Dorsal Blastomeres Research Articles

XTcf-3 Transcription Factor Mediates β-Catenin-Induced Axis Formation in Xenopus Embryos

XTcf-3 is a maternally expressed Xenopus homolog of the mammalian HMG box factors Tcf-1 and Lef-1. The N-terminus of XTcf-3 binds to β-catenin. Microinjection of XTcf-3 mRNA in embryos results in nuclear translocation of β-catenin. The β-catenin–XTcf-3 complex activates transcription in a transient reporter gene assay, while XTcf-3 by itself is silent. N-terminal deletion of XTcf-3 (ΔN) abrogates the interaction with β-catenin, as well as the consequent transcription activation. This dominant-negative ΔN mutant suppresses the induction of axis duplication by microinjected β-catenin. It also suppresses endogenous axis specification upon injection into the dorsal blastomeres of a 4-cell-stage embryo. We propose that signaling by β-catenin involves complex formation with XTcf-3, followed by nuclear translocation and activation of specific XTcf-3 target genes.

Anti-Dorsalizing Morphogenetic Protein is a novel TGF-β homolog expressed in the Spemann organizer

We have identified a novel growth factor in Xenopus, which is most closely related to human Bone Morphogenetic Protein-3. Its expression peaks during gastrulation, most prominently in the Spemann organizer, and persists in the posterior neural floor plate and prechordal plate during neurulation. Injection of the corresponding mRNA into dorsal blastomeres results in dose-dependent suppression of dorsal and anterior structures, even in the presence of lithium chloride. Overexpression of the gene downregulates the dorsalizing factors noggin, goosecoid and follistatin, as well as the dorsal markers NCAM, muscle actin and MyoD; conversely, ventral markers are induced. We therefore designate this gene product Anti-Dorsalizing Morphogenetic Protein (ADMP). Though development of dorsoanterior structures is suppressed when exogenous ADMP is injected, the gene is induced by lithium chloride treatment or activin, both of which are known to produce the opposite effect. Thus, the expression of ADMP resembles that of several dorsalizing signals, but its product exerts dorsal-suppressing activity. This suggests that ADMP may moderate organizer-associated dorsalizing influences. These findings are also consistent with the recently advanced proposal of dorsally expressed inhibitory activin-like signals.

Blastomere derivation and domains of gene expression in the Spemann Organizer of Xenopus laevis.

Spemann's Organizer, located in the dorsal marginal zone of the amphibian gastrula, induces and differentiates dorsal axial structures characteristic of this and other vertebrates. To trace the cellular origins of the Xenopus Organizer, we labelled dorsal blastomeres of three of the four tiers (A, B and C) of the 32-cell embryo with green, red and blue fluorescent lineage tracers. A strong vegetalward displacement of labelled clones occurs between the late blastula and early gastrula stages but clones mix only slightly at their borders. The typical early gastrula Organizer is composed of approximately 10% A1 progeny in its animalmost region, 70% B1 progeny in the central region, and 20% C1 progeny in vegetal and deep regions. Variability in the composition of the early gastrula Organizer results from variability in the position of early cleavage planes and in pregastrulation movements. As the Organizer involutes during gastrulation, forming dorsal axial mesoderm, clonal boundaries are greatly dispersed by cell intermixing. Within a clone, deep cells are displaced and intermixed more than superficial cells. Variability in the distribution of progeny in the dorsal axial mesoderm of the late gastrula results mostly from variable intermixing of cells during gastrulation. Experiments to perturb later developmental events by molecular or embryonic manipulations at an early stage must take this variability into account along with the majority distributions of the fate map. Within the early gastrula Organizer, the genes Xbra, goosecoid, noggin and xNR3 are expressed differently in the animal-vegetal and superficial-deep dimensions. In situ hybridization and lineage labelling define distinct regions of the dorsal marginal zone. By the end of gastrulation, dorsal axial mesoderm cells derived from the Organizer have altered their expression of the genes Xbra, goosecoid, noggin and xNR3. At a given stage, a cell's position in the embryo rather than its lineage may be more important in determining which genes it will express.

