A micromere induction signal is activated by beta-catenin and acts through notch to initiate specification of secondary mesenchyme cells in the sea urchin embryo

Development ◽  
2000 ◽  
Vol 127 (23) ◽  
pp. 5113-5122 ◽  
Author(s):  
D.R. McClay ◽  
R.E. Peterson ◽  
R.C. Range ◽  
A.M. Winter-Vann ◽  
M.J. Ferkowicz
Keyword(s):  
Sea Urchin ◽  
Notch Pathway ◽  
Notch Receptor ◽  
Cell Diameter ◽  
Beta Catenin ◽  
Animal Pole ◽  
Induction Signal ◽  
Vegetal Pole ◽  
Mesenchyme Cells ◽  
To Receive

At fourth cleavage of sea urchin embryos four micromeres at the vegetal pole separate from four macromeres just above them in an unequal cleavage. The micromeres have the capacity to induce a second axis if transplanted to the animal pole and the absence of micromeres at the vegetal pole results in the failure of macromere progeny to specify secondary mesenchyme cells (SMCs). This suggests that micromeres have the capacity to induce SMCs. We demonstrate that micromeres require nuclear beta-catenin to exhibit SMC induction activity. Transplantation studies show that much of the vegetal hemisphere is competent to receive the induction signal. The micromeres induce SMCs, most likely through direct contact with macromere progeny, or at most a cell diameter away. The induction is quantitative in that more SMCs are induced by four micromeres than by one. Temporal studies show that the induction signal is passed from the micromeres to macromere progeny between the eighth and tenth cleavage. If micromeres are removed from hosts at the fourth cleavage, SMC induction in hosts is rescued if they later receive transplanted micromeres between the eighth and tenth cleavage. After the tenth cleavage addition of induction-competent micromeres to micromereless embryos fails to specify SMCs. For macromere progeny to be competent to receive the micromere induction signal, beta-catenin must enter macromere nuclei. The macromere progeny receive the micromere induction signal through the Notch receptor. Signaling-competent micromeres fail to induce SMCs if macromeres express dominant-negative Notch. Expression of an activated Notch construct in macromeres rescues SMC specification in the absence of induction-competent micromeres. These data are consistent with a model whereby beta-catenin enters the nuclei of micromeres and, as a consequence, the micromeres produce an inductive ligand. Between the eighth and tenth cleavage micromeres induce SMCs through Notch. In order to be receptive to the micromere inductive signal the macromeres first must transport beta-catenin to their nuclei, and as one consequence the Notch pathway becomes competent to receive the micromere induction signal, and to transduce that signal. As Notch is maternally expressed in macromeres, additional components must be downstream of nuclear beta-catenin in macromeres for these cells to receive and transduce the micromere induction signal.

Identification and localization of a sea urchin Notch homologue: insights into vegetal plate regionalization and Notch receptor regulation

Development ◽  
1997 ◽  
Vol 124 (17) ◽  
pp. 3363-3374 ◽  
Author(s):  
D.R. Sherwood ◽  
D.R. McClay
Keyword(s):  
Sea Urchin ◽  
Basolateral Membrane ◽  
Notch Pathway ◽  
Cell Types ◽  
Dorsal Side ◽  
Notch Receptor ◽  
Molecular Organization ◽  
Intercellular Signaling ◽  
Vegetal Pole ◽  
Numerous Cell

The specifications of cell types and germ-layers that arise from the vegetal plate of the sea urchin embryo are thought to be regulated by cell-cell interactions, the molecular basis of which are unknown. The Notch intercellular signaling pathway mediates the specification of numerous cell fates in both invertebrate and vertebrate development. To gain insights into mechanisms underlying the diversification of vegetal plate cell types, we have identified and made antibodies to a sea urchin homolog of Notch (LvNotch). We show that in the early blastula embryo, LvNotch is absent from the vegetal pole and concentrated in basolateral membranes of cells in the animal half of the embryo. However, in the mesenchyme blastula embryo LvNotch shifts strikingly in subcellular localization into a ring of cells which surround the central vegetal plate. This ring of LvNotch delineates a boundary between the presumptive secondary mesoderm and presumptive endoderm, and has an asymmetric bias towards the dorsal side of the vegetal plate. Experimental perturbations and quantitative analysis of LvNotch expression demonstrate that the mesenchyme blastula vegetal plate contains both animal/vegetal and dorsoventral molecular organization even before this territory invaginates to form the archenteron. Furthermore, these experiments suggest roles for the Notch pathway in secondary mesoderm and endoderm lineage segregation, and in the establishment of dorsoventral polarity in the endoderm. Finally, the specific and differential subcellular expression of LvNotch in apical and basolateral membrane domains provides compelling evidence that changes in membrane domain localization of LvNotch are an important aspect of Notch receptor function.

