Competence of the animal cap to react with the inductive signal from micromere descendants in the hatching blastula stage of echinoid embryos

Zygote ◽  
1999 ◽  
Vol 8 (S1) ◽  
pp. S81-S81
Author(s):  
Yasuhiro Ishizuka ◽  
Shonan Amemiya
Keyword(s):  
Sea Urchin ◽  
Blastula Stage ◽  
Cell Stage ◽  
Scaphechinus Mirabilis ◽  
Glass Needle ◽  
Kanagawa Prefecture ◽  
Rhodamine B Isothiocyanate ◽  
Okayama Prefecture ◽  
Inductive Signal ◽  
Hemicentrotus Pulcherrimus

Micromere signalling is a key to understanding the developmental mechanisms underlying endomesoderm differentiation along the A-V axis in sea urchin embryos. A recent study has shown that micromere activity in inducing endo-mesoderm differentiation of mesomere descendants is, unexpectedly, maximal at the hatching blastula (HB) stage in the echinoids Scaphechinus mirabilis and Hemicentrotus pulcherrimus. This study focused mainly on the timing of emission of the inductive signal by the micromere descendants. The timing of animal cap competent to react to the inductive signal from micromeres was not specifically investigated.In the present study, we examined the competence of mesomere descendants at the HB stage to react to the inductive signal of micromere descendants by producing recombinant embryos of the descendants of mesomeres and micromeres.Adults of the sand dollar Scaphechinus mirabilis and the sea urchin Hemicentrotus pulcherrimus were collected along the shore of Shiraishi Island, Okayama Prefecture and in the vicinity of the Misaki Marine Biological Station, Kanagawa Prefecture, respectively. The fertilised eggs were separated into two groups immediately after removal of the fertilisation membranes. One group was cultured in normal artificial seawater (ASW), and the other in ASW containing 50 mg/ml of rhodamine B isothiocyanate (RITC). At the early 16-cell stage unlabelled embryos were transferred to calcium-free seawater (CFSW) and dissected in the equatorial plane with a glass needle to isolate animal caps consisting of eight mesomeres. Embryos labeled with RITC were also transferred to CFSW at the same stage and dissected to isolate four micromeres. An isolated animal cap and a quartet of micromeres were cultured separately in ASW and recombined at the stage corresponding to the HB of control (undisturbed) embryos.

scholarly journals Involvement of Huntingtin in Development and Ciliary Beating Regulation of Larvae of the Sea Urchin, Hemicentrotus pulcherrimus

2021 ◽  
Vol 22 (10) ◽  
pp. 5116
Author(s):  
Hideki Katow ◽  
Tomoko Katow ◽  
Hiromi Yoshida ◽  
Masato Kiyomoto
Keyword(s):  
Sea Urchin ◽  
The Body ◽  
Brdu Incorporation ◽  
Wild Type ◽  
Blastula Stage ◽  
Ciliary Beating ◽  
Marker Proteins ◽  
Disease Protein ◽  
Pattern Regulation ◽  
Hemicentrotus Pulcherrimus

The multiple functions of the wild type Huntington’s disease protein of the sea urchin Hemicentrotus pulcherrimus (Hp-Htt) have been examined using the anti-Hp-Htt antibody (Ab) raised against synthetic oligopeptides. According to immunoblotting, Hp-Htt was detected as a single band at around the 350 kDa region at the swimming blastula stage to the prism larva stage. From the 2-arm pluteus stage (2aPL), however, an additional smaller band at the 165 kDa region appeared. Immunohistochemically, Hp-Htt was detected in the nuclei and the nearby cytoplasm of the ectodermal cells from the swimming blastula stage, and the blastocoelar cells from the mid-gastrula stage. The Ab-positive signal was converged to the ciliary band-associated strand (CBAS). There, it was accompanied by several CBAS-marker proteins in the cytoplasm, such as glutamate decarboxylase. Application of Hp-Htt morpholino (Hp-Htt-MO) has resulted in shortened larval arms, accompanied by decreased 5-bromo-2-deoxyuridin (BrdU) incorporation by the ectodermal cells of the larval arms. Hp-Htt-MO also resulted in lowered ciliary beating activity, accompanied by a disordered swirling pattern formation around the body. These Hp-Htt-MO-induced deficiencies took place after the onset of CBAS system formation at the larval arms. Thus, Hp-Htt is involved in cell proliferation and the ciliary beating pattern regulation signaling system in pluteus larvae.