Modifications of cell fate specification in equal-cleaving nemertean embryos: alternate patterns of spiralian development

The nemerteans belong to a phylum of coelomate worms that display a highly conserved pattern of cell divisions referred to as spiral cleavage. It has recently been shown that the fates of the four embryonic cell quadrants in two species of nemerteans are not homologous to those in other spiralian embryos, such as the annelids and molluscs (Henry, J. Q. and Martindale, M. Q. (1994a) Develop. Genetics 15, 64-78). Equal-cleaving molluscs utilize inductive interactions to establish quadrant-specific cell fates and embryonic symmetry properties following fifth cleavage. In order to elucidate the manner in which cell fates are established in nemertean embryos, we have conducted cell isolation and deletion experiments to examine the developmental potential of the early cleavage blastomeres of two equal-cleaving nemerteans, Nemertopsis bivittata and Cerebratulus lacteus. These two species display different modes of development: N. bivittata develops directly via a non-feeding larvae, while C. lacteus develops to form a feeding pilidium larva which undergoes a radical metamorphosis to give rise to the juvenile worm. By examining the development of certain structures and cell types characteristic of quadrant-specific fates for each of these species, we have shown that isolated blastomeres of the indirect-developing nemertean, C. lacteus, are capable of generating cell fates that are not a consequence of that cell's normal developmental program. For instance, dorsal blastomeres can form muscle fibers when cultured in isolation. In contrast, isolated blastomeres of the direct-developing species, N. bivittata do not regulate their development to the same extent. Some cell fates are specified in a precocious manner in this species, such as those that give rise to the eyes. Thus, these findings indicate that equal-cleaving spiralian embryos can utilize different mechanisms of cell fate and axis specification. The implications of these patterns of nemertean development are discussed in relation to experimental work in other spiralian embryos, and a model is presented that accounts for possible evolutionary changes in cell lineage and the process of cell fate specification amongst these protostome phyla.

Bone morphogenetic protein 2 in the early development of Xenopus laevis

The temporal and spatial transcription patterns of the Xenopus laevis Bone morphogenetic protein 2 (BMP-2) gene have been investigated. Unlike the closely related BMP-4 gene, the BMP-2 gene is strongly transcribed during oogenesis. Besides some enrichment within the animal half, maternal BMP-2 transcripts are ubiquitously distributed in the early cleavage stage embryos but rapidly decline during gastrulation. Zygotic transcription of this gene starts during early neurulation and transcripts are subsequently localized to neural crest cells, olfactory placodes, pineal body and heart anlage. Microinjection of BMP-2 RNA into the two dorsal blastomeres of 4-cell stage embryos leads to ventralization of developing embryos. This coincides with a decrease of transcripts from dorsal marker genes (β-tubulin, α-actin) but not from ventral marker genes (α-globin). BMP-2 overexpression inhibits transcription of the early response gene XFD-1, a fork head/HNF-3 related transcription factor expressed in the dorsal lip, but stimulates transcription of the posterior/ventral marker gene Xhox3, a member of the helix-turn-helix family. Activin A incubated animal caps from BMP-2 RNA injected embryos show transcription of ventral but an inhibition of dorsal marker genes; thus, BMP-2 overrides the dorsalizing activity of activin A. The results demonstrate that BMP-2 overexpression exerts very similar effects as have previously been described for BMP-4, and they suggest that BMP-2 may act already as a maternal factor in ventral mesoderm formation.

Effect of activin and lithium on isolated Xenopus animal blastomeres and response alteration at the midblastula transition