GSK3beta/shaggy mediates patterning along the animal-vegetal axis of the sea urchin embryo

Development ◽  
1998 ◽  
Vol 125 (13) ◽  
pp. 2489-2498 ◽  
Author(s):  
F. Emily-Fenouil ◽  
C. Ghiglione ◽  
G. Lhomond ◽  
T. Lepage ◽  
C. Gache
Keyword(s):  
Sea Urchin ◽  
Wnt Pathway ◽  
Dominant Negative ◽  
Early Gene ◽  
Expression Domain ◽  
Animal Pole ◽  
Axial Patterning ◽  
Sea Urchin Embryo ◽  
Vegetal Pole ◽  
Dominant Negative Activity

In the sea urchin embryo, the animal-vegetal axis is defined before fertilization and different embryonic territories are established along this axis by mechanisms which are largely unknown. Significantly, the boundaries of these territories can be shifted by treatment with various reagents including zinc and lithium. We have isolated and characterized a sea urchin homolog of GSK3beta/shaggy, a lithium-sensitive kinase which is a component of the Wnt pathway and known to be involved in axial patterning in other embryos including Xenopus. The effects of overexpressing the normal and mutant forms of GSK3beta derived either from sea urchin or Xenopus were analyzed by observation of the morphology of 48 hour embryos (pluteus stage) and by monitoring spatial expression of the hatching enzyme (HE) gene, a very early gene whose expression is restricted to an animal domain with a sharp border roughly coinciding with the future ectoderm / endoderm boundary. Inactive forms of GSK3beta predicted to have a dominant-negative activity, vegetalized the embryo and decreased the size of the HE expression domain, apparently by shifting the boundary towards the animal pole. These effects are similar to, but even stronger than, those of lithium. Conversely, overexpression of wild-type GSK3beta animalized the embryo and caused the HE domain to enlarge towards the vegetal pole. Unlike zinc treatment, GSK3beta overexpression thus appeared to provoke a true animalization, through extension of the presumptive ectoderm territory. These results indicate that in sea urchin embryos the level of GSKbeta activity controls the position of the boundary between the presumptive ectoderm and endoderm territories and thus, the relative extent of these tissue layers in late embryos. GSK3beta and probably other downstream components of the Wnt pathway thus mediate patterning both along the primary AV axis of the sea urchin embryo and along the dorsal-ventral axis in Xenopus, suggesting a conserved basis for axial patterning between invertebrate and vertebrate in deuterostomes.

Scanning electron microscopy of gastrulation in a sea urchin (Anthocidaris crassispina)

Development ◽  
1982 ◽  
Vol 67 (1) ◽  
pp. 27-35
Author(s):  
Shonan Amemiya ◽  
Koji Akasaka ◽  
Hiroshi Terayama
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Cell Formation ◽  
Cell Movement ◽  
Animal Pole ◽  
Vegetal Pole ◽  
Mesenchyme Cell ◽  
Major Motive ◽  
Mesenchyme Cells ◽  
Scanning Electron

Gastrulation in Anthocidaris was investigated by observing the inside and the outside of embryos by scanning electron microscopy. Furrows which possibly rėflect changes in intercellular interactions were observed on the outer surface (hyaline layer side) of embryos twice in development: firstly at the time of primary mesenchyme cell formation, and secondly at the time of vegetal plate indentation. In the latter case, the cells within and surrounding the vegetal plate appeared to change their shapes differently; the former (within the plate) having broader surfaces on the blastocoel side whereas the latter (surrounding the plate) having broader surfaces on the hyaline layer side. This suggests that the first phase of indentation may be mediated by the autonomous change of cell shape and intercellular adhesiveness, accompanied by an autonomous cell movement in the vegetal pole region. Although some pseudopodial linkages were observed between secondary mesenchyme cells on the top of the invaginating archenteron and the animal pole in the mid-gastrula and later stage embryos, they were thinner and smaller in number as compared to those in the Pseudocentrotus embryos. The rate of invagination appeared rather constant throughout gastrulation in contrast to the accelerated invagination in other embryos with larger blastocoel cavities. Moreover, the number of columnar cells on the dissected surface of embryos remained unaltered. These findings suggest that the secondary mesenchyme cells may act as a linker between the archenteron tip and the animal pole, but they may not generate major motive forces for archenteron invagination at least in the Anthocidaris embryos.

