XB-ART-21318Development. May 1, 1994; 120 (5): 1179-89.
The cleavage stage origin of Spemann''s Organizer: analysis of the movements of blastomere clones before and during gastrulation in Xenopus.
Recent investigations into the roles of early regulatory genes, especially those resulting from mesoderm induction or first expressed in the gastrula, reveal a need to elucidate the developmental history of the cells in which their transcripts are expressed. Although fates both of the early blastomeres and of regions of the gastrula have been mapped, the relationship between the two sets of fate maps is not clear and the clonal origin of the regions of the stage 10 embryo are not known. We mapped the positions of each blastomere clone during several late blastula and early gastrula stages to show where and when these clones move. We found that the dorsal animal clone (A1) begins to move away from the animal pole at stage 8, and the dorsal animal marginal clone (B1) leaves the animal cap by stage 9. The ventral animal clones (A4 and B4) spread into the dorsal animal cap region as the dorsal clones recede. At stage 10, the ventral animal clones extend across the entire dorsal animal cap. These changes in the blastomere constituents of the animal cap during epiboly may contribute to the changing capacity of the cap to respond to inductive growth factors. Pregastrulation movements of clones also result in the B1 clone occupying the vegetal marginal zone to become the primary progenitor of the dorsal lip of the blastopore (Spemann''s Organizer). This report provides the fundamental descriptions of clone locations during the important periods of axis formation, mesoderm induction and neural induction. These will be useful for the correct targeting of genetic manipulations of early regulatory events.
PubMed ID: 8026328
Article link: Development.
Grant support: NS23158 NINDS NIH HHS
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|Fig. 1. The location and nomenclature of blastomeres used in this study. (A) 16-cell embryo (Hirose and Jacobson, 1979). (B) 32-cell embryo labeled with the nomenclature of Jacobson and Hirose (1981). (C) 32-cell embryo labeled with the nomenclature of Nakamura and Kishiyama (1971).|
|Fig. 2. Camera-lucida drawings of labeled surface cells (stippled), members of clones of 16-cell blastomeres, at stage 7. Each clone retains the shape and position of its progenitor blastomere, with interdigitation of cells at the edge of the clone. (A) D1.1, animal view, dorsal to the top. (B) D1.2, animal view, dorsal to the top. (C) D2.1, vegetal view, dorsal to the top. (D) V2.1, view of ventral midline, animal to the right.|
|Fig. 3. Camera-lucida drawings of labeled members of 16-cell clones (stippled) in parasagittal (A), sagittal (B,D) and coronal (C) sections of stage 7 embryos. (A) The D1.2 clone mixes with neighboring clones by one or two cell diameters at the marginal zone edge. (B) The D1.1 clone contains numerous unlabeled cells 3- to 6-cell diameters from the edge of the clone. This is especially notable at the animal cap border. (C) The D2.1 clone, viewed from just below the floor of the blastocoel, mixes by one cell diameter with neighboring clones at several sites. (D) The V2.1 clone barely interdigitates with the clone of its lateral neighbor.|
|Fig. 6. Summary diagrams illustrating the locations of the clones derived from the midline blastomeres of the 32-cell embryo (A) at stage 8 (B), stage 9 (C), stage 10 (D), stage 11 (E), and stage 12.5-13 (F). Data were derived from tissue sections in which two adjacent blastomere clones were labeled with different lineage dyes. These illustrations are composite summaries of at least three embryos per blastomere pair. Diagrams are oriented as in Fig. 1. Clones are represented by the following colors: lilac, C4; orange, B4; green, A4; yellow, A1; red, B1; blue, C1; purple, D1. Illustrations in B-F are based on midsagittal photomicrographs by Hausen and Riebesell (1991).|