Dorsoventral mesoderm patterning in the amphibian embryo involves a series of interactions mediated by several peptide growth factors. Animal blastomeres isolated at the 8-cell stage are useful for studying mesoderm patterning, since they contain the prospective (uninduced) mesoderm region and allow examination of the default state of animal cells. When activin is applied to these dorsal and ventral animal half explants, a competence prepattern for responding to activin is observed. In order to investigate the characteristics of prepatterning, we treated animal blastomeres with the embryo dorsalizing agent LiCl. Treatment with lithium alone did not induce normal trunk mesoderm in either blastomere. Lithium did, however, alter the competence of animal blastomeres to activin. Dorsal mesoderm was formed in the ventral blastomeres, as well as in the dorsal blastomeres. This result reveals that the early dorsoventral polarity in the animal hemisphere is not fixed. Using goosecoid(gsc) and Xwnt-8 genes as dorsal and ventral mesoderm markers, it was verified that lithium modifies the competence to activin. Unexpectedly, lithium treatment on its own resulted in gsc expression in the animal half explants. This suggests that embryo goosecoid expression may be induced by the effect of dorsal determination activity, but not by mesoderm induction. However, lithium induced also the expression of brachyury (Xbra) gene at very low levels. This would indicate the formation of dorsal-anterior mesoderm, which was not identified by the tissue observations. Expression of Xwnt-8, a ventral mesoderm marker usually induced in blastula animal caps by activin, was hardly induced in the blastomere explants. We isolated whole animal half explants at the 8-cell stage and exposed to activin at different stages. It was found that the same concentration of activin induces gsc before the midblastula stage, and induces Xwnt-8 at later stages. This suggests that the response of animal blastomeres alters depending on the stage of activin signaling.

Linkage of cardiac left-right asymmetry and dorsal-anterior development in Xenopus

The left-right body axis is defined relative to the dorsal-ventral and anterior-posterior body axes. Since left-right asymmetries are not randomly oriented with respect to dorsal-ventral and anterior-posterior spatial patterns, it is possible that a common mechanism determines all three axes in a coordinate manner. Two approaches were undertaken to determine whether alteration in dorsal-anterior development perturbs the left-right orientation of heart looping. Treatments known to decrease dorsal-anterior development in Xenopus laevis, UV irradiation during the first cell cycle or Xwnt-8 DNA injections into dorsal blastomeres, caused an increase in cardiac left-right reversals. The frequency of left-right reversal was correlated with the severity of dorsal-anterior perturbation and with the extent of anterior notochord regression. Injection of Xwnt-8 DNA into dorsal midline cells resulted in decreased dorsal-anterior development and a correlated increase in cardiac left-right reversals. In contrast, injection of Xwnt-8 DNA into cardiac progenitor blastomeres did not result in left-right reversals, and dorsal-anterior development and notochord formation were normal. Disrupting development of dorsal-anterior cells, including cells that give rise to the Organizer region and the notochord, results in the randomization of cardiac left-right asymmetry. These results suggest dorsal-anterior development and the regulation of left-right orientation are linked.

Differential perturbations in the morphogenesis of anterior structures induced by overexpression of truncated XB- and N-cadherins in Xenopus embryos.

Cadherins, a family of Ca-dependent adhesion molecules, have been proposed to act as regulators of morphogenetic processes and to be major effectors in the maintenance of tissue integrity. In this study, we have compared the effects of the expression of two truncated cadherins during early neurogenesis in Xenopus laevis. mRNA encoding deleted forms of XB- and N-cadherin lacking most of the extracellular domain were injected into the four animal dorsal blastomeres of 32-cell stage Xenopus embryos. These truncated cadherins altered the cohesion of cells derived from the injected blastomeres and induced morphogenetic defects in the anterior neural tissue to which they chiefly contributed. Truncated XB-cadherin was more efficient than N-cadherin in inducing these perturbations. Moreover, the coexpression of both truncated cadherins had additive perturbation effects on neural development. The two truncated cadherins can interact with the three known catenins, but with distinct affinities. These results suggest that the adhesive signal mediated by cadherins can be perturbed by overexpressing their cytoplasmic domains by competing with different affinity with catenins and/or a common anchor structure. Therefore, the correct regulation of cadherin function through the cytoplasmic domain appears to be a crucial step in the formation of the neural tissue.

Progressive determination of cell fates along the dorsoventral axis in the sea urchin Heliocidaris erythrogramma.

