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Vertebrate development > Pre and post-implantation > Flashcards

Pre and post-implantation Flashcards

1
Q

Characteristics of mammalian embryos

A

> small egg
no yolk
fertilisation + development in maternal environment
slow early division
cleavage -> formation of equal blastomeres
lineage commitment occurs early in development
regulated development

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2
Q

Zona pellucida

- what is it?

A

=Translucent matrix of glycoproteins

  • Surrounds mammalian oocyte
  • Critical to successful fertilisation
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3
Q

Zona pellucida

- function

A

> Only allows species-specific fertilisation
Prevents polyspermy
Enables acrosome reaction for successful adhesion + penetration of sperm

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4
Q

ZP proteins

  • what are they?
  • types?
A

Bind to capacitated spermatozoa

ZP1, ZP2 + ZP3

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5
Q

Roles of different ZPs

A 
ZP3 = allows species-specific sperm binding
ZP2 = mediates subsequent sperm binding
ZP1 = cross-links ZP2 +ZP3
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6
Q

Aim of early stages of development

A

Fertilised egg -> multicellular state

Set aside cells that form embryo + those that form extraembryonic membranes

Formation of maternal foetal connection
= placenta

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7
Q

Types of cleavage depend on…

A

Yolk content

Distribution of yolk

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8
Q

Cleavage in mammalian embryos

- 1st and 2nd cleavage

A

1st = meridional
- pole to pole

2nd = equatorial
- rotational

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9
Q

Cleavage in mice

A

Rotational + holoblastic

Gives rise to equal size blastomeres

Each of blastomeres at 2 + 4 cell stage = totipotent

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10
Q

Stages in mouse pre-implantation development

A
  1. Cleavage gives rise to blastomeres
  2. Compaction + formation of morula
  3. 1st differentiation of cells at morula + blastocyst stages
    - lineage allocation
  4. Hatching out of zona pellucida + implantation
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11
Q

Cleavage

- define

A

Cell division without intervening growth

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12
Q

Cell cycle during first 2 cleavage stages

in mice

A

Slow

- approx 2 days

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13
Q

Pre-natal diagnosis

A

16 cell stage
- culture for 6-8hrs

  1. Single cell removed
  2. Genotyping by PCR
  3. If healthy, transfer embryo to to uterus
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14
Q

How can you take a cell out of an embryo?

A

Embryo is regulated

  • can take 1 cell out and won’t affect development
  • > regulated back to normal size

(Same for adding 1 cell)

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15
Q

Pre-implantation development

- compaction

A

Cells change shape

Gap and tight junctions form

Outer cells become differentiated from inner cells
(diff genes expressed)

Tightly packed mass

Polar cells

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16
Q

Polarity

A

Asymmetric organisation of several cellular components

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17
Q

Setting up the 2 main lineages

A

Outer cells of morula form Trophectoderm (TE)

Inner cells form Inner Cell Mass (ICM)
-> differentiates into Primitive Endoderm (PE) and Epiblast (EPI)

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18
Q

Inside-outside model

A

Cells on inside + outside receive different amounts of cell contact
-> differences in TF expression

Low cell contact
-> Cdx2 expressed

High cell contact
-> Cdx2 inhibited

19
Q

Cell polarity model

A

Presence or absence of an apical domain
-> differences in TF expression

Outside = apical domain
-> Cdx2 expressed

Inside = No apical domain
-> Cdx2 inhibited

20
Q

Hippo signalling

  • apical domain
  • outside cells
A

Apical domain

  • > inhibits hippo signalling
  • > Yap not phosphorylated by Lats1/2
  • > Yap moves into nucleus
  • > Tead4 expresses Cdx2
21
Q

Hippo signalling

  • No apical domain
  • inside cells
A
  • > Hippo signalling occurs
  • > Lat1/2 phosphorylate Yap
  • > can’t move into nucleus
  • > Tead4 can’t express Cdx2
22
Q

Molecular mechanisms underlying the 1st lineage segregation

- TE

A

Hippo pathway

  1. No hippo pathway
  2. Yap activates Tead4
  3. Tead4 expresses Cdx2 + Gata3

Ras/MAPK pathway

  1. Asymmetric cell division
  2. Ras actuvates Erk2
  3. Erk2 activates Tcfap2c
  4. –> expresses Cdx2
23
Q

Molecular mechanisms underlying the 1st lineage segregation

- ICM

A

Hippo pathway

  1. Cell contact + polarity factors
  2. Hippo pathway actuvated
  3. Activates Lats1/2
  4. Phosphorylation of Yap
  5. Prevents Tead4 activation
  6. Prevents Cdx2 + Gata3 expression
24
Q

Molecular players in the formation of the first 3 lineages in the blastocyst
- Oct4

