur until most cells are functional progeny and only a few mother cells remain to regenerate tissue
Explain how WNT signaling leads to the formation of the neural fold. (3 marks) What would happen if there was a mutation in the NLS of beta catenin? (2 marks)
Wnt 2 signaling pathways:
Canonical - beta-catenin transcriptional co-activation = generation of neural progenitors and specialized subtypes
Non-canonical - disheveled can activate rho/rock and alter cytoskeleton = neural tube closure and cytoskeletal dynamics
Rho signaling is activated through the Wnt pathway - Rho-GTPases promote invagination of epi cells - RhoA mediates actin/myosin contraction of adhesion actin belt = invagination
B-catenin NLS directs their transport to the nucleus - mutation would not allow this to heppen ; unable to activate transcription of target genes needed for wnt signalling - would interfere with formation of neural fold/neural tube closure - can result in severe birth defects
Explain how morphogenic signaling leads to the patterning and development of the various types of neurons within the spinal cord along the dorsal ventral axis. (Be sure to mention the location of the signaling and the development of sensory vs motor neurons) (5 marks)
Shh dominates at ventral side of neural tube ; aids in formation of floor plate (notochord), mesencephalon, motor neurons, dopaminergic neurons
BMP and WNT dominate at dorsal side of neural tube ; aids in formation of roof plate, neural tube (becomes CNS) and the development and migration of neural crest cells out of neural tube that becomes the PNS also paraxial mesoderm development (becomes muscle)
BMP and WNT released from roof plate and diffuse down, Shh released from floor plate and notochord and will diffuse up - gives unique gradient of signals along neural tube allows cells in different areas to be exposed to different levels or morphogens
High WNT/BMP / low Shh at dorsal side = sensory neurons
Low WNT/BMP / high Shh at ventral side = motor neurons
Explain what radial glial cells are and how they support the formation of a developing neocortex. (4 marks)
-Stem cells that are able to give rise to neurons and glial cells
-Newly born neurons or glial cells that are produced from radial glial cells are made in ventral proliferative zone and can migrate up the radial glial cell towards the growth cone
-Can deposit where needed and become what they need to become based on location they are in - (how the brain is actually developed)
-Will extend further (via growth cone) and keep populating the area with different cells along the way
-Will build each layer of tissue as it extends. Next round of neurons can migrate past the first to sequentially grow the brain - results in formation of layers in newly developing neocortex
Why do commissural axons in the developing neural tube turn when the cross the midline? (2 marks)
Once commissural neurons cross midline netrin signalling is out competed by slit proteins
-Netrin diffuses far enough from floor plate to reach neuron and act as a chemoattractant - one neuron gets closer it comes in contact with split
-Slit = chemorepellant, also released from floor plate but cant diffuse very far - only acts locally
-Causes migrating axon to start moving to the brain
You think you may have identified a small molecule that promotes differentiation of intestinal stem cells to absorptive cells. You decide to perform a BrdU experiment to label dividing stem cells in the intestinal crypt of mice exposed to your small molecule and see what they become. You pulse the mice with BrdU but are unsure how long to wait after the pulse before staining the tissue for markers of absorptive cells. Your advisor suggests that you wait 3-5 days. Why do think they recommended this time frame? (2 marks)
Remember the timeline of proliferation within the crypt
-bottom of crypt after 24 hours
-still be in proliferation zone after 48 hours
-cells become epithelium and migrate out of crypt (reach villi) - wnt in crypt so cant differentiate, now it can)
-after 5 days = apoptosis
In your lab you study the epithelial mesenchymal transition and how it relates to the metastasis of lung cancer to the brain. You are testing a potential drug which your student found that appears to inhibit this transition, but your student does not know the mechanism through which it works. Through various experiments you find altered WNT signaling when the drug is administered to the cells. Explain how altered WNT signaling might inhibit the epithelial mesenchymal transition in these cells and result in blocking metastasis. (4 marks)
TGFB and WNT must both be present to support EMT
-TGFB: SMAD pathway - drives expression of EMT-TFs (eg. snail)
-WNT: frizzled/ b-catenin activate EMT-TFs through GSK3b inhibition -
*GSK3b inhibition BLOCKS snail degradation
Snail: active in nucleus; drives repression of E-cadherins and expression of N-cadherin and vimentin - facilitates switch to mesenchymal
Without WNT signalling snail is degraded - switch to mesenchymal cant occur
-snail expression does not matter if they are being degraded!!!!
Cancer: EMT allows cells to start migrating out of epi layer - if cells have accumulated cancerous mutations (many in epi cell layer of gut have which is why they undergo apoptosis) - cancer cells can migrate out and become metastatic
Blocking of WNT stops EMT - prevents this from occurring
Explain how the stem cell population is maintained in the crypt of the gut epithelium while still supporting a fast turnover rate of the differentiated functional cells. Be sure to mention WNT and Notch signaling. If the cells that line the gut have a high turnover rate how can cancer develop? (6 marks)
-Wnt and notch control the gut, wnt maintains notch expression
-Paneth cells secrete WNT - crypt is under WNT influence - active WNT = notch express
-Stem cells express notch, paneth cells express delta to inhibit stem cells
-Stem cells use asymmetric division to create more stem cells and progenitor cells which will proliferate faster
-Progenitor cells continue to proliferate and migrate up and out of the crypt which is when they differentiate into functional cells
-Notch expressing cells will become absorptive cells and delta expressing cells will become secretory cells
-Cancer happens when mutation in APC complex: allows for overactivation of WNT pathway by preventing degradation of beta catenin - inhibits cells ability to migrate out of the crypt but continue to proliferate uncontrolled
-Create accumulation of cell protrusions called adenomas
-Not cancerous but harsh environment makes it likely for them to accumulate mutations (because they do not undergo mutation in the crypt since they cannot migrate out - can lead to cancer)