In the direct-developing sea urchin Heliocidaris erythrogramma the first cleavage division bisects the dorsoventral axis of the developing embryo along a frontal plane. In the two-celled embryo one of the blastomeres, the ventral cell (V), gives rise to all pigmented mesenchyme, as well as to the vestibule of the echinus rudiment. Upon isolation, however, the dorsal blastomere (D) displays some regulation, and is able to form a small number of pigmented mesenchyme cells and even a vestibule. We have examined the spatial and temporal determination of cell fates along the dorsoventral axis during subsequent development. We demonstrate that the dorsoventral axis is resident within both cells of the two-celled embryo, but only the ventral pole of this axis has a rigidly fixed identity this early in development. The polarity of this axis remains the same in half-embryos developing from isolated ventral (V) blastomeres, but it can flip 180° in half-embryos developing from isolated dorsal (D) blastomeres. We find that cell fates are progressively determined along the dorsoventral axis up to the time of gastrulation. The ability of dorsal half-embryos to differentiate ventral cell fates diminishes as they are isolated at progressively later stages of development. These results suggest that the determination of cell fates along the dorsoventral axis in H. erythrogramma is regulated via inductive interactions organized by cells within the ventral half of the embryo.

Progressive determination of cell fates along the dorsoventral axis in the sea urchin Heliocidaris erythrogramma

In the direct-developing sea urchinHeliocidaris erythrogramma the first cleavage division bisects the dorsoventral axis of the developing embryo along a frontal plane. In the two-celled embryo one of the blastomeres, the ventral cell (V), gives rise to all pigmented mesenchyme, as well as to the vestibule of the echinus rudiment. Upon isolation, however, the dorsal blastomere (D) displays some regulation, and is able to form a small number of pigmented mesenchyme cells and even a vestibule. We have examined the spatial and temporal determination of cell fates along the dorsoventral axis during subsequent development. We demonstrate that the dorsoventral axis is resident within both cells of the two-celled embryo, but only the ventral pole of this axis has a rigidly fixed identity this early in development. The polarity of this axis remains the same in half-embryos developing from isolated ventral (V) blastomeres, but it can flip 180° in half-embryos developing from isolated dorsal (D) blastomeres. We find that cell fates are progressively determined along the dorsoventral axis up to the time of gastrulation. The ability of dorsal half-embryos to differentiate ventral cell fates diminishes as they are isolated at progressively later stages of development. These results suggest that the determination of cell fates along the dorsoventral axis inH. erythrogramma is regulated via inductive interactions organized by cells within the ventral half of the embryo.

Competence Prepattern in the Animal Hemisphere of the 8-Cell-Stage Xenopus Embryo

Activins are known to be potentially important regulators in Xenopus developmental processes. It has been shown that activins exist maternally in the egg and can induce mesodermal tissues in blastula animal cap explants. However, the blastula ectoderm is known to possess a predisposed local response pattern to activin, and the process of the prepatterning is not understood. We isolated animal hemispheres from late 8-cell-stage embryos and treated them briefly with activin A. Expression of the muscle-specific actin gene was induced after a 30-min activin treatment, even when it was followed by treatment with follistatin, an activin-specific binding protein. This suggests that the animal-half blastomeres become competent to activin A before the 16-cell stage. In the normal embryo, the 8-cell stage animal dorsal blastomeres populate neural ectoderm and most of the dorsal lip of the gastrula blastopore, the region of Spemann's organizer, and are the major progenitor for dorsal mesodermal tissues. When the dorsal and ventral animal-half blastomeres of the 8-cell stage were isolated and treated with activin independently, significant differences in tissue differentiation were observed. Dorsal blastomeres gave rise to trunk and tail structures containing dorsal mesoderm, whereas the ventral blastomere explants formed spheres containing solely ventral mesoderm. Further, both muscle actin transcription and goosecoid transcription were induced primarily in dorsal blastomeres. Our results suggest that a competence prepattern of response to activin exists as early as the 8-cell stage.