A

Observed in all blastomeres throughout early cleavage stages due to maternally encoded protein

8-cell stage = all blastomeres contain Oct4

Blastocyst stage = Oct4 gradually down regulated in outer TE cells by Cdx2

25
Q

Molecular players in the formation of the first 3 lineages in the blastocyst
- Cdx2

A

Cdx2 protein detected beginning at 8-16 cell stage
- initial expressin = stochastic

Early modula - early blastocyst
= Cdx2 expression ubiquitous but higher in outer, apically polarised cells

Blastocyst
= Restricted expression in outer TE cells

26
Q

Molecular players in the formation of the first 3 lineages in the blastocyst
- Nanog + Gata6

A

Detected from 8 cell stage

Both expressed uniformly in all cells until early blastocyst stage

Late blastocyst stage
- ICM cells express either Nano + Gata6 exclusively

27
Q

Molecular players in the formation of the first 3 lineages in the blastocyst
- Nanog

A

Expression down regulated in outer cells by Cdx2

+ in subpopulation of ICM by Grb2-dependent signalling

28
Q

Molecular players in the formation of the first 3 lineages in the blastocyst
- Gata6

A

Expression maintained by Grb6-dependent signalling

29
Q

Changes during blastocyst formation

A

Epithelialisation of out layer of cells and establishment of polarity

Vectorial fluid transport
-> Blastocoel expands

TE cells give rise to trophoblast

ICM form small cluster
-> give rise to entire org later on

30
Q

Mechanisms underlying 2nd lineage segregation

PE vs EPI

A

Gata6-expressing cells form PE

Nanog-expressing cells form EPI

PE + EPI progenitors initially distributed in ‘salt + pepper’ formation
->sort into 2 organised layers

31
Q

PE vs EPI organised into layers

A

Based on differential adhesions + signals coming from either TE or blastocoel

Nanog-expressing cells
= high adhesion to each other _ low adhesion to Gata6 cells

Gata6 cells
= intermediate adhesion to each other

32
Q

PE vs EPI

- positioning

A

Accurate positon achieved by adding forces exerted by blastocoel or the TE

33
Q

PE vs EPI

- Differential Nanog + Gata6 expression depends on

A

Growth factor Fgf4 + receptor Fgfr2

34
Q

PE vs EPI
- signalling

Nanog-expressing cells

A

Fgfr2 activates expression of Gata6
-> antagonises Nanog

Nanog-expressing cells up-regulate Fgf4 but lose Fgfr2
- inhibits Gata6 transcription

Leads to complete loss of Gata6 in EPI

35
Q

PE vs EPI
-signalling

Gata6-expressing cells

A

Fgfr2 activates expression of Gata6
-> antagonises Nanog

Gata6-expressing cells retain expression on Fgfr2
- Fgfr2 activated by Fgf4 secreted by EPI cells

Nanog in these cells inhibited by Fgdr2 + Gata6

  • Gata6 enhances own expression
  • > loss of Nanog + activation of PE-specifying genes Sox17 + Gata4
36
Q

Which proteins form a regulatory loop required to maintain PE identity?

A

Fgfr2
Gata6
Soz17
Gata4

37
Q

Blastocyst hatching

A

Volume of blastocyst increases 2-3fold
-> trophoblast epithelium stretches due to intake of fluid
= increases hydrostatic pressure within blastocoel

38
Q

Why does blastocyst hatching need to occur?

A

Zona pellucida prevents blastocyst being sticky + implanting

39
Q

Cell lineage formation from egg to egg cylinder

A 
1 cell
2 cell
8 cell
Early morula
(Blastomeres): 
Late morula
Blastocyst 
Late blastocyst 
Early egg cylinder
40
Q

Morula

A

8 cell

Compacted

41
Q

Emergence of asymmetry during pre-implantation development

- blastocyst

A

Site of tethering of 2nd polar body

Tilting of ICM

Position of Cer1 + Lefty-1 expressing cells in primitive endoderm

42
Q

Emergence of asymmetry during pre-implantation development

- Early egg cylinder

A

Tilting of ectoplacental cone away from proximodistal axis

Prospective AP axis aligns with shorter diameter of cylinder

43
Q

Emergence of asymmetry during pre-implantation development

- Embryo with DVE

A

Lopsided distribution of Cer1 + Lefty1-expressing cells in DVE

Localisation of Wnt3 expression on 1 side of visceral endoderm

44
Q

Emergence of asymmetry during pre-implantation development

- embryo with AVE

A

Tilting of ectoplacental cone

Prospective AP axis aligns with longer diameter of cylinder r

Regionalisation of gene expression in the AVE and posterior epiblast

Vertebrate development (12 decks)

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