Origin and behaviour of pigment cells in sea urchin embryos

Zygote ◽  
1999 ◽  
Vol 8 (S1) ◽  
pp. S42-S43 ◽  
Author(s):  
Tetsuya Kominami
Keyword(s):  
Sea Urchin ◽  
Pigment Cell ◽  
Cell Lineage ◽  
Pigment Cells ◽  
Cleavage Stage ◽  
Fate Map ◽  
Blastula Stage ◽  
Cell Stage ◽  
Stage Embryo ◽  
Founder Cells

Sea urchin pluteus larvae contain dozens of pigment cells in their ectoderm. These pigment cells are the descendants of the veg2 blastomeres of the 60-cell stage embryo. According to the fate map made by Ruffins and Ettensohn, the prospective pigment cells occupy the central region of the vegetal plate. Most of these prospective pigment cells exclusively give rise to pigment cells. Therefore, specification of the pigment cell lineage should occur at some point between the 60-cell and mesenchyme blastula stage. However, the detailed process of the specification of the pigment lineage is unknown.When are pigment cells specified? Are cell interactions necessary for the specification? Do founder cells exist? To answer these questions, I treated embryos with Ca2+-free seawater during the cleavage stage and examined the number of pigment cells observed in pluteus larvae. Treatment at 5.5–8.5 h and especially 7.5–10.5 h postfertilisation markedly reduced the number of pigment cells. The decrease was statistically significant. On the other hand, the treatment at 3.5–6.5 h or 9.5–12.5 h never reduced the number of pigment cells. By examining the frequency of the appearance of embryos whose numbers of pigment cells were less than 20, it was also found that the numbers of pigment cells were frequently in multiples of 4. Embryos having 4, 8, 12, 16 and 20 pigment cells were more frequently observed. Statistics indicated that the frequency of appearance was not random. These results indicated that cell contacts are necessary for the specification of pigment cells and that the specification occurs from 7 to 10 h postfertilisation. The results also suggest that the founder cells, if they exist, divide twice before they differentiate into pigment cells.

Spatial and temporal expression pattern during sea urchin embryogenesis of a gene coding for a protease homologous to the human protein BMP-1 and to the product of the Drosophila dorsal-ventral patterning gene tolloid

Development ◽  
1992 ◽  
Vol 114 (1) ◽  
pp. 147-163 ◽  
Author(s):  
T. Lepage ◽  
C. Ghiglione ◽  
C. Gache
Keyword(s):  
Sea Urchin ◽  
Catalytic Domain ◽  
Intracellular Localization ◽  
Regulation Of Gene Expression ◽  
Drosophila Embryo ◽  
Limited Area ◽  
Fate Map ◽  
Blastula Stage ◽  
Cell Stage ◽  
Short Period

A cDNA clone coding for a sea urchin embryonic protein was isolated from a prehatching blastula lambda gt11 library. The predicted translation product is a secreted 64 × 10(3) Mr enzyme designated as BP10. The protein contains several domains: a signal peptide, a putative propeptide, a catalytic domain with an active center typical of a Zn(2+)-metalloprotease, an EGF-like domain and two internal repeats similar to repeated domains found in the C1s and C1r serine proteases of the complement cascade. The BP10 protease is constructed with the same domains as the human bone morphogenetic protein BMP-1, a protease described as a factor involved in bone formation, and as the recently characterized product of the tolloid gene which is required for correct dorsal-ventral patterning of the Drosophila embryo. The transcription of the BP10 gene is transiently activated around the 16- to 32-cell stage and the accumulation of BP10 transcripts is limited to a short period at the blastula stage. By in situ hybridization with digoxygenin-labelled RNA probes, the BP10 transcripts were only detected in a limited area of the blastula, showing that the transcription of the BP10 gene is also spatially controlled. Antibodies directed against a fusion protein were used to detect the BP10 protein in embryonic extracts. The protein is first detected in early blastula stages, its level peaks in late cleavage, declines abruptly before ingression of primary mesenchyme cells and remains constant in late development. The distribution of the BP10 protein during its synthesis and secretion was analysed by immunostaining blastula-stage embryos. The intracellular localization of the BP10 staining varies with time. The protein is first detected in a perinuclear region, then in an apical and submembranous position just before its secretion into the perivitelline space. The protein is synthesized in a sharply delimited continuous territory spanning about 70% of the blastula. Comparison of the size and orientation of the labelled territory in the late blastula with the fate map of the blastula stage embryo shows that the domain in which the BP10 gene is expressed corresponds to the presumptive ectoderm. Developing embryos treated with purified antibodies against the BP10 protein and with synthetic peptides derived from the EGF-like domain displayed perturbations in morphogenesis and were radialized to various degrees. These results are consistent with a role for BP10 in the differentiation of ectodermal lineages and subsequent patterning of the embryo. On the basis of these results, we speculate that the role of BP10 in the sea urchin embryo might be similar to that of tolloid in Drosophila. We discuss the idea that the processes of spatial regulation of gene expression along the animal-vegetal in sea urchin and dorsal-ventral axes in Drosophila might have some similarities and might use common elements.