The retinal fate of Xenopus cleavage stage progenitors is dependent upon blastomere position and competence: studies of normal and regulated clones

The clonal origin of the stage 43-44 Xenopus retina from cleavage stage precursors was quantitatively assessed with lineage tracing techniques. The retina descends from a specific subset of those blastomeres that form forebrain. The most animal dorsal midline cell (D1.1.1) produced about half of the retina, the three other dorsal ipsilateral blastomeres each produce about an eighth of the retina, and the four contralateral dorsal blastomeres and an ipsilateral ventral-animal cell together produce the remaining eighth of the retina. There was no significant spatial segregation of the clones derived from different progenitors in either the anterior-posterior or dorsal-ventral axes of the retina and no boundaries between clones were observed. Instead, the clones intermixed to form multiple radial subclones that were equivalent to those demonstrated by marking optic vesicle progenitor cells (Holt et al., 1988; Wetts and Fraser, 1988). This mosaic pattern was initiated by the beginning of gastrulation, advanced in the neural plate, and virtually complete in the optic vesicle. At optic vesicle stages cell movement within subclones was restricted, resulting in the formation of lineally related columns of cells in the mature retina. To determine if the blastomere progenitors are determined to produce these retinal lineage patterns, the major retinal progenitor (D1.1.1) was deleted bilaterally. About 60% of the tadpoles developed normal-appearing eyes; of these the retinas in two-thirds were normal in size and the rest were smaller. The blastomeres surrounding the deleted D1.1.1 progenitors changed their contributions to retina in different ways to effect a complete or partial restoration. Ventral blastomeres, which normally contribute mainly to the tail, produced substantial amounts of the retina while dorsal blastomeres, which normally contribute mainly to the head, decreased their contribution to the retina. To determine whether these changes in retinal lineage were due to changes in blastomere position after the surgery, various other blastomeres were deleted prior to lineage mapping. Dorsal-animal blastomeres took over the retinal fate of their dorsal-vegetal neighbors after those neighbors were deleted, but did not change fate after the deletion of their ventral-animal neighbors. This result suggests that dorsal-animal blastomeres change positional values in only one direction (dorsal to vegetal) after neighbor cell deletion, and that retinal fate is dictated by blastomere position. To test this hypothesis directly, different ventral and vegetal blastomeres, which normally do not produce retina, were transplanted to the position of D1.1.1.(ABSTRACT TRUNCATED AT 400 WORDS)

The homeobox gene goosecoid controls cell migration in Xenopus embryos

goosecold (gsc), a homeobox gene expressed specifically in the dorsal blastopore lip of the Xenopus gastrula, is considered to play an important role in Spemann's organizer phenomenon. Lineage tracing and time-lapse microscopy were used to follow the fate of embryonic cells microinjected with gsc mRNA. Microinjected gsc has non-cell autonomous effects, recruiting neighboring uninjected cells into a twinned dorsal axis. Ectopic expression of gsc mRNA in ventral blastomeres as well as overexpression of gsc in dorsal blastomeres leads to cell movement toward the anterior of the embryo. The results suggest a function for gsc in the control of gastrulation movements in groups of cells, but not in dissociated cells, and demonstrate that a vertebrate homeobox gene can regulate region-specific cell migration.

Autonomous differentiation of dorsal axial structures from an animal cap cleavage stage blastomere in Xenopus

Dorsal or ventral blastomeres of the 16- and 32-cell stage animal hemisphere were labeled with a lineage dye and transplanted into the position of a ventral, vegetal midline blastomere. The donor blastomeres normally give rise to substantial amounts of head structures and central nervous system, whereas the blastomere which they replaced normally gives rise to trunk mesoderm and endoderm. The clones derived from the transplanted ventral blastomeres were found in tissues appropriate for their new position, whereas those derived from the transplanted dorsal blastomeres were found in tissues appropriate for their original position. The transplanted dorsal clones usually migrated into the host's primary axis (D1.1, 92%; D1.1.1, 69%; D1.1.2, 100%), and in many cases they also induced and populated a secondary axis (D1.1, 43%; D1.1.1, 67%; D1.1.2, 63%). Bilateral deletion of the dorsal blastomeres resulted in partial deficits of dorsal axial structures in the majority of cases, whereas deletions of ventral midline blastomeres did not. When the dorsal blastomeres were cultured as explants they elongated. Notochord and cement glands frequently differentiated in these explants. These studies show that the progeny of the dorsal, midline, animal blastomeres: (1) follow their normal lineage program to populate dorsal axial structures after the blastomere is transplanted to the opposite pole of the embryo; (2) induce and contribute to a secondary axis from their transplanted position in many embryos; (3) are important for the normal formation of the entire length of the dorsal axis; and (4) autonomously differentiate in the absence of exogenous growth factor signals. These data indicate that by the 16-cell stage, these blastomeres have received instructions regarding their fate, and they are intrinsically capable of carrying out some of their developmental program.

