Index
Very early development: ..................................................................................................................... 3
Gastrulation and mesoderm induction ................................................................................................... 4
Patterning of the germ layers.......................................................................................................... 4
Mesoderm induction: ...................................................................................................................... 4
How to put a break on the mesoderm induction ............................................................................ 5
Early development and germ layers in mouse ................................................................................ 5
Cleavage and formation of the blastula .......................................................................................... 6
A-P patterning in mice ..................................................................................................................... 7
Symmetry breaking by migration of visceral endoderm to anterior............................................... 7
Vertebrate axis formation ............................................................................................................... 8
Mesoderm induction in mouse ....................................................................................................... 8
In mice, the same BMP antagonist are common then in Xenopus ................................................. 9
Morphogenesis ...................................................................................................................................... 10
Cadherins ....................................................................................................................................... 10
Immunoglobulin superfamily ........................................................................................................ 11
Integrins ......................................................................................................................................... 11
Actin filaments............................................................................................................................... 12
Involution (Xenopus): .................................................................................................................... 12
Convergent extension ................................................................................................................... 12
Epiboly ........................................................................................................................................... 14
Gastrulation in zebrafish ............................................................................................................... 14
Morphogenesis ...................................................................................................................................... 15
A-P patterning and Hox gene function .......................................................................................... 15
Temporal order of somite formation ............................................................................................ 15
FGF signalling ................................................................................................................................. 16
Retinoic Acid (RA) .......................................................................................................................... 16
FGF and RA gradients during somite formation ............................................................................ 16
The clock and wavefront model .................................................................................................... 17
Molecular mechanism of segmentation ....................................................................................... 18
Hox gene function ......................................................................................................................... 18
Function of several members of Hox genes .................................................................................. 19
Ectopic expression of Hox genes ................................................................................................... 20
Molecular control of Hox gene expression ................................................................................... 20
Organogenesis ....................................................................................................................................... 21
1
, Antero-Posterior Patterning and Organogenesis .......................................................................... 21
AER is required for limb outgrowth............................................................................................... 21
FGF signaling for limb outgrowth .................................................................................................. 21
Proximo-distal patterning.............................................................................................................. 21
Zone of polarizing activity (ZPA) .................................................................................................... 22
Sonic Hedgehog signaling in limb (Shh)......................................................................................... 22
Dorso-Ventral patterning .............................................................................................................. 23
Gene regulation in limb development and disease....................................................................... 24
Germ cells .............................................................................................................................................. 26
The development of germ cells ..................................................................................................... 26
Meiosis .......................................................................................................................................... 29
Genomic imprinting ............................................................................................................................... 31
Fertilization............................................................................................................................................ 34
Fertilization of a mammalian egg .................................................................................................. 34
Calcium wave at fertilization ......................................................................................................... 36
Assisted reproductive technology ................................................................................................. 37
Determination of sexual phenotype ............................................................................................. 38
Determination sex of the germ cells ............................................................................................. 40
Summary: ...................................................................................................................................... 41
Dosage compensation ................................................................................................................... 41
How do cells count and choose chromosomes for inactivation?.................................................. 42
Cell differentiation................................................................................................................................. 44
Transcription regulation in cell differentiation ............................................................................. 44
Cells stop dividing during differentiation ...................................................................................... 48
Cell cycle and differentiation factors are biochemically linked .................................................... 48
Stem cells and their niche ............................................................................................................. 49
How would this work molecularly in response to physiological needs? ....................................... 50
Different modes of self-renewal contribute to tissue homeostasis.............................................. 51
Cellular (re)programming .............................................................................................................. 52
Another way to induce cells to change fate: ................................................................................. 54
Cloning by nuclear transfer ........................................................................................................... 54
Stem cells in homeostasis, repair & regenerative medicine ......................................................... 55
Genetic lineage tracing can be used to follow cell fate ................................................................ 56
Evo devo ................................................................................................................................................ 58
2
, - Specification: Cells have received sufficient information to develop according to their normal
fate, for example when isolated. However, they can still be re-specified to a different fate
when transplanted to a different position, meaning they are not yet determined.
- Determination: Stable change in fate (irreversible). Cells can no longer be re-specified.
- Differentiation: Cell become functionally different, become different cell types.
