The allocation of early blastomeres to the ectoderm and endoderm is variable in the sea urchin embryo

Development ◽  
1997 ◽  
Vol 124 (11) ◽  
pp. 2213-2223 ◽  
Author(s):  
C.Y. Logan ◽  
D.R. McClay
Keyword(s):  
Sea Urchin ◽  
Low Frequency ◽  
Initial Step ◽  
Cell Types ◽  
Lytechinus Variegatus ◽  
Cell Fates ◽  
Cell Stage ◽  
Sea Urchin Embryo

During sea urchin development, a tier-to-tier progression of cell signaling events is thought to segregate the early blastomeres to five different cell lineages by the 60-cell stage (E. H. Davidson, 1989, Development 105, 421–445). For example, the sixth equatorial cleavage produces two tiers of sister cells called ‘veg1′ and ‘veg2,’ which were projected by early studies to be allocated to the ectoderm and endoderm, respectively. Recent in vitro studies have proposed that the segregation of veg1 and veg2 cells to distinct fates involves signaling between the veg1 and veg2 tiers (O. Khaner and F. Wilt, 1991, Development 112, 881–890). However, fate-mapping studies on 60-cell stage embryos have not been performed with modern lineage tracers, and cell interactions between veg1 and veg2 cells have not been shown in vivo. Therefore, as an initial step towards examining how archenteron precursors are specified, a clonal analysis of veg1 and veg2 cells was performed using the lipophilic dye, DiI(C16), in the sea urchin species, Lytechinus variegatus. Both veg1 and veg2 descendants form archenteron tissues, revealing that the ectoderm and endoderm are not segregated at the sixth cleavage. Also, this division does not demarcate cell type boundaries within the endoderm, because both veg1 and veg2 descendants make an overlapping range of endodermal cell types. The allocation of veg1 cells to ectoderm and endoderm during cleavage is variable, as revealed by both the failure of veg1 descendants labeled at the eighth equatorial division to segregate predictably to either tissue and the large differences in the numbers of veg1 descendants that contribute to the ectoderm. Furthermore, DiI-labeled mesomeres of 32-cell stage embryos also contribute to the endoderm at a low frequency. These results show that the prospective archenteron is produced by a larger population of cleavage-stage blastomeres than believed previously. The segregation of veg1 cells to the ectoderm and endoderm occurs relatively late during development and is unpredictable, indicating that later cell position is more important than the early cleavage pattern in determining ectodermal and archenteron cell fates.

Cell-cell interactions regulate skeleton formation in the sea urchin embryo

Development ◽  
1993 ◽  
Vol 119 (3) ◽  
pp. 833-840 ◽  
Author(s):  
N. Armstrong ◽  
J. Hardin ◽  
D.R. McClay
Keyword(s):  
Sea Urchin ◽  
Cell Interactions ◽  
Sea Urchin Embryo ◽  
Skeletal Pattern ◽  
Tissue Sensitivity ◽  
Mesenchyme Cells ◽  
Larval Skeleton ◽  
Skeleton Formation

In the sea urchin embryo, the primary mesenchyme cells (PMCs) make extensive contact with the ectoderm of the blastula wall. This contact is shown to influence production of the larval skeleton by the PMCs. A previous observation showed that treatment of embryos with NiCl2 can alter spicule number and skeletal pattern (Hardin et al. (1992) Development, 116, 671–685). Here, to explore the tissue sensitivity to NiCl2, experiments recombined normal or NiCl2-treated PMCs with either normal or NiCl2-treated PMC-less host embryos. We find that NiCl2 alters skeleton production by influencing the ectoderm of the blastula wall with which the PMCs interact. The ectoderm is responsible for specifying the number of spicules made by the PMCs. In addition, experiments examining skeleton production in vitro and in half- and quarter-sized embryos shows that cell interactions also influence skeleton size. PMCs grown in vitro away from interactions with the rest of the embryo, can produce larger spicules than in vivo. Thus, the epithelium of the blastula wall appears to provide spatial and scalar information that regulates skeleton production by the PMCs.

scholarly journals β-Catenin is essential for patterning the maternally specified animal-vegetal axis in the sea urchin embryo