Specification of endoderm and mesoderm in the sea urchin

Zygote ◽  
1999 ◽  
Vol 8 (S1) ◽  
pp. S41-S41 ◽  
Author(s):  
David R. McClay
Keyword(s):  
Sea Urchin ◽  
Wnt Pathway ◽  
Special Significance ◽  
Nuclear Entry ◽  
Cell Stage ◽  
Animal Pole ◽  
Secondary Axis ◽  
Sea Urchin Embryo ◽  
Vegetal Pole

It has long been recognized that micromeres have special significance in early specification events in the sea urchin embryo. Micromeres have the ability to induce a secondary axis if transferred to the animal pole at the 16-cell stage of sea urchin embryos (Hörstadius, 1939). Without micromeres an isolated animal hemisphere develops into an ectodermal ball called a dauer blastula. Addition of micromeres to an animal half rescues a normal pluteus larva, including endoderm (Hörstadius, 1939). Despite these well-known experiments, however, neither the molecular basis of that induction nor the endogenous inductive role of micromeres in development was known. In recent experiments we learned that if one eliminates micromeres from the vegetal pole at the 16-cell stage the resulting embryo makes no secondary mesenchyme. Earlier it had been found that β-catenin is crucial for specification events that lead to mesoderm and endoderm (Wikra-manayake et al., 1998; Emily-Fenouil et al., 1998; Logan et al., 1999). We noticed that at the 16-cell stage β-catenin enters the nuclei of micromeres, then enters the nuclei of macromeres at the 32-cell stage (Logan et al., 1999). Since nuclear entry of β-catenin is known to be important for its signalling function in the Wnt pathway, we asked whether β-catenin functions in the micromere induction pathway.

Studies of the cleavage in the frog egg

Development ◽  
1969 ◽  
Vol 21 (1) ◽  
pp. 119-129
Author(s):  
T. Kubota
Keyword(s):  
Sea Urchin ◽  
Sea Urchins ◽  
Cleavage Furrow ◽  
Mitotic Apparatus ◽  
Animal Pole ◽  
Sea Urchin Eggs ◽  
Rana Nigromaculata ◽  
Vegetal Pole ◽  
Egg Cortex

In sea-urchin eggs, once karyokinesis reaches metaphase or anaphase, the cleavage furrow can be formed even if the mitotic apparatus is destroyed (Swann & Mitchison, 1953) or removed (Hiramoto, 1956). A similar result was obtained in frog eggs (Kubota, 1966). In amphibian eggs a much longer time is available for performing experiments than in sea urchins as the furrow first appears at the animal pole and slowly travels toward the vegetal pole. Taking advantage of this situation, Waddington (1952) and Dan & Kuno-Kojima (1963) performed various kinds of operations to elucidate the roles of the egg cortex and the inner cytoplasm in furrow formation, and Selman & Waddington (1955) also made cytological observations of the process. In the present paper a shift of the inner cytoplasm relative to the cortex and its influence on the course of the furrow was analysed for eggs of the frog Rana nigromaculata.

Nuclear beta-catenin is required to specify vegetal cell fates in the sea urchin embryo

Development ◽  
1999 ◽  
Vol 126 (2) ◽  
pp. 345-357 ◽  
Author(s):  
C.Y. Logan ◽  
J.R. Miller ◽  
M.J. Ferkowicz ◽  
D.R. McClay
Keyword(s):  
Cell Fate ◽  
Sea Urchin ◽  
Beta Catenin ◽  
Cell Fate Specification ◽  
Cell Fates ◽  
Cell Stage ◽  
Animal Pole ◽  
Fate Specification ◽  
The Relationship ◽  
Vegetal Cell

