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Fig. 5. Noelin-4 over-expression results in expansion of neural fate. Embryos injected with Noelin-4 exhibit enlarged and ectopic neural tissue. Embryos were injected with 500 pg (A - G, P - S) or 1 ng (H - O) of Noelin-4 mRNA, 500 pg Noelin-3 (T, U), and 100 pg of lac Z mRNA as a lineage tracer. h-galactosidase activity was assessed with X-gal (turquoise, A-G) or Magenta-Gal (magenta, H-O) and embryos were stained for Sox-2 expression. (A-C) Neural plate stage embryos injected with (A) Noelin-4 (AMY), (B) Noelin-2 (AMZ) or (C) lacZ. Injected side is to the left, embryos are photographed from the dorsal side, oriented with anterior pointing up. Anterior neural plate is indicated with an arrowhead. Note expansion of the neural plate in panel A. (D - G) Embryos were harvested at stage 29 and stained for Sox-2 expression. (D) Control side, (E) injected side of same embryo. Neural retina is revealed by Sox-2 staining, note that injected side eye is larger than control side (arrowheads). (F) Cross section through midbrain region; (G) comparison of neural retinas from both sides of the embryo magnified to the same degree. (H - O) Examples of embryos harvested at stage 35 and stained for Sox-2 expression. (H, L) Control side; (I, M) injected side. Panels J, N, and O are cross- sections through the head regions with the section level indicated on panels I and M. Panel K is a magnification of the injected side in panel J. Note expansion of neural tissue and ectopic Sox-2 staining (arrowhead). Panel O shows an ectopic cement gland ventral to the eye (arrow) that is discontinuous with the endogenous cement gland (see arrowhead in panel M). (P-S) N-tubulin staining of embryos injected with Noelin-4 (P, R) or Noelin-1 (Q, S). (P, Q) Neural plate stage embryos, with injected side to the left, anterior pointing to top. (R, S) Embryos at stage 24 with anterior to the left. Arrowheads indicate bulging tissue on the injected side in panels P, R. (T, U) Noelin-3 causes similar neural territory expansion. Noelin-3 (BMY) injection, Sox-2 staining. Bars and brackets outline width of neural structures in two different embryos on the injected side (bottom half) and uninjected control side (top half). |