1998 ◽  
Vol 95 (16) ◽  
pp. 9343-9348 ◽  
Author(s):  
Athula H. Wikramanayake ◽  
Ling Huang ◽  
William H. Klein
Keyword(s):  
Sea Urchin ◽  
Molecular Mechanisms ◽  
Early Embryo ◽  
Cell Fates ◽  
Cell Stage ◽  
Signal Transduction Cascade ◽  
Sea Urchin Embryos ◽  
Sea Urchin Embryo ◽  
Vegetal Pole ◽  
Vegetal Cell

In sea urchin embryos, the animal-vegetal axis is specified during oogenesis. After fertilization, this axis is patterned to produce five distinct territories by the 60-cell stage. Territorial specification is thought to occur by a signal transduction cascade that is initiated by the large micromeres located at the vegetal pole. The molecular mechanisms that mediate the specification events along the animal–vegetal axis in sea urchin embryos are largely unknown. Nuclear β-catenin is seen in vegetal cells of the early embryo, suggesting that this protein plays a role in specifying vegetal cell fates. Here, we test this hypothesis and show that β-catenin is necessary for vegetal plate specification and is also sufficient for endoderm formation. In addition, we show that β-catenin has pronounced effects on animal blastomeres and is critical for specification of aboral ectoderm and for ectoderm patterning, presumably via a noncell-autonomous mechanism. These results support a model in which a Wnt-like signal released by vegetal cells patterns the early embryo along the animal–vegetal axis. Our results also reveal similarities between the sea urchin animal–vegetal axis and the vertebrate dorsal–ventral axis, suggesting that these axes share a common evolutionary origin.

Analysis of clonal restriction of cell mingling in Xenopus

1985 ◽  
Vol 312 (1153) ◽  
pp. 57-65 ◽  
Keyword(s):  
Horseradish Peroxidase ◽  
Cell Types ◽  
Cell Group ◽  
Cell Stage ◽  
Wide Range ◽  
Cell Groups ◽  
Cell Layers ◽  
Single Blastomeres

Cell fates were traced by injecting horseradish peroxidase into single blastomeres of Xenopus embryos at 2- to 512-cell stages. At later stages the number, types and locations of all labelled progeny were observed. Progeny of a single labelled ancestral cell divided coherently until the 12th cell generation, the onset of gastrulation, and then dispersed and mingled with unlabelled cells. Cell mingling was restricted at mediolateral and anterior—posterior boundaries. These boundaries were always respected by progeny of any blastomere labelled at the 512-cell stage but they were frequently crossed by progeny of blastomeres labelled at the 256-cell or earlier stages. The boundaries defined seven morphological compartments each populated exclusively by a group of ancestral cells at the 512-cell stage. Each blastomere that contributed progeny to the nervous system also gave rise to a wide range of cell types in all three primary germ cell layers but the clone was restricted to a single compartment. Analysis of clonal restriction of cell mingling was done in vitro . Twenty to thirty blastomeres were excised from one ancestral cell group at the 512-cell stage and combined in vitro with 20-30 blastomeres from another group. One group of blastomeres labelled with horseradish peroxidase was placed in contact with another group of unlabelled blastomeres, maintained in vitro for up to 2 days, and then processed histologically to show the distribution of labelled and unlabelled cells. Mingling was significantly greater in combinations of two of the same ancestral cell groups than in combinations of two different ancestral cell groups. A similar result was observed when a single labelled cell was combined with either the same or different ancestral cells. In all experiments the cells were significantly larger in combinations of different ancestral cell groups, indicating that they had undergone fewer divisions. These results are consistent with the hypothesis that boundaries observed in vivo are lines of clonal restriction formed by mutual inhibition of cell motility and cell division following contact between progeny of different ancestral cell groups.

scholarly journals Proteolysis of the major yolk glycoproteins is regulated by acidification of the yolk platelets in sea urchin embryos

1992 ◽  
Vol 117 (6) ◽  
pp. 1211-1221 ◽  
Author(s):  
SK Mallya ◽  
JS Partin ◽  
MC Valdizan ◽  
WJ Lennarz
Keyword(s):  
Early Development ◽  
Sea Urchin ◽  
Ph Value ◽  
Total Protein Content ◽  
Developmentally Regulated ◽  
Cell Stage ◽  
Ph Optimum ◽  
Sea Urchin Embryos

