| |
Fig. 3. Recombinations of Spemann organiser (SO) and ventralised embryos (containing only non-organiser mesoderm) as indicated in the schematic drawing
(A). (B –F) Stage 10 to 10+ organiser tissue was implanted into the marginal zone of stage 10 ventralised embryo. Analysis at stage 27 (lateral views). A nontreated
control embryo (con), a ventralised embryo without graft (UV) and ventralised embryos implanted with organiser mesoderm (UV + SO). Probe
combinations are indicated. Arrowheads show the distance between the most posterior Krox-20 expression and the anterior Hox expression boundary. The
normal spatially colinear Hox sequence is restored by organiser transplantation. (G–N) Organiser tissue was explanted from dorsal blastopore lips of stage 10
to 10+ and stage 11.5, respectively, and implanted into the marginal zone of stage 10+ ventralised embryos. Analysis at stage 27 (lateral views). Shown are nontreated
controls (con), ventralised embryos without grafts (UV) and recombinations with early organiser (10 + UV + 10 + SO) and late organiser (10 + UV +
11.5 SO), respectively. Probe combinations are indicated. Arrowheads show the distance between the most posterior Krox-20 expression and the anterior Hox
expression boundary. |
| |
Fig. 4. Timed interactions between the Spemann organiser and the non-organiser mesoderm (NOM). (A) Ageing the non-organiser mesoderm (isolated in ventralised embryos). A ventralised embryo with no
implant (UV), an untreated control embryo (con), and recombinations of organiser mesoderm from stage 10 (0h SO) with ventralised embryos of different ages after the beginning of gastrulation (0h, 2h, 4h, 6h
NOM). Embryos are positioned with their head up and dorsal to the right. They were analysed with WISH using axial markers, including En-2 (midbrain –hindbrain border), Krox-20 (hindbrain), Hoxb-4 (posterior
hindbrain), Hoxc-6 and Hoxa-7 (anterior spinal cord), Hoxd-13 (posterior spinal cord). Expression of Krox-20 (arrow heads) and Hoxd-13 illustrates the results. Pictograms indicate restored part of axis (based on
conclusion from all markers). (B) Ageing the Spemann organiser. A ventralised embryo without implant (UV), an untreated control (con), and recombinations of stage 10 ventralised embryos (0h NOM) with
organiser tissue (SO) aged for 0h, 2h, 4h, 6h after beginning of gastrulation. Embryos orientated and WISH analysed as in (A). Krox-20 expression (arrowheads) and Hoxd-13 illustrate the results. Pictograms
indicate restored part of axis. The age of the organiser implant does not affect the restored axial values. (C) Timed restoration of organiser functions by Noggin protein (nog) injection. Ventralised embryos were
injected with Noggin protein into the blastocoel (schematic drawing) at different blastula and gastrula stages. Embryos were analysed as above. Left panel stained for En-2/Krox-20/Hoxc-6/Xbra, right panel for
Krox-20/Hoxd-13. Embryos are orientated as in (A), arrows point to Krox-20 expression. Top, non-injected ventralised embryos (UV). Rows 2– 5 show ventralised embryos injected with Noggin at the indicated
stages. Bottom, control embryos (con). Early-treated embryos restore head (grey colour in the corresponding pictograms) and anterior trunk (Krox-20 expression, blue colors in pictograms). Later-treated embryos
show progressively less head (grey) and more trunk (anterior trunk marked by Krox-20 and blue colors in pictograms, posterior trunk marked by Hox genes and Xbra, yellow and red colors in pictograms). Very
late on, there is an extensive zone of Hoxd-13 expression (posterior trunk) and anterior trunk markers (e.g., Krox-20) have reached the anterior end of the embryo. |
