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EMBRYO SQ 11-12 FULL.doc

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University of Waterloo
BIOL 303
Dragana Miskovic

1 1. What parts do each somite become divided into (5 parts)? - Sclerotome (vertebrae, ribs, rib cartilage) - myotome (musculature of back, ribs, limb muscle) - dermatome (back dermis) - endothelial cells (vascular cells in dorsal aorta) - syndotome (tendon) 2. Which proteins control somite boundaries? - Notch – receptor protein, has signal pathway associated with gene expression for boundary formation o Stimulate gene expression of Hairy1 - Delta-like3 – ligand for notch to initiate signal pathway (Juxtacrine signaling) - FGF – expression trigger oscillating clock signal to make Notch - Hairy1 – define somite posterior end - Ephrin-B2 – actual ligand for somite and unsegmented paraxial mesoderm boundary formation - EphA4 (tyrosine kinase) – receptor in anterior of unsegmented paraxial mesoderm 3. Explain the results of the following experiment involving protein Notch 1. - Quail presumptive boundary cells transplanted into unsegmented paraxial mesoderm of chick - Result = boundary formed 2 2. - Quail non-boundary cells transplanted into unsegemented paraxial mesoderm of chick - Result = no boundary induced 3. - Quail non-boundary cells tramsplated into unsegmented paraxial mesoderm of chick followed by LOCAL ECTOPIC (unusual place) EXPRESSION OF NOTCH via induction of graft by electroporation to activate Notch - Result = somite boundary induced 4. How does protein Notch influence gene expression? - Notch is receptor for delta ligand - Binding cause notch domain to be cleaved and that polypeptide piece goes into the nucleus to influence gene expression (i.e. form gaps b/w segments) 5. What is the current theory explaining somite formation involving Notch and Hairy 1 (among other factors)? - Theory = clock and wave mechanism that follows Hensen node retraction - FGF expression trigger oscillating “clock” signal that causes o Notch’s (as transcription factor) gene expression of Hairy1 (transcription factor) and Notch inhibitor  Hairy1 = posteriorizes somite  As more notch is created from Wnt pathway, more notch inhibitor made = decreasing levels of notch = decreasing levels of inhibitor = eventual re-increase of notch = wave • Simple negative feedback loop (sinusoidal expression) o Expression of Hairy1 allow for determination of the posterior of each somite 6. What is the role of ephrin and ephrin recepror in somite formation? 3 - The ephrin ligand (ephrin-b2) and tyrosine kinase receptor (EphA4) allows somites to separate from unsegmented paraxial mesoderm o allows for somite repulsion and separation from unsegmented paraxial mesoderm o ephrin prevents NCC movement in certain areas  expression modification can cause cytoskeleton rearrangements 4 Describe the process of somite epithelization. What are the roles of fibronectin and N- cadherin? - After somite formation Ectodermal signals initiates mesenchymal-to- epithelial transition (MET) - Form basal membrane, sub-apical surface - Fibronectin – ECM organizing protein - N-cadherin – adhesion protein o Work with fibronectin to organize outer layers into epithelium posterior to anterior (for chickens) - Epithelialization occur in posterior part of anterior somite (top of gap to bottom of gap o Central cells remain mesenchymal while outer cells become epithelial layer - Somite forms ANTERIOR TO POSTERIOR - Epithelialization forms POSTERIOR TO ANTERIOR - Gap forms within anterior mesenchymal Presomitic mesoderm (PSM) Cells posterior to the gap undergo epithelialization at later stage than cells at the gap anterior Cells at anterior part of gap will complete epithelialization first and become the POSTERIOR edge of somite 5 7. Do somites continue to exist in the body, or are they only temporary present in the embryo? - Somites are only temporarily present in the embryo (transient structures) - Important for subsequent development o Vertebrae, ribs, dorsal skin dermis, back skeletal muscle, body walls, limbs (bones, muscle, skin, limbs) o Determine migration path of NCC and spinal nerve axons (function) 8. Draw the relative geometric shapes and positions of the dermatome, sclerotome and two subdivisions of the myotome in respect to the notochord, neural tube, lateral plate mesoderm and overlaying epidermis. Why is this particular arrangement important? This particular arrangement is important because the placement of morphogen generating structures determines what types of somites get exposed to what and at what concentration. Those factors determine the developmental fate of the somites. 9. Somites first subdivide into sclerotome, dermatome and myotome. What signals are involved in these subdivisions and where are they coming from? L12S9 Structure Exposed to Becomes High conc. Sonic Hedgehog Sclerotome (shh) from notochord, Cartilage, vertebrae floor plate (Pax1) Induces sclerotome cells to secrete Pax1 transcription factor Dermatome Neurotropin3 (NT3) and adipocyte Wnt1 (Shh antagonist) from roof plate Primaxial – low Myotome concentration of Wnt Dermatome, myotome, sclerotome Abaxial – Wnt (epidermal), BMP4, Fgf5 (lateral plate mesoderm) 6 10. Which cells (derivatives of what) are forming an anterior part of
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