The precise function of the yolk platelets of sea urchin embryos during early development is unknown. We have shown previously that the chemical composition of the yolk platelets remains unchanged in terms of phospholipid, triglyceride, hexose, sialic acid, RNA, and total protein content after fertilization and early development. However, the platelet is not entirely static because the major 160-kD yolk glycoprotein YP-160 undergoes limited, step-wise proteolytic cleavage during early development. Based on previous studies by us and others, it has been postulated that yolk platelets become acidified during development, leading to the activation of a cathepsin B-like yolk proteinase that is believed to be responsible for the degradation of the major yolk glycoprotein. To investigate this possibility, we studied the effect of addition of chloroquine, which prevents acidification of lysosomes. Consistent with the postulated requirement for acidification, it was found that chloroquine blocked YP-160 breakdown but had no effect on embryonic development. To directly test the possibility that acidification of the yolk platelets over the course of development temporally correlated with YP-160 proteolysis, we added 3-(2,4-dinitroanilo)-3-amino-N-methyldipropylamine (DAMP) to eggs or embryos. This compound localizes to acidic organelles and can be detected in these organelles by EM. The results of these studies revealed that yolk platelets did, in fact, become transiently acidified during development. This acidification occurred at the same time as yolk protein proteolysis, i.e., at 6 h after fertilization (64-cell stage) in Strongylocentrotus purpuratus and at 48 h after fertilization (late gastrula) in L. pictus. Furthermore, the pH value at the point of maximal acidification of the yolk platelets in vivo was equal to the pH optimum of the enzyme measured in vitro, indicating that this acidification is sufficient to activate the enzyme. For both S. purpuratus and Lytechinus pictus, the observed decrease in the pH was approximately 0.8 U, from 7.0 to 6.2. The trypsin inhibitor benzamidine was found to inhibit the yolk proteinase in vivo. By virtue of the fact that this inhibitor was reversible we established that the activity of the yolk proteinase is developmentally regulated even though the enzyme is present throughout the course of development. These findings indicate that acidification of yolk platelets is a developmentally regulated process that is a prerequisite to initiation of the catabolism of the major yolk glycoprotein.

Altering cell fates in sea urchin embryos by overexpressing SpOtx, an orthodenticle-related protein

Development ◽  
1996 ◽  
Vol 122 (5) ◽  
pp. 1489-1498 ◽  
Author(s):  
C.A. Mao ◽  
A.H. Wikramanayake ◽  
L. Gan ◽  
C.K. Chuang ◽  
R.G. Summers ◽  
...  
Keyword(s):  
Cell Fate ◽  
Sea Urchin ◽  
Biological Effects ◽  
Gene Activation ◽  
Related Protein ◽  
Cell Fates ◽  
Cell Stage ◽  
Morphological Alterations ◽  
Transactivation Domains ◽  
Sea Urchin Embryo

While many general features of cell fate specification in the sea urchin embryo are understood, specific factors associated with these events remain unidentified. SpOtx, an orthodenticle-related protein, has been implicated as a transcriptional activator of the aboral ectoderm-specific Spec2a gene. Here, we present evidence that SpOtx has the potential to alter cell fates. SpOtx was found in the cytoplasm of early cleavage stage embryos and was translocated into nuclei between the 60- and 120-cell stage, coincident with Spec gene activation. Eggs injected with SpOtx mRNA developed into epithelial balls of aboral ectoderm suggesting that SpOtx redirected nonaboral ectoderm cells to an aboral ectoderm fate. At least three distinct domains on SpOtx, the homeobox and regions in the N-terminal and C-terminal halves of the protein, were required for the morphological alterations. These same N-terminal and C-terminal regions were shown to be transactivation domains in a yeast transactivation assay, indicating that the biological effects of overexpressing SpOtx were due to its action as a transcription factor. Our results suggest that SpOtx is involved in aboral ectoderm differentiation by activating aboral ectoderm-specific genes and that modulating its expression can lead to changes in cell fate.

SpSoxB1, a maternally encoded transcription factor asymmetrically distributed among early sea urchin blastomeres

Development ◽  
1999 ◽  
Vol 126 (23) ◽  
pp. 5473-5483 ◽  
Author(s):  
A.P. Kenny ◽  
D. Kozlowski ◽  
D.W. Oleksyn ◽  
L.M. Angerer ◽  
R.C. Angerer
Keyword(s):  
Transcription Factor ◽  
Sea Urchin ◽  
Protein Distribution ◽  
Blastula Stage ◽  
Cell Stage ◽  
Mrna Accumulation ◽  
Cis Element ◽  
Sea Urchin Embryo ◽  
Asymmetrically Distributed

