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NEUR3005 > Lecture 3 - gene regulation > Flashcards

Lecture 3 - gene regulation Flashcards

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1
Q

Early embryo development stages

A
  1. Single fertilisation
  2. Cell division - blastocyst forms
  3. Gastrulation - form 3 germlines
    Ecto/Meso/Endoderm
  4. Organogenesis
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2
Q

Why do we want to compare embryos?

A

Early stages are very similar across species.
Mouse models have been useful in research

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3
Q

Early CNS formation

A
  1. Neural Plate
  2. Folds to form Neural Groove
  3. Then Neural Tube
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4
Q

What stage are we at in a 3-4 week embryo?

A

3 Vesicle stage

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5
Q

What stage are we at in a 5 week embryo?

A

5 Vesicle Stage

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6
Q

Dorsoventral patterning

A

Mainly controlled by opposing signaling gradients of WNT/BMP from the roof plate, and SHH from floor plate cells

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7
Q

Shh pathway is strongly associated with…

A

Development of the neural tube, patterning of the ventral structures and ventral forebrain, neuronal differentiation, proliferation

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8
Q

WNT-signalling controls…

A

Neural tube rostrocaudal patterning
* progresive caudalisation from fore to hindbrain

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9
Q

What does neuroectodermal expression of Dickkopf1 (Dkk1) do?

A

Antagonises and inhibits Wnt signalling in the ANTERIOR

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10
Q

What signalling component is highly expressed in the anterior?

A

Sfrp1

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11
Q

What is the role of notch signalling?

A

Maintains neural progenitors/stem cells in developing brain

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12
Q

What aspects of neurogenesis does notch signalling impact?

A

Morphology, migration, synaptic plasticity, maintenance of mature/immature neurons + radial glia , and dendrite development.

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13
Q

What does notch signalling promote and inhibit?

A

Inhibit: Neurons and Oligodendrocytes
Promotes: Astrocytes

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14
Q

What is essential for a “prepattern” of neural induction?

A

Fibroblast growth factor (FGF) signals from precursors of organiser prior to gastrulation

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15
Q

How does FGF work?

A

Activates Sox3 and early response to neural induction

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16
Q

How is the process of transcription initiated?

A

RNA polymerase binds upstream of the gene on its promoter
* 1 TF binds to one of these promoter sequences, initiating a series of interactions.

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17
Q

What are Transcription factors (TFs)?

A

Regulatory proteins that activate (inhibit is rare) transcription of DNA by binding to SPECIFIC DNA sequences

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18
Q

How are TFs categorised?

A

They have highly conserved DNA binding domains which catagorise them into families (MADS box-containing proteins, SOX proteins, and POU factors)

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19
Q

Key transcription factors involved in determining specific neural fates

A

Homeodomain proteins and basic helix-loop-helix (bHLH) TFs
* provide regional identity (A-P and V-D axes)

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20
Q

Basic-helix-loop-helix (bHLH) Transcription Factors

A

Ensure that appropriate no. of specific neuronal and glial cell types are produced
* bind E-box motifs with the consensus sequence CANNTG.

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21
Q

What neural lineages are bHLH genes expressed in?

A

Neurogenin, Neurod, Atonal and Olig families

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22
Q

What does Neurod1 do?

A

Differentiation of inner ear sensory neurons and granule cells in the cerebellum and hippocampus

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23
Q

What does Neurod2 and 6 do?

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A

Both required for formation of callosal connections in the Cerebral Cx.

24
Q

What does Bhlhe22 do?

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A

Differentiation of neurons in several CNS domains, incl. dorsal horn of the spinal cord, dorsal cochlear nucleus in the brainstem and retinal amacrine cells

25
Homeoproteins role in CNS development
Essential for *formation* of **borders**
26
What significance does OTX2 and GBX2 have in mice brain development?
There is a ***sharp border*** between these proteins. If it is altered: * More **OTX2** > *larger* **midbrain** * More **GBX2** > *larger* **hindbrain**
27
What does areal development refer to?
*Formation* of *distinct* functional **brain regions**
28
What does laminar development refer to?
*Formation* of *distinct* **layers** within each brain region, creating a layered structure known as a "**lamina**."
29
What is Pax6 and what does it do?
*TF* which promotes development of **rostro-lateral** regions
30
What is Emx2
*TF* which promotes **caudal-medial** regions
31
In what axis does Pax6 and Emx2 create an opposing gradient. How?
**Anteroposterior** axis * **EMX2** directly ***represses*** **PAX6** expression and vice versa
32
What effect did removing Emx2 expression in mice have?
Zone of **Pax6** expression is ***enlarged*** * *Increase* in primary motor (**M1**) and sensory (**S1**) areas of the *anterior* cortex.
33
What effect did removing Pax6 expression in mice have?
The primary visual (**V1**) cortical area is ***increased***.
34
What are HOX genes?
Conserved family of **homeodomain TFs** with roles in (**A-P**) patterning and the early **NS development**
35
What regulates HOX genes?
Expression of ***RA, FGF, WNT***
36
Outline where Hox1-5 and Hox4-11 are expressed
Hox**1**-Hox**5**: expressed in the ***hindbrain*** Hox**4**-Hox**11** detected in the ***spinal cord***
37
SOX2
**TF** expression ***marks*** the **CNS** from the earliest developmental stages.
38
SOX4
*Promote* neuronal ***differentiation*** both in the **adult** and **embryonic** neural **progenitors**
39
SOX5
Controls the *sequential* generation of distinct **corticofugal neuron** subtypes by ***preventing*** **premature emergence**.
40
SOX9
*Differentiation* of neural **stem** cells (NSCs) into ***glial*** cells, rather than neurons.
41
Transgenesis
Introducing **new** gene into mouse genome to ***overexpress*** specific protein
42
Gene knockout
***Deleting*** specific gene
43
Conditional knockout
Allow gene **deletion** in *specific* cell types at *specific* developmental **stage** using **Cre-loxP** system
44
Point mutations
Introduce specific ***changes*** in gene to ***mimic*** human genetic **mutations** associated with **neurological disorders**
45
Where is Twist1 expressed?
In **Neural Crest Cells** (NCCs)
46
What happens when you knockout Twist1 only?
**Not too much** morphological difference
47
Twist1 AND Chd7 knocked out?
*Extreme* morphological **difference**, indicating they are in the **same** pathway of development
48
Twist1 AND Chd8 knocked out?
Neural tube ***does not close*** properly, nerves are not formed properly, **not enough forming**.
49
What happens when Otx2 is knocked out in mouse models?
Change in the **amount** and **location** of expression of ***telencephalon markers*** **Morphology** is also changed
50
What human cells can we use to study NDDs?
Human ***embryonic*** stem cells (hESCs) and human ***induced pluripotent stem*** cells (hiPSCs)
51
Brain organoids
Take ***hSCs*** and put in **tissue culture** Make **embryoid bodies** and *induce* them into **neural** cells
52
RNA sequencing
***Reverse transcribe*** RNA fragments > **DNA** fragments > create **library** and compare to ***human genome***
53
Bulk RNA sequencing
Measures the ***average*** gene **expression** level across *all* cells in a sample
54
Single cell RNA sequencing
Examines the gene **expression** profile of ***individual*** cells, when you want to study the *diversity* of cell types within a tissue
55
What are the limitations of single cell RNA sequencing
Input has to be a single cell ***suspension***, **time**, **cost**, **loss** of data.

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