|Fig. 4. At stage 7, mixing of the D1.1 clone is primarily with its contralateral neighbor. One D1.1 blastomere was labeled with FDA (green), the contralateral D1.1 blastomere was labeled with TRDA (red). Transverse section, dorsal is to the top. Bar equals 200 mm in all color photomicrographs. Fig. 5. At stage 8, descendants of A4 (red) were found one or two cell diameters on the dorsal side of the animal pole (line), while the animalmost descendants of A1 (green) no longer reached the animal pole. Sagittal section, dorsal is to the right. Fig. 7. At stage 9, the A4 clone (green) has moved into the dorsal part of the animal cap. Its deep cells underlie more superficial cells of the B4 clone (red) in the blastocoel roof. Sagittal section, dorsal is to the left, animal pole is marked by line. Fig. 8. At stage 9 both the B1 clone (red) and the C1 clone (green) have moved into the vegetal marginal zone. A few descendants of C1 can be found three or four cell diameters within the clone of B1. Parasagittal section, dorsal is to the right. Dotted line indicates level of the blastocoel. Fig. 9. At stage 10, the A4 clone (green) has spread over most of the blastocoel roof. The B4 clone (red) extends from the animal cap to the noninvoluting marginal zone (solid arrow), contributing some deep cells to the involuting mesoderm (open arrow). Sagittal section, dorsal is in the lower right corner. Animal pole is marked by line. Fig. 10. At stage 11, the clones of A1 (green) and B1 (red) are distinct in preinvolution mesoderm, but mix as they approach the site of invagination (arrow) and enter the postinvolution mesoderm. Sagittal section, dorsal is to the top. Fig. 11. At stage 10, the B1 clone (red) occupies the dorsal blastopore lip to the site of invagination (arrow). The C1 clone (green) stretches from the yolk plug to the floor of the blastocoel. Figs 12, 13. Two examples of stage 10 embryos in which the B1 clone (red) is the primary contributor to the marginal zone of the dorsal blastopore lip, but members of the C1 clone (green) also make a small contribution. Arrows mark the site of invagination. Orientation is the same as in Fig. 11. Fig. 14. At stage 12.5, the A1 clone (green) has reached the blastopore lip (arrow) and contributes extensively to the entire neural ectoderm. The B1 clone (red) extends throughout the anterior/posterior extent of both neural and mesodermal layers. a, anterior; p, posterior. Sagittal section dorsal is to the top. Fig. 15. At stage 12, the A4 clone (green) and the B4 clone (red) extend across the ventral epidermis from anterior (a) to posterior (p). The ventral blastopore lip is at the upper left edge of the micrograph (arrow). The clones intermix extensively in the posterior half of the deep layer. Sagittal section dorsal is to the top.|
|Fig. 16. Camera-lucida drawings of the positions of 16-cell clones in the vegetal hemisphere during gastrulation. In the top row clones of D2.2, V2.1, D1.2, and V1.1 are shown. In the bottom row clones of D1.1, D2.1, V2.2, and V1.2 are shown. At stage 10 (left), the four vegetal clones are in the positions of the original blastomeres. In addition D1.1 has moved from the animal hemisphere to form the dorsal lip of the blastopore (bottom, stippled). Note that cells from the D2.2 clone form the lateral edge of the lip (top, stippled) and that a few cells from the D2.1 clone are in the dorsal lip (bottom, white). At stage 11 (middle), the two lateral animal clones (D1.2, top; V1.2, bottom) have extended into the vegetal hemisphere and reached the blastopore lip. The D1.1 clone has undergone convergent extension to become a narrow midline stripe. At stage 12 (right), the dorsal clones have narrowed due to convergent extension and the ventral clones have spread over a larger surface area. A small portion of the V1.1 clone (top, black) approaches the ventral lip at the midline, but does not involute.|
|Table 1. Positions of 16- and 32-cell clones in the stage 10 gastrula and their fates The locations of the blastomere clones at stage 10 were described according to the regions defined by Keller (1975, 1976). The fates of these regions (Keller, 1975, 1976) and the observed phenotypes of the blastomeres clones in the tailbud embryo (Moody, 1987a,b) are compared. It should be noted that as a consequence of mixing between clones, as described in the Results section, each clone also makes minor contributions to neighboring regions not listed in this table.|