Very early development:
- Maternal to zygotic transition (MTZ)
o Mid-blastula stage (xenopus & zebrafish)
o Maternal RNA degradation → zygotic genome activation (ZGA)
o Mid-blastula transition: when transcription starts. MBT used in flies, fish, frogs
MBT: Transition at the mid-blastula stage, involving (1) onset of mbryonic transcription, (2)
cell cycle lengthening, (3) acquisition of cell motility
- Dorsal ventral body axis (after ZGA)
o Frogs: sperm entry (future ventral side) → cortical rotation (V→D) with wnt (11 and
5a) signals to the nucleus via B-catenin → accumulation of B-catenin in nuclei on the
future dorsal side of the blastula, VegT is also present to express the transcription
factor Siamosis which activate goosecoids (important in the process for making the
organizer) which defines the Nieuwkoop centre
When there is free β-catenin, normally, in the absence of Wnt, it is degraded by the APC/Axin/GSK-
3/CK1γ complex. This way β-catenin cannot be transported to the nucleus. When Wnt is present, Wnt
binds to Frizzled (Wnt receptor) and the APC/Axin/GSK-3/CK1γ complex binds to phosphorylated
LRP6/Frizzled. Β-catenin accumulates and binds in the nucleus to the transcription factor TCF.
Transcription takes place in the nucleus. On the future dorsal side, the Wnt signalling pathway is
activated. Wnt11b mRNA and other determinants (Dishevelled) relocate to prospective dorsal region.
There is dorsal β-catenin accumulation. The sperm entry point, which is random, will determine where
dorsal and ventral sides will be.
The Tcf3 transcription factor translocate its cofactor β-catenin to the nucleus.
• β-catenin can induce dorsal axis at ventral side
• Knockdown of fzd7 (Xfz7, Wnt receptor): Loss of dorsal structures
• Also: Wnt inhibitors ventral (pre-MBT
o Zebrafish: sperm entry not involved, no cortical rotation. Wnt 8a mRNA is
transported to the future dorsal side by oriented parallel arrays of microtubules and
enters the blastoderm (animal side). At the mid-blastula transition: B-catenin moves
into the nuclei of the cells of the dorsal yolk syncytial layer → accumulation → where
it induces the expression of a zygotic gene VegT encoding the transcription factor
Dharma → induction of the organizer/shield genes
Sia1 (frogs) and Dharma (zebrafish) is activated at the MBT in response to Wnt signaling.
Nieuwkoop centre (blastula stage) and Spemann Mangold organizer (gastrula stage). The
Nieuwkoop centre is the organizer of the organizer/shield.
Differences in mesoderm induction by dorsal and ventral vegetal regions.
The dorsal vegetal region of the Xenopus blastula, which contains the
Nieuwkoop center, induces notochord and muscle from animal cap tissues,
whereas ventral vegetal cells induce blood and associated tissues
3
, Gastrulation and mesoderm induction
➔ Fate map, gastrulation
➔ Mesoderm induction
Endoderm & ectoderm is maternally specified.
Patterning of the germ layers
Mesoderm induction: Vegetal region
Maternal VegT in the vegetal region activates the transcription of nodal-related genes (Xnr and
Derrière). The presence of nuclear β-catenin on the dorsal side stimulates nodal-related transcription,
resulting in a dorsal-to-ventral gradient in the Nodal related proteins. These induce mesoderm and, at
high doses, specify the Spemann organizer on the dorsal side(where the most intense Nodal signaling
is).
Mesoderm induction:
Mesoderm in Xenopus is specified by endoderm signals in prospective ectoderm. Mesoderm is not
maternally specified. How do you specify mesoderm?:
Bmp4 helps patterning the mesoderm, specifying ventral mesodermal fate.
• Ectodermin inhibits its signaling in the animal pole, important for ectodermal fate, by ubiquitinating Smad4 (the “co-Smad”). This
restricts both BMP and Nodal signaling to equatorial zone which then will form mesoderm.