Beta-catenin is thought to mediate cell fate specification events by localizing to the nucleus where it modulates gene expression. To ask whether beta-catenin is involved in cell fate specification during sea urchin embryogenesis, we analyzed the distribution of nuclear beta-catenin in both normal and experimentally manipulated embryos. In unperturbed embryos, beta-catenin accumulates in nuclei that include the precursors of the endoderm and mesoderm, suggesting that it plays a role in vegetal specification. Using pharmacological, embryological and molecular approaches, we determined the function of beta-catenin in vegetal development by examining the relationship between the pattern of nuclear beta-catenin and the formation of endodermal and mesodermal tissues. Treatment of embryos with LiCl, a known vegetalizing agent, caused both an enhancement in the levels of nuclear beta-catenin and an expansion in the pattern of nuclear beta-catenin that coincided with an increase in endoderm and mesoderm. Conversely, overexpression of a sea urchin cadherin blocked the accumulation of nuclear beta-catenin and consequently inhibited the formation of endodermal and mesodermal tissues including micromere-derived skeletogenic mesenchyme. In addition, nuclear beta-catenin-deficient micromeres failed to induce a secondary axis when transplanted to the animal pole of uninjected host embryos, indicating that nuclear beta-catenin also plays a role in the production of micromere-derived signals. To examine further the relationship between nuclear beta-catenin in vegetal nuclei and micromere signaling, we performed both transplantations and deletions of micromeres at the 16-cell stage and demonstrated that the accumulation of beta-catenin in vegetal nuclei does not require micromere-derived cues. Moreover, we demonstrate that cell autonomous signals appear to regulate the pattern of nuclear beta-catenin since dissociated blastomeres possessed nuclear beta-catenin in approximately the same proportion as that seen in intact embryos. Together, these data show that the accumulation of beta-catenin in nuclei of vegetal cells is regulated cell autonomously and that this localization is required for the establishment of all vegetal cell fates and the production of micromere-derived signals.

Commitment along the dorsoventral axis of the sea urchin embryo is altered in response to NiCl2

Development ◽  
1992 ◽  
Vol 116 (3) ◽  
pp. 671-685 ◽  
Author(s):  
J. Hardin ◽  
J.A. Coffman ◽  
S.D. Black ◽  
D.R. McClay
Keyword(s):  
Sea Urchin ◽  
Major Effect ◽  
Specific Gene ◽  
Animal Pole ◽  
Dorsoventral Axis ◽  
Dorsoventral Polarity ◽  
Normal Number ◽  
Sea Urchin Embryo ◽  
Mesenchyme Cells ◽  
Primary Mesenchyme Cells

Few treatments are known that perturb the dorsoventral axis of the sea urchin embryo. We report here that the dorsoventral polarity of the sea urchin embryo can be disrupted by treatment of embryos with NiCl2. Lytechinus variegatus embryos treated with 0.5 mM NiCl2 from fertilization until the early gastrula stage appear morphologically normal until the midgastrula stage, when they fail to acquire the overt dorsoventral polarity characteristic of untreated siblings. The ectoderm of normal embryos possesses two ventrolateral thickenings just above the vegetal plate region. In nickel-treated embryos, these become expanded as a circumferential belt around the vegetal plate. The ectoderm just ventral to the animal pole normally invaginates to form a stomodeum, which then fuses with the tip of the archenteron to produce the mouth. In nickel-treated embryos, the stomodeal invagination is expanded to become a circumferential constriction, and it eventually pinches off as the tip of the archenteron fuses with it to produce a mouth. Primary mesenchyme cells form a ring in the lateral ectoderm, but as many as a dozen spicule rudiments can form in a radial pattern. Dorsoventral differentiation of ectodermal tissues is profoundly perturbed: nickel-treated embryos underexpress transcripts of the dorsal (aboral) gene LvS1, they overexpress the ventral (oral) ectodermal gene product, EctoV, and the ciliated band is shifted to the vegetal margin of the embryo. Although some dorsoventral abnormalities are observed, animal-vegetal differentiation of the archenteron and associated structures seems largely normal, based on the localization of region-specific gene products. Gross differentiation of primary mesenchyme cells seems unaffected, since nickel-treated embryos possess the normal number of these cells. Furthermore, when all primary mesenchyme cells are removed from nickel-treated embryos, some secondary mesenchyme cells undergo the process of “conversion” (Ettensohn, C. A. and McClay, D. R. (1988) Dev. Biol. 125, 396–409), migrating to sites where the larval skeleton would ordinarily form and subsequently producing spicule rudiments. However, the skeletal pattern formed by the converted cells is completely radialized. Our data suggest that a major effect of NiCl2 is to alter commitment of ectodermal cells along the dorsoventral axis. Among the consequences appears to be a disruption of pattern formation by mesenchyme cells.