Segregation of fate during cleavage of frog (Xenopus laevis) blastomeres

A detailed fate map of all the progeny derived from each of the blastomeres of the 4- and 8-cell stage South African clawed frog (Xenopus laevis) embryo is presented. Each "identified" blastomere that results from stereotypic cleavages has a characteristic set of progeny that distinguishes it from the other blastomeres of the embryo. The 4-cell dorsal (D) blastomere is the major progenitor of the stomodeum, cement gland, retina, notochord, head somite, pharynx and liver. The 4-cell ventral (V) blastomere is the major progenitor of the trunk and fin epidermis, ventral somite, nephrotome, lateral plate mesoderm and proctodeum. The other organs are derived from both blastomeres. At the next cell division, the animal hemisphere daughters of both blastomeres (D1 and V1, respectively) become the major progenitors for head ectodermal and mesodermal structures, and the vegetal hemisphere daughters become the major progenitors for trunk mesodermal (D2) or trunk endodermal (V2) structures. Semiquantitative lineage diagrams, using data from this and from previous studies demonstrate that as cleavage proceeds from the 2- to the 32-cell stage, the progenitors for particular organs or for specific regions of organs segregate into defined regions of the blastula. To determine whether this segregation is related to the position of the blastomere or to its geneological lineage, we compared the fates of radial 8-cell blastomeres to those of stereotypic 8-cell blastomeres. Radial blastomeres have fates nearly equivalent to the sum of the two 16-cell blastomeres that occupy the same position in the embryo, demonstrating that fate depends upon blastomere position rather than lineage.

Lithium induces dorsal-type migration of mesodermal cells in the entire marginal zone of urodele amphibian embryos.

The effect of lithium (Li+) on gastrulation movements was investigated during the development of the urodele amphibianPleurodeles waltl. Attention was focused on mesodermal cell migration. Under conditions of Li+ treatment providing a maximal enhancement of dorsoanterior structures, it was found that the dorsoventral polarity of gastrulation was abolished. In particular, vital staining and scanning electron microscopy observations on embryo fractures showed that mesodermal cells migrated radially after Li+ treatment, which led to the formation of rounded embryos. Epiboly movements thus were accelerated. Nevertheless, contrasting with the precocious disappearance of the early-formed yolk plug, archenteron invagination was constantly retarded and commenced with a delay of several hours as compared to control gastrulae. Cell-lineage analysis of the progenies from ventral or dorsal equatorial blastomeres of 32-cell-stage embryos provided evidence that both dorsal and ventral mesoderm contributed to notochordal tissue after Li+ treatment. Dorsalization of the entire marginal zone was confirmed by the ability of the entire mesoderm rudiment to behave as a dorsal organiser after Li+ treatment. Comparison of the migratory behaviour of isolated animal hemispheres from Li+-treated or control embryos cultured on fibronectin-coated substrate indicated that all marginal cells acquired the autonomous capacity for migration of dorsal marginal cells under the action of lithium.

Cell-cell interactions determine the dorsoventral axis in embryos of an equally cleaving opisthobranch mollusc.