• BMP signaling antagonists like noggin, chordin, Cerberus inhibit its signaling at the dorsal side (Organizer), restricting ventral fate to
ventral and lateral positions in the margin
Signals:
- Foxi1e (foxi1): activated ectoderm formation and controls cell position in Xenopus blastula
(prevent mixing)
- VegtT: specifies endoderm
- Ectodermin (Trimm33): specify ectoderm
- Vg-1 (gdf1) TGFB family: mesoderm induction
Transcription factors respond to TGFB family gradients and act as master regulators
4
Very early development: ..................................................................................................................... 3
Gastrulation and mesoderm induction ................................................................................................... 4
Patterning of the germ layers.......................................................................................................... 4
Mesoderm induction: ...................................................................................................................... 4
How to put a break on the mesoderm induction ............................................................................ 5
Early development and germ layers in mouse ................................................................................ 5
Cleavage and formation of the blastula .......................................................................................... 6
A-P patterning in mice ..................................................................................................................... 7
Symmetry breaking by migration of visceral endoderm to anterior............................................... 7
Vertebrate axis formation ............................................................................................................... 8
Mesoderm induction in mouse ....................................................................................................... 8
In mice, the same BMP antagonist are common then in Xenopus ................................................. 9
Morphogenesis ...................................................................................................................................... 10
Cadherins ....................................................................................................................................... 10
Immunoglobulin superfamily ........................................................................................................ 11
Integrins ......................................................................................................................................... 11
Actin filaments............................................................................................................................... 12
Involution (Xenopus): .................................................................................................................... 12
Convergent extension ................................................................................................................... 12
Epiboly ........................................................................................................................................... 14
Gastrulation in zebrafish ............................................................................................................... 14
Morphogenesis ...................................................................................................................................... 15
A-P patterning and Hox gene function .......................................................................................... 15
Temporal order of somite formation ............................................................................................ 15
FGF signalling ................................................................................................................................. 16
Retinoic Acid (RA) .......................................................................................................................... 16
FGF and RA gradients during somite formation ............................................................................ 16
The clock and wavefront model .................................................................................................... 17
Molecular mechanism of segmentation ....................................................................................... 18
Hox gene function ......................................................................................................................... 18
Function of several members of Hox genes .................................................................................. 19
Ectopic expression of Hox genes ................................................................................................... 20
Molecular control of Hox gene expression ................................................................................... 20
Organogenesis ....................................................................................................................................... 21
1
, Antero-Posterior Patterning and Organogenesis .......................................................................... 21
AER is required for limb outgrowth............................................................................................... 21
FGF signaling for limb outgrowth .................................................................................................. 21
Proximo-distal patterning.............................................................................................................. 21
Zone of polarizing activity (ZPA) .................................................................................................... 22
Sonic Hedgehog signaling in limb (Shh)......................................................................................... 22
Dorso-Ventral patterning .............................................................................................................. 23
Gene regulation in limb development and disease....................................................................... 24
Germ cells .............................................................................................................................................. 26
The development of germ cells ..................................................................................................... 26
Meiosis .......................................................................................................................................... 29
Genomic imprinting ............................................................................................................................... 31
Fertilization............................................................................................................................................ 34
Fertilization of a mammalian egg .................................................................................................. 34
Calcium wave at fertilization ......................................................................................................... 36
Assisted reproductive technology ................................................................................................. 37
Determination of sexual phenotype ............................................................................................. 38
Determination sex of the germ cells ............................................................................................. 40
Summary: ...................................................................................................................................... 41
Dosage compensation ................................................................................................................... 41
How do cells count and choose chromosomes for inactivation?.................................................. 42
Cell differentiation................................................................................................................................. 44
Transcription regulation in cell differentiation ............................................................................. 44
Cells stop dividing during differentiation ...................................................................................... 48
Cell cycle and differentiation factors are biochemically linked .................................................... 48
Stem cells and their niche ............................................................................................................. 49
How would this work molecularly in response to physiological needs? ....................................... 50
Different modes of self-renewal contribute to tissue homeostasis.............................................. 51
Cellular (re)programming .............................................................................................................. 52
Another way to induce cells to change fate: ................................................................................. 54
Cloning by nuclear transfer ........................................................................................................... 54
Stem cells in homeostasis, repair & regenerative medicine ......................................................... 55
Genetic lineage tracing can be used to follow cell fate ................................................................ 56
Evo devo ................................................................................................................................................ 58
2
, - Specification: Cells have received sufficient information to develop according to their normal
fate, for example when isolated. However, they can still be re-specified to a different fate
when transplanted to a different position, meaning they are not yet determined.
- Determination: Stable change in fate (irreversible). Cells can no longer be re-specified.
- Differentiation: Cell become functionally different, become different cell types.
