Early Development Amphibians

Cleavage in a frog Describe the progression of events. differences in size of blastomeres morula and blastula stages; presence of blastocoel cell held together by EP-cadherin

Gastrulation in Xenopus What have fate maps told us about tissue origins?

Gastrulation in Xenopus What are VegT and Vg1 and what influcence do they have? VegT – transscription factor; Vg1 – paracrine factor;mRNA’s located in cortical cytoplasm of unfertilized egg needed for prospective fate development

Gastrulation in Xenopus What type of cell movement initiates gastrulation? invagination of cell in grey crescent region on future dorsal side; shape chages, formation of bottle cells

Gastrulation in Xenopus What happens next? involution of marginal zone cells; epiboly of animal pole cells always new cells at dorsal lip; involution of marginal zone cells first cells are cells that become pharyngeal cells, then precursor of head mesoderm, then chordomesoderm get displacement of blastocoel, formation of archenteron; involution of lateral and ventral lips; yolk plug

Gastrulation in Xenopus What happens at the mid-blastula transition in preparation for gastrulation? How is dorsal-ventral axis established? What happens if cortical rotation is blocked? demethylation of certain promoters; allows transcription factors like VegT to initiate new transcription activation of zygote genome dorsal side forms 180 degrees from sperm entry site; blastopore no dorsal development rotation of egg can change site of gastrulation

Gastrulation in Xenopus How did Gimlich and Gerhart demonstrate that certain cells were responsible for inducing dorsal lip formation?

Gastrulation in Xenopus What appears to be the major factor directing movement of cells into embryo? vegetal rotation – superficial layer of marginal cells is pulled inward because it is attached to actively migrating deep cells

Gastrulation in Xenopus What other cell movements occur? intercalation of IMZ leading to convergent extension

Gastrulation in Xenopus What two factors appear to drive convergent extension? adhesion proteins paraxial and axial protocadherin are expressed changes in calcium concentrations

Gastrulation in Xenopus At what point in gatrulation do notocordal cells become distinct? toward the end – perhaps because of different adhesion molecules

Gastrulation in Xenopus What mechanism appears to be responsible for epiboly of the ectoderm? increase in cell number and intercalation of several layers of cells

Axis Formation How did Hans Spemann first demonstrate the importance of the grey crescent? newt –Triturus taeniatus

Axis Formation What did Spemann do to demonstrate changes in cell potency during gastrulation?

Axis Formation What classic experiment was done by Spemann and Mangold that led to the concept of the organizer? Triturus taeniatus and Triturus cristatus called primary embryonic induction

Axis Formation How did work of Nieuwkoop and Nakamura contribute to an understanding of how the “organizer” worked? combined animal and vegetal caps to generate mesodermal tissue

Axis Formation What is the Nieuwkoop center? How is it formed? dorsalmost vegetal cells capable of inducing the organizer created by cytoplasmic rotation at fertilization

Axis Formation What is involved in the mechanism of induction of mesoderm by endoderm? What appears to be the role of β-catenin? What is the connection between β-catenin and glycogen synthase kinase – 3 (GSK-3)? endodermal cells secrete activin like (derriere) factor that causes expression of Brachyury gene in mesodermal cells Brachyury protein acts as a transcription factor to induce eFGF that maintains Xbra expression and activates genes that produce mesodermal specific proteins factor forming Nieuwkoop center; key player in formation of dorsal structures GSK-3 negatively regulates β-catenin

Axis Formation How does β-catenin become concentrated in dorsal cells of embryo? initially made throughout embryo; degraded by GSK-3 in ventral cells disheveled, an inhibitor of GSK-3 is translocated to doral side during cortical rotation

Axis Formation How does β-catenin contribute to formation of the organizer? TGF-β factor may be Vg1 or nodal proteins

Axis Formation What model was developed by Agius to explain mesodermal induction?

The Organizer What are the four major functions of this tissue? differentiate into dorsal mesoderm induce formation of paraxial mesoderm induce formation of neural tube initiate gastrulation movements

The Organizer What is the role of the goosecoid gene? Abilitiy of goosecoid to induce new axis a)uninjected embryo – one dorsal lip b) injected with goosecoid into ventral vegetal blastomeres – two dorsal lips c) top embryos show dorsal axis d) twinned embryo from injection gene codes DNA binding protein that activates migration properties of cells, determines fate of dorsal mesodermal cells, recruits neighboring cells into dorsal axis, activates a gene needed for brain formation goosecoid works in nucleus of cells

The Organizer How does the organizer actually induce formation of neural tissue? induction actually lead to formation of epidermal tissue from ectoderm – via BMP organizer secretes molecules that block induction – get default neural tissue

The Organizer What are the proteins secreted by the organizer?

The Organizer What can the secreted protein noggin do? What other proteins exhibit similar effects? rescue UV irradiated embryos induces dorsal ectoderm to form neural tissue and dorsalizes mesodermal cells binds to BMP-2 and BMP-4 and prevents their binding to receptors chordin and nodal-related 3, follistatin

The Organizer What are some of the important effects of BMP-4? activates expression of genes encoding transcription factors that induce expression of epidermal and ventral mesodermal differentiation – suppress genes that produce neural phenotype low dose causes muscle to form; intermediate dose causes kidney cells; high dose causes blood cells to form

The Organizer What are the functions of the diffusable proteins cerberus, dickkopf and Frzb? How do these proteins work? secreted by pharyngeal endoderm and prechordal mesoderm and induce most anterior head structures block wnt pathway and BMP-4

The Organizer

The Organizer Why is neurogenin so important? produces neural phenotype

Induction How did Otto Mangold demonstrate the regional specificity of neural induction? portion of archenteron roof transplanted determined neural structures that formed

Induction Toivonen and Saxen postulated two gradients in amphibian embryos dorsal gradient – “neuralizing” activity caudal gradient – “posteriorizing” activity What is the primary protein involved in posteriorizing the neural tube? Xwnt8 protein – can add to developing embryos in anterior region and alter course of development; can suppress wnt signaling and get more neural cells BMP specifies dorsal-ventral axis; wnt specifies anterior-posterior axis other factors like FGF and retinoic acid may be involved in a-p axis

Induction What positive signal promotes anterior head development? insulin-like growth factors

Left-Right Axis What appears to be major event in left-right axis formation in Xenopus? expression of Xnr1 in lateral plate mesoderm on left side may be initiated at fertilization via Vg1 protein on left side