On ‘the Stationary Surface Ring’ in Heart-Shaped Cleavage

1958 ◽  
Vol 35 (2) ◽  
pp. 400-406
Author(s):  
KATSUMA DAN
Keyword(s):  
Cell Surface ◽  
Sea Urchin ◽  
Mitotic Spindle ◽  
Cleavage Furrow ◽  
Sand Dollar ◽  
Animal Pole ◽  
Sea Urchin Eggs ◽  
Stationary Surface ◽  
Vegetal Pole

1. The eggs of the sand dollar, Astriclypeus manni, and the medusa, Spirocodon saltatrix, were used for the reason that they cleave in heart shape, the cleavage furrow appearing earlier at the animal than at the vegetal pole. 2. By the superposition of drawings showing contours and astral centres as well as the positions of carbon markers on the cell surface, the presence of a pair of stationary circular zones of the cortex can be demonstrated. These remain absolutely stationary through successive stages of cleavage, as was shown to be true of regularly cleaving sea-urchin eggs. 3. The two planes determined by this pair of stationary surface rings tilt toward each other on the animal pole side in linear proportion to the eccentricity of the mitotic spindle within the cell, and the loci of the astral centres tend to slant toward the animal pole. 4. The above phenomena can be explained by the previously proposed theory for heart-shaped cleavage; i.e. the primary cause of heart-shaped cleavage is the eccentric position of the spindle, which in turn causes the rotation of the asters and the bending of the spindle.

Ca(2+) in specification of vegetal cell fate in early sea urchin embryos

2001 ◽  
Vol 204 (5) ◽  
pp. 823-834
Author(s):  
I. Yazaki
Keyword(s):  
Cell Fate ◽  
Sea Urchin ◽  
Small Cells ◽  
Beta Catenin ◽  
Cell Stage ◽  
Sea Urchin Embryos ◽  
Mesoderm Specification ◽  
New Findings ◽  
Vegetal Pole ◽  
Vegetal Cell

In sea urchin embryos, the first specification of cell fate occurs at the fourth cleavage, when small cells (the micromeres) are formed at the vegetal pole. The fate of other blastomeres is dependent on the receipt of cell signals originating from the micromeres. The micromeres are fated to become skeletogenic cells and show the ability to induce the endoderm (the archenteron) in the neighbouring cells during the 16- to 60-cell stage. Several molecules involved in signaling pathways, i.e. Notch for mesoderm specification, bone morphogenic protein (BMP) for ectoderm specification and beta-catenin for endoderm specification, are spatially and temporally expressed during development. In the micromeres, beta-catenin increases and subsequently localizes to the nuclei under the regulation of TCF, a nuclear binding partner of beta-catenin, until the 60-cell stage. However, the mechanisms activating these signaling substances are still unclear. In this article, I demonstrate some specific properties of the membrane and cytoplasm of micromeres including new findings on intracellular Ca(2+) concentration, and propose a mechanism by which the functional micromeres are autonoumously formed. The possible roles of these in the specification of vegetal cell fate in early development are discussed.

LvNotch signaling mediates secondary mesenchyme specification in the sea urchin embryo

Development ◽  
1999 ◽  
Vol 126 (8) ◽  
pp. 1703-1713 ◽  
Author(s):  
D.R. Sherwood ◽  
D.R. McClay
Keyword(s):  
Sea Urchin ◽  
Cell Types ◽  
Notch Signaling Pathway ◽  
Notch Receptor ◽  
Dominant Negative ◽  
Sea Urchin Embryo ◽  
Numerous Cell ◽  
Vertebrate Embryos ◽  
Mesenchyme Cells ◽  
Molecular Bases

Cell-cell interactions are thought to regulate the differential specification of secondary mesenchyme cells (SMCs) and endoderm in the sea urchin embryo. The molecular bases of these interactions, however, are unknown. We have previously shown that the sea urchin homologue of the LIN-12/Notch receptor, LvNotch, displays dynamic patterns of expression within both the presumptive SMCs and endoderm during the blastula stage, the time at which these two cell types are thought to be differentially specified (Sherwood, D. R. and McClay, D. R. (1997) Development 124, 3363–3374). The LIN-12/Notch signaling pathway has been shown to mediate the segregation of numerous cell types in both invertebrate and vertebrate embryos. To directly examine whether LvNotch signaling has a role in the differential specification of SMCs and endoderm, we have overexpressed activated and dominant negative forms of LvNotch during early sea urchin development. We show that activation of LvNotch signaling increases SMC specification, while loss or reduction of LvNotch signaling eliminates or significantly decreases SMC specification. Furthermore, results from a mosaic analysis of LvNotch function as well as endogenous LvNotch expression strongly suggest that LvNotch signaling acts autonomously within the presumptive SMCs to mediate SMC specification. Finally, we demonstrate that the expansion of SMCs seen with activation of LvNotch signaling comes at the expense of presumptive endoderm cells, while loss of SMC specification results in the endoderm expanding into territory where SMCs usually arise. Taken together, these results offer compelling evidence that LvNotch signaling directly specifies the SMC fate, and that this signaling is critical for the differential specification of SMCs and endoderm in the sea urchin embryo.

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