Very early development:
- Maternal to zygotic transition (MTZ)
o Mid-blastula stage (xenopus & zebrafish)
o Maternal RNA degradation → zygotic genome activation (ZGA)
o Mid-blastula transition: when transcription starts. MBT used in flies, fish, frogs
MBT: Transition at the mid-blastula stage, involving (1) onset of mbryonic transcription, (2)
cell cycle lengthening, (3) acquisition of cell motility
- Dorsal ventral body axis (after ZGA)
o Frogs: sperm entry (future ventral side) → cortical rotation (V→D) with wnt (11 and
5a) signals to the nucleus via B-catenin → accumulation of B-catenin in nuclei on the
future dorsal side of the blastula, VegT is also present to express the transcription
factor Siamosis which activate goosecoids (important in the process for making the
organizer) which defines the Nieuwkoop centre
When there is free β-catenin, normally, in the absence of Wnt, it is degraded by the APC/Axin/GSK-
3/CK1γ complex. This way β-catenin cannot be transported to the nucleus. When Wnt is present, Wnt
binds to Frizzled (Wnt receptor) and the APC/Axin/GSK-3/CK1γ complex binds to phosphorylated
LRP6/Frizzled. Β-catenin accumulates and binds in the nucleus to the transcription factor TCF.
Transcription takes place in the nucleus. On the future dorsal side, the Wnt signalling pathway is
activated. Wnt11b mRNA and other determinants (Dishevelled) relocate to prospective dorsal region.
There is dorsal β-catenin accumulation. The sperm entry point, which is random, will determine where
dorsal and ventral sides will be.
The Tcf3 transcription factor translocate its cofactor β-catenin to the nucleus.
• β-catenin can induce dorsal axis at ventral side
• Knockdown of fzd7 (Xfz7, Wnt receptor): Loss of dorsal structures
• Also: Wnt inhibitors ventral (pre-MBT
o Zebrafish: sperm entry not involved, no cortical rotation. Wnt 8a mRNA is
transported to the future dorsal side by oriented parallel arrays of microtubules and
enters the blastoderm (animal side). At the mid-blastula transition: B-catenin moves
into the nuclei of the cells of the dorsal yolk syncytial layer → accumulation → where
it induces the expression of a zygotic gene VegT encoding the transcription factor
Dharma → induction of the organizer/shield genes
Sia1 (frogs) and Dharma (zebrafish) is activated at the MBT in response to Wnt signaling.
Nieuwkoop centre (blastula stage) and Spemann Mangold organizer (gastrula stage). The
Nieuwkoop centre is the organizer of the organizer/shield.
Differences in mesoderm induction by dorsal and ventral vegetal regions.
The dorsal vegetal region of the Xenopus blastula, which contains the
Nieuwkoop center, induces notochord and muscle from animal cap tissues,
whereas ventral vegetal cells induce blood and associated tissues
3
, Gastrulation and mesoderm induction
➔ Fate map, gastrulation
➔ Mesoderm induction
Endoderm & ectoderm is maternally specified.
Patterning of the germ layers
Mesoderm induction: Vegetal region
Maternal VegT in the vegetal region activates the transcription of nodal-related genes (Xnr and
Derrière). The presence of nuclear β-catenin on the dorsal side stimulates nodal-related transcription,
resulting in a dorsal-to-ventral gradient in the Nodal related proteins. These induce mesoderm and, at
high doses, specify the Spemann organizer on the dorsal side(where the most intense Nodal signaling
is).
Mesoderm induction:
Mesoderm in Xenopus is specified by endoderm signals in prospective ectoderm. Mesoderm is not
maternally specified. How do you specify mesoderm?:
Bmp4 helps patterning the mesoderm, specifying ventral mesodermal fate.
• Ectodermin inhibits its signaling in the animal pole, important for ectodermal fate, by ubiquitinating Smad4 (the “co-Smad”). This
restricts both BMP and Nodal signaling to equatorial zone which then will form mesoderm.
• BMP signaling antagonists like noggin, chordin, Cerberus inhibit its signaling at the dorsal side (Organizer), restricting ventral fate to
ventral and lateral positions in the margin
Signals:
- Foxi1e (foxi1): activated ectoderm formation and controls cell position in Xenopus blastula
(prevent mixing)
- VegtT: specifies endoderm
- Ectodermin (Trimm33): specify ectoderm
- Vg-1 (gdf1) TGFB family: mesoderm induction
Transcription factors respond to TGFB family gradients and act as master regulators
4