scholarly journals A Cytochemical Study of the Sulfhydryl Groups of Sea Urchin Eggs during the First Cleavage

1958 ◽  
Vol 4 (5) ◽  
pp. 615-620 ◽  
Author(s):  
Naoko Kawamura ◽  
Katsuma Dan
Keyword(s):  
Sea Urchin ◽  
Azo Dye ◽  
Sea Water ◽  
Color Intensity ◽  
Cell Stage ◽  
Cytochemical Study ◽  
Sea Urchin Eggs ◽  
Sh Groups ◽  
Precursor Material ◽  
Hemicentrotus Pulcherrimus

In the eggs of four species of echinoderms, Mespilia globulus, Pseudocentrotus depressus, Hemicentrotus pulcherrimus and Clypeaster japonicus, changes in the distribution of protein-bound SH groups from fertilization to the 2 cell stage have been studied cytochemically by use of a mercaptide-forming azo dye. In the eggs of these species, the color intensity in the cytoplasm increased upon fertilization. The astral centers and spindle during mitosis were stained deeply. When the aster formation was suppressed by ether, hyaline spots appeared in the egg cytoplasm instead of well formed astral centers and these spots were stained by the SH-specific dye. Upon recovery of such eggs in pure sea water, and when cleavage ensued, such spots disappeared and two new astral centers were reorganized. The SH-protein occurring in the centrosphere is considered to be the precursor material for the asters and spindle, and this material is apparently derived from the cytoplasm.

scholarly journals Size classes of replication units in DNA from sea urchin embryos.

1979 ◽  
Vol 81 (3) ◽  
pp. 698-703 ◽  
Author(s):  
M P Kurek ◽  
D Billig ◽  
P Stambrook
Keyword(s):  
Sea Urchin ◽  
Blastula Stage ◽  
Substantial Fraction ◽  
Base Pairs ◽  
Cell Stage ◽  
Size Classes ◽  
Detectable Difference ◽  
Bubble Sizes ◽  
Two Stages ◽  
Sea Urchin Sperm

Sea urchin DNA containing replication structures was isolated from two to four cell stage and blastula stage embryos, and examined by electron microscopy. In addition to the expected eye forms, we also observed molecules with large internal single-stranded gaps. Such structures were not present in DNA devoid of replicating molecules such as that isolated from sea urchin sperm. When the size of eye forms and interbubble distances between the two stages were compared, there was no detectable difference. In both stages, we observed two distinct size classes of bubbles and of interbubble distances. In the case of bubble sizes, the smaller size class was comprised of clustered microbubbles that ranged from 200 base pairs to 1 Kilobase (kb) with a mean of 432 base pairs. The large eye forms measured 1--35 kb with a mean of 6.8 kb. Interbubble distances also yielded two distinct populations, with the smaller class ranging from 400 base pairs to 2.3 kb (mean = 1.1 kb) and the larger population ranging from 2.8 to 36 kb (mean = 10.9 kb). Although other possibilities cannot be entirely excluded, the data support the contention that a substantial fraction of the larger eye-form population arises from the fusion of the clustered microbubbles.

scholarly journals Cyclin D and cdk4 Are Required for Normal Development beyond the Blastula Stage in Sea Urchin Embryos

2002 ◽  
Vol 22 (13) ◽  
pp. 4863-4875 ◽  
Author(s):  
Jennifer C. Moore ◽  
Jan L. Sumerel ◽  
Bradley J. Schnackenberg ◽  
Jason A. Nichols ◽  
Athula Wikramanayake ◽  
...  
Keyword(s):  
Sea Urchin ◽  
Normal Development ◽  
Sea Urchins ◽  
Ectopic Expression ◽  
Cyclin D ◽  
Blastula Stage ◽  
Cell Stage ◽  
Cell Cycles ◽  
Early Embryos ◽  
Cdk4 Expression

ABSTRACT cdk4 mRNA and protein are constitutively expressed in sea urchin eggs and throughout embryonic development. In contrast, cyclin D mRNA is barely detectable in eggs and early embryos, when the cell cycles consist of alternating S and M phases. Cyclin D mRNA increases dramatically in embryos at the early blastula stage and remains at a constant level throughout embryogenesis. An increase in cdk4 kinase activity occurs concomitantly with the increase in cyclin D mRNA. Ectopic expression of cyclin D mRNA in eggs arrests development before the 16-cell stage and causes eventual embryonic death, suggesting that activation of cyclin D/cdk4 in cleavage cell cycles is lethal to the embryo. In contrast, blocking cyclin D or cdk4 expression with morpholino antisense oligonucleotides results in normal development of early gastrula-stage embryos but abnormal, asymmetric larvae. These results suggest that in sea urchins, cyclin D and cdk4 are required for normal development and perhaps the patterning of the developing embryo, but may not be directly involved in regulating entry into the cell cycle.

