Developmental Biology – Biology 4361 Axis Formation and Mesoderm Induction October 27, 2005.
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Transcript of Developmental Biology – Biology 4361 Axis Formation and Mesoderm Induction October 27, 2005.
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Developmental Biology – Biology 4361
Axis Formation andMesoderm Induction
October 27, 2005
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Amphibian anteroposterior specification
- polarized eggs – animal/vegetal
- localized cytoplasmic components
- pigment
- yolk v. clear cytoplasm
- mitochondrial cloud
- germinal vesicle
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Figure 8.25
RNA localization – Xenopus oocytes
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Figure 9.7
Anteroposterior axis – VegT depletion
normal
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Figure 9.7
depletion of VegT = - shift from endoderm to mesoderm and ectoderm - mesoderm replaced with ectoderm - animal region forms only epidermis and no nervous system
Anteroposterior axis – VegT depletion
normal VegT - depleted
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Figure 9.18
Dorsalization - Xenopus
UV = ventralized
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Figure 9.15
Transplantation of dorsalizing activity
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Figure 9.19
Early Dorsoventral Determination
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Gray crescent formation
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Cortical rotation and Disheveled
sperm
Dsh
Disheveledprotein (Dsh)
1. Fertilization
2. Cortical rotation
3. Dorsal enrichment of Dsh
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Figure 9.21
gylcogen synthasekinase-3
Disheveled activity
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Figure 9.21
gylcogen synthasekinase-3
Disheveled protein
blocks GSK-3 phosphorylation of -catenin
Disheveled activity
Transcription factor
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Molecular basis of dorsoventral axis
-catenin stabilized
Repressed
TGF-bsignalingpathway
-catenin degraded
Tcf-3proteins
siamoisgene
transcription
transcription
goosecoidgene
Goosecoidprotein
-cateninproteins
siamoisgene
Siamoisprotein
Activated
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Organizer transplant
Spemann’s organizer – dorsal lip of the blastopore
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Organizer transplant
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Gilbert: Developmental Biology, 7th ed (2003) Table 10.2.
chordin
noggin
nodal-related proteins
(several)
XLim1
Xnot
Otx2
XFD1
XANF1
Goosecoid
Cerberus
Follistatin
Frzb
Secreted ProteinsNuclear Proteins
“Organizer” proteins
- expressed almost exclusively in the organizer
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goosecoid mRNA can induce a second dorsal axis:
goosecoid mRNA injection causes formation of a second dorsal blastopore lip
Gilbert: Developmental Biology, 7th ed (2003) Fig 10.28.
Organizer gene activity
produces embryo with two dorsal axes and two sets of head structures
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Rescue of dorsal structures by noggin protein:
Gilbert: Developmental Biology, 7th ed (2003) Fig 10.30.
“overdose” of noggin mRNA causes formation of dorsal structures at the expense of ventral structures
dose-dependent induction of dorsal structures by injection of noggin mRNA
ventralized embryo without dorsal structures (UV-irradiated)
Organizer gene activity
noggin binds to bone morphogenic proteins (BMP2 & BMP4) - inhibits binding BMP receptor binding
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chordin mRNA is localized in the ‘organizer’:
Gilbert: Developmental Biology, 7th ed (2003) Fig 10.32.
Organizer gene activity
- inhibition of BMP4 & BMP2 induces formation of the neural tube in adjacent ectoderm
- chordin protein binds to BMP4 and BMP2 – inhibits receptor BMP-receptor binding
- late in gastrulation, chordin is localized in the dorsal mesoderm of the notochord
- chordin mRNA is found in the dorsal lip
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Figure 9.8
Mesoderm induction - Xenopus
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Figure 9.8
Mesoderm induction - Xenopus
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Figure 9.8
Mesoderm induction - Xenopus
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Figure 9.9a, b
Mesoderm induction - Xenopus
mesoderm inducers:
bFGF
Vg1
activin
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Mesoderm induction, Organizer formation
β-catenin
VegT, Vg1
Nodalrelatedhigh
Nodalrelatedlow
Organizer
Ventralmesoderm
1. β-catenin acts with VegT and Vg1 to activate Xnr genes (Xenopus Nodal-related)
2. Organizer originates in the region where VegT & Vg1 and β-catenin overlap
3. Gradient of Xnr protein specifies mesoderm: low Xnr ventral mesoderm
4. High Xnr levels activate goosecoid and other ‘organizer’ genes
BMP4high
BMP4low
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Left-right asymmetry
Most animals are bilaterally symmetrical (Bilateria)
- however, individuals deviate to some degree from true bilateral symmetry:
- regular asymmetry or directed asymmetry: sidedness is fixed for a species or for a higher taxon
e.g. in humans: - heart on left side - stomach curves to the left - liver & spleen on right side
- fluctuating asymmetry: non-heritable minor left-right differences
- antisymmetry: heritable morphological left-right differences - sidedness is randomly distributed (ca. 50% each)
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- situs inversus: complete reversal of left-right symmetry in all organs- heterotaxis: some organs reversed- isomerism: normally asymmetrical organs duplicated or missing
Left-right asymmetry
Deviation from directed asymmetry is often lethal!
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Left-Right Asymmetry
Mechanistic basis for establishing asymmetry:
- translated into left-right differences at the level of cells, tissues and the whole organism
- chiral molecules may cause “symmetry-breaking” event
(specific orientation of stereoisomeric molecules relative to the body axes)
Candidate chiral molecule: Dynein
- motor protein complex associated with axonemes, cilia
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Fig 2.7.
Dyneins - microtubule-associated motor protein complexes
- chiral: curve clockwise (from base) = ‘handedness’- mediate sliding between adjacent microtubules in cilia or flagella- cause cilia to rotate in a specific direction (clockwise)- monocilia (at Hensen’s node - mouse) generate oriented flow of signal molecules to the left side of the embryo- signal molecules activate or inhibit patterning genes on left side
Axonemal dyneins:
Left-Right Asymmetry
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Iv+ and Inv+
- iv protein is a left-right dynein
- iv-/iv- = no motility, no fluid flow
- randomized L-R asymmetry (lethal)
iv+: ‘situs inversus viscerum’
- wild type & heterozygous embryos turn clockwise
inv+: “inversion of embryonic turning”
- inv-/inv- turn counterclockwise in amniotic cavity
- mechanism of inv action is unknown
- 100 % of homozygotes for inv show situs inversus
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nodal expression in mouse:
Nodal activated by iv,inv
wild type ectopic
Nodal
- nodal is involved in determining left-right asymmetry in mice, frogs, chicken & zebrafish
- ectopic expression of nodal on right side randomizes location of the heart
- mesoderm adjacent to nodal expression develops into asymmetrical organs
- nodal protein synthesized in left lateral plate mesoderm
- nodal gene activated by iv and inv genes
- intracellular protein - TGF-β family
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pitx2 injection in Xenopus:
Pitx2 & lefty activated by iv, ivn, nodal
pitx2+ and lefty+ genes :
- pitx2 expression depends on iv, ivn and nodal genes- pitx2 and lefty encode homeobox transcription factors that regulate genes - both are expressed primarily on left side of vertebrate embryos have been found in all vertebrates studied- injection of ptx2 on right side of embryo - can cause a complete reversal of gut coiling and heart looping
nodal, pitx2 and lefty form an evolutionary conserved signaling system that is involved in regulating left-right asymmetry in all vertebrates