We have identified a Sox family transcription factor, SpSoxB1, that is asymmetrically distributed among blastomeres of the sea urchin embryo during cleavage, beginning at 4th cleavage. SpSoxB1 interacts with a cis element that is essential for transcription of SpAN, a gene that is activated cell autonomously and expressed asymmetrically along the animal-vegetal axis. In vitro translated SpSoxB1 forms a specific complex with this cis element whose mobility is identical to that formed by a protein in nuclear extracts. An anti-SpSoxB1 rabbit polyclonal antiserum specifically supershifts this DNA-protein complex and recognizes a single protein on immunoblots of nuclear proteins that comigrates with in vitro translated SpSoxB1. Developmental immunoblots of total proteins at selected early developmental stages, as well as EMSA of egg and 16-cell stage proteins, show that SpSoxB1 is present at low levels in unfertilized eggs and progressively accumulates during cleavage. SpSoxB1 maternal transcripts are uniformly distributed in the unfertilized egg and the protein accumulates to similar, high concentrations in all nuclei of 4- and 8-cell embryos. However, at fourth cleavage, the micromeres, which are partitioned by asymmetric division of the vegetal 4 blastomeres, have reduced nuclear levels of the protein, while high levels persist in their sister macromeres and in the mesomeres. During cleavage, the uniform maternal SpSoxB1 transcript distribution is replaced by a zygotic nonvegetal pattern that reinforces the asymmetric SpSoxB1 protein distribution and reflects the corresponding domain of SpAN mRNA accumulation at early blastula stage (approximately 150 cells). The vegetal region lacking nuclear SpSoxB1 gradually expands so that, after blastula stage, only cells in differentiating ectoderm accumulate this protein in their nuclei. The results reported here support a model in which SpSoxB1 is a major regulator of the initial phase of asymmetric transcription of SpAN in the nonvegetal domain by virtue of its distribution at 4th cleavage and is subsequently an important spatial determinant of expression in the early blastula. This factor is the earliest known spatially restricted regulator of transcription along the animal-vegetal axis of the sea urchin embryo.

scholarly journals ACTIVE SUPPRESSION AS A POSSIBLE MECHANISM OF TOLERANCE IN TETRAPARENTAL MICE

1973 ◽  
Vol 137 (2) ◽  
pp. 291-300 ◽  
Author(s):  
S. Michael Phillips ◽  
Thomas G. Wegmann
Keyword(s):  
Experimental System ◽  
Cell Types ◽  
Parental Cell ◽  
Spleen Cells ◽  
Cell Stage ◽  
Self Tolerance ◽  
Blocking Antibodies ◽  
Mixed Lymphocyte Reactions

Previous work has indicated that tetraparental mice, chimeric since the eight-cell stage because of embryo fusion using histoincompatible strain combinations, possess autospecific immune cells and blocking antibodies. Although this phenomenon has been demonstrated in vitro, it may have relevance to the self-tolerance shown by these mice in vivo. The experiments described here indicate that spleen cells from tetraparental mice can block mixed lymphocyte reactions between the two parental cell types, but not between unrelated strains. Furthermore, this suppressive ability is not affected by an otherwise effective treatment of the tetraparental spleen cells with anti-θ antibody and complement. The in vitro experimental system elaborated here should help to characterize the cell type responsible for the suppression.

2 THE IN VIVO DEVELOPMENTAL POTENTIAL OF PORCINE SKIN-DERIVED PROGENITORS

2012 ◽  
Vol 24 (1) ◽  
pp. 112 ◽  
Author(s):  
M. T. Zhao ◽  
X. Yang ◽  
K. Lee ◽  
J. Mao ◽  
J. M. Teson ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Lineage ◽  
Pcr Amplification ◽  
Cell Types ◽  
Blastocyst Stage ◽  
Cell Stage ◽  
Germ Layers ◽  
Developmental Potential