Dorsoventral polarity in molluscan embryos can arise by two distinct mechanisms, where the mechanism employed is strongly correlated with the cleavage pattern of the early embryo. In species with unequal cleavage, the dorsal lineage, or "D quadrant", is determined in a cell-autonomous manner by the inheritance of cytoplasmic determinants. However, in gastropod molluscs with equal cleavage, cell-cell interactions are required to specify the fate of the dorsal blastomere. During the fifth cleavage interval in equally cleaving embryos, one of the vegetal macromeres makes exclusive contacts with the animal micromeres, and this macromere will give rise to the mesodermal precursor cell at the next division, thereby identifying the dorsal quadrant. This study examines D-quadrant determination in an equally cleaving species from a group of previously uninvestigated gastropods, the subclass Opisthobranchia. Blastomere ablation experiments were performed on embryos of Haminoea callidegenita to (i) determine the developmental potential of macromeres before and after fifth cleavage, and (ii) examine the role of micromere-macromere interactions in the establishment of bilateral symmetry. The results suggest that the macromeres are developmentally equivalent prior to fifth cleavage, but become nonequivalent soon afterward. The dorsoventral axis corresponds to the displacement of the micromeres over one macromere early in the fifth cleavage interval. This unusual cellular topology is hypothesized to result from constraints imposed on micromere-macromere interactions in an embryo that develops from a large egg and forms a stereoblastula (no cleavage cavity). Ablation of the entire first quarter of micromeres results in embryos which remain radially symmetrical in the vegetal hemisphere, indicating that micromere-macromere interactions are required for the elaboration of bilateral symmetry properties. Therefore, inductive interactions between cells may represent a general strategy for dorsoventral axis determination in equally cleaving gastropods.

Dorsal Blastomeres in the Equatorial Region of the 32-Cell Xenopus Embryo Autonomously Produce Progeny Committed to the Organizer: (Xenopus/32-cell embryo/blastomere transplantation/determinant localization/head organizer).

For testing the autonomic differentiation abilities of dorsal equatorial blastomeres of 32-cell Xenopus embryos, their roles in head formation in normal development and the organizer-inducing capabilities of the dorsal-most vegetal cells, interspecific transplantations were made using Xenopus borealis and X. laevis. When transplanted into the ventral region, the dorsal blastomeres produced descendants that differentiated into prechordal mesoderm, notochord and somites in the recipient according to their fates. They induced formation of the secondary embryo with the head and tail. The prechordal mesoderm and notochord in the secondary structure consisted of progeny of the graft, whereas somites and the CNS were chimeric and the pronephros was composed of host cells. Replacement of the dorsal blastomeres by ventral equatorial cells caused complete arrest of head formation in the recipient. Without exception, the notochord was completely absent or very thin. These results confirm the assumption that the presumptive head organizer in the Xenopus embryo is localized in the dorsal equatorial region at the 32-cell stage and comes into existence not under the inductive influence of the dorsal-most vegetal cells, but owing to allocation of morphogenetic determinants residing in the fertilized egg to the dorsal equatorial blastomeres of the 32-cell embryo.

Regional differences of proteins in isolated cells of early embryos of Xenopus laevis

Regional differences of proteins were studied by two-dimensional gel electrophoresis in early embryos of Xenopus laevis. Pairs of blastomeres on the dorso-ventral axis were isolated from 16- and 32-cell embryos. Some dorso-ventral differences have been detected at 32-cell embryos. The proteins which were clearly detectable in the vegetal cells of the ventral marginal zone were only faintly detectable or undetectable in those of the dorsal marginal zone, and a regionally specific spot was detected in dorsal blastomeres.

DIFFERENTIAL INITIATION OF rRNA GENE ACTIVITY IN PROGENIES OF DIFFERENT BLASTOMERES OF EARLY XENOPUS EMBRYOS: EVIDENCE FOR REGULATED SYNTHESIS OF rRNA.

The dorsal and ventral blastomeres of 4-celled embryos of Xenopus luevis were separated. During the next 14 hr in cultnre, the cell numbers of the progeny cell aggregates of the dorsal and ventral blastomeres respectively, were the same. Synthesis of 4 S RNA started after about 4 hr of culture in both kinds of progeny cell aggregates. However, a clear-cut difference was found in the time when rRNA synthesis started in these aggregates: it began after 10 hr in cell aggregates derived from the dorsal blastomeres, and after about 14 hr in those derived from the ventral blastomeres. After this, rRNA synthesis became more and more active in both types of aggregates. This is the first demonstration of differential initiation of rRNA synthesis in aggregates derived from different blastomeres but containing the same number of cells. The results provide unequivocal evidence for developmental regulation of rRNA synthesis.