Skin-derived progenitors (SKP) are capable of generating both neural and mesodermal progeny in vitro: neurons, Schwann cells, adipocytes, osteocytes and chondrocytes, thus exhibiting characteristics similar to embryonic neural crest stem cells. SKP show distinct transcriptional profiles when compared with neurospheres/neural stem cells in the central nervous system (CNS) and skin-derived fibroblasts, indicating a novel type of multipotent stem cell derived from the dermis of the skin. However, it remains unclear whether SKP cells can produce ectoderm and mesoderm lineages or other germ layers in vivo, although oocyte-like structures can be induced from porcine SKP in vitro. Embryonic chimeras are a well-established tool for investigating cell lineage determination and cell potency through normal embryonic development. Thus the purpose of this study was to investigate the in vivo developmental potential of porcine SKP by chimera production. Porcine SKP cells and fibroblasts were isolated from the back skin of Day 35 to 50 GFP transgenic fetuses. Individual cells or clusters of male GFP transgenic SKP and skin-derived GFP-expressing fibroblasts were injected into pre-compact in vitro-fertilized (IVF) embryos, respectively and then transferred into corresponding surrogates 24 h post-injection. Additional injected embryos were cultured in PZM3 medium for another 2 days until the blastocyst stage and subsequently stained with Hoechst 33342. Interestingly, in some of the chimeras the injected SKP cells migrated and dispersed into different locations of the host blastocysts, whereas in others they remained as a cluster of cells within the chimeric blastocysts. In contrast, the fibroblast cells were not observed to spread around the host blastocysts. Two chimeric fetuses were recovered at the middle of gestation and a litter of viable piglets was born. Genomic DNA was extracted from various tissues of chimeric piglets and subjected to PCR amplification. Two chimeric fetuses and 2 out of 6 piglets carried the GFP transgene in SKP-derived chimeras, but GFP was not present in the fibroblast-derived chimeric fetuses (n = 6). Surprisingly, the GFP transgene was present in various tissues of two SKP-derived chimeric piglets, including lung, heart, liver, artery, kidney, brain, skin, muscle, gut, ovary, pancreas and stomach, thus representing the 3 germ layers (ectoderm, mesoderm and endoderm). In addition, SRY was detected in several tissues of the two GFP-positive female chimeric piglets, confirming the chimerism of these piglets. Therefore, it appears that porcine SKP can contribute to various cell types of the 3 germ layers and have a broader developmental potency than previously expected. Alternatively, pre-compact (4-cell and 8-cell stage) embryos may provide a unique environment for reprogramming skin-derived progenitors into a more primitive state by the process of embryonic compaction. This study was funded by NIH National Center for Research Resources (R01RR013438) and Food for the 21st Century at the University of Missouri.

A fate map of the vegetal plate of the sea urchin (Lytechinus variegatus) mesenchyme blastula

Development ◽  
1996 ◽  
Vol 122 (1) ◽  
pp. 253-263 ◽  
Author(s):  
S.W. Ruffins ◽  
C.A. Ettensohn
Keyword(s):  
Sea Urchin ◽  
Single Cells ◽  
Cell Types ◽  
Lineage Tracing ◽  
Lytechinus Variegatus ◽  
Fate Map ◽  
Blastula Stage ◽  
Cell Stage ◽  
Mesodermal Cell ◽  
Single Blastomeres

Previous lineage tracing experiments have shown that the vegetal blastomers of cleavage stage embryos give rise to all the mesoderm and endoderm of the sea urchin larva. In these studies, vegetal blastomers were labeled no later than the sixth cleavage division (60-64 cell stage). In an earlier study we showed that single cells in the vegetal plate of the blastula stage Lytechinus variegatus embryo could be labeled in situ with the fluorescent, lipophilic dye, DiI(C18), and that cells labeled in the central region of the vegetal plate of the mesenchyme blastula primarily gave rise to homogeneous clones consisting of a single secondary mesenchyme cell (SMC) type (Ruffins and Ettensohn (1993) Dev. Biol. 160, 285–288). Our clonal labeling showed that a detailed fate map could be generated using the DiI(C18) labeling technique. Such a fate map could provide information about the spatial relationships between the precursors of specific mesodermal and endodermal cell types and information concerning the movements of these cells during gastrulation and later embryogenesis. We have used this method to construct the first detailed fate map of the vegetal plate of the sea urchin embryo. Ours is a latitudinal map; mapping from the plate center, where the mesodermal precursors reside, through the region which contains the endodermal precursors and across the ectodermal boundary. We found that the precursors of certain SMC types are segregated in the mesenchyme blastula stage vegetal plate and that prospective germ layers reside within specific boundaries. To determine whether the vegetal plate is radially symmetrical with respect to mesodermal cell fates, single blastomeres of four cell stage embryos were injected with lysyl-rhodamine dextran (LRD). The resulting ectodermal labeling patterns were classified and correlated with the SMC types labeled. This analysis indicates that the dorsal and ventral blastomers do not contribute equally to SMC derivatives in L. variegatus.

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