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.

T-brain homologue (HpTb) is involved in the archenteron induction signals of micromere descendant cells in the sea urchin embryo

Development ◽  
2002 ◽  
Vol 129 (22) ◽  
pp. 5205-5216 ◽  
Author(s):  
Takuya Fuchikami ◽  
Keiko Mitsunaga-Nakatsubo ◽  
Shonan Amemiya ◽  
Toshiya Hosomi ◽  
Takashi Watanabe ◽  
...  
Keyword(s):  
Wnt Signaling ◽  
Sea Urchin ◽  
Nuclear Localization ◽  
Signaling Cascade ◽  
Gastrula Stage ◽  
Blastula Stage ◽  
Sea Urchin Embryo ◽  
Mesenchyme Cells ◽  
Antisense Morpholino ◽  
Hemicentrotus Pulcherrimus

Signals from micromere descendants play a crucial role in sea urchin development. In this study, we demonstrate that these micromere descendants express HpTb, a T-brain homolog of Hemicentrotus pulcherrimus. HpTb is expressed transiently from the hatched blastula stage through the mesenchyme blastula stage to the gastrula stage. By a combination of embryo microsurgery and antisense morpholino experiments, we show that HpTb is involved in the production of archenteron induction signals. However, HpTb is not involved in the production of signals responsible for the specification of secondary mesenchyme cells, the initial specification of primary mesenchyme cells, or the specification of endoderm.HpTb expression is controlled by nuclear localization ofβ-catenin, suggesting that HpTb is in a downstream component of the Wnt signaling cascade. We also propose the possibility that HpTbis involved in the cascade responsible for the production of signals required for the spicule formation as well as signals from the vegetal hemisphere required for the differentiation of aboral ectoderm.

Morphogenesis of exogut isolated from vegetalised embryo of sea urchin

Zygote ◽  
1999 ◽  
Vol 8 (S1) ◽  
pp. S84-S84
Author(s):  
Yasuyuki Kamata ◽  
Kazuyuki Endo ◽  
Hiroyuki Nozaki ◽  
Akiko Fujiwara ◽  
Ikuo Yasumasu
Keyword(s):  
High Activity ◽  
Sea Urchin ◽  
Normal Development ◽  
Sea Urchins ◽  
Cell Stage ◽  
Minimum Concentration ◽  
Spherical Structure ◽  
Chloroform Methanol ◽  
Early Gastrula ◽  
Hemicentrotus Pulcherrimus

It is well known that sea urchin embryos treated with lithium chloride (LiC1) develop to abnormally into vegetalised embryos, in which differentiation of ectodermal cells is inhibited. When embryos of the sea urchins, Hemicentrotus pulcherrimus and Anthocidaris crassispina were treated with 20 mM LiC1 from the 8-cell stage to the corresponding early gastrula stage, they developed to vegetalised embryos with a large exogut 45 h after fertilisation. In these vegetalised embryos, high activity of alkaline phosphatase (AP) was detected histochemically at the end of the exogut where it is attached to the embryo body. High activity of AP is known to be detected specifically in the gut of sea urchin pluteus larvae by the same procedure as used in this study. Hence, we concluded that this part of the exogut is composed of the cells which develop into the cells of the gut in normal development.When exogut isolated from vegetalised embryos was cultured in the extract obtained from eggs or embryos, the end composed of the cells in which high AP activity was detected, expanded during culture and formed a large spherical structure about 24 h after the initiation of culture. The minimum concentration of extract to cause expansion of isolated exogut was 5 × 103 egg or embryo equivalent/ml ASW (artificial seawater). The extract boiled at 95 °C for 1 h also caused expansion of isolated exogut at the same concentrations as non-boiled extract. On the other hand, the extract obtained from eggs or embryos by chloroform–methanol extraction did not cause any expansion of exogut, but the aqueous phase, heat-dried and dissolved in ASW, induced expansion of isolated exogut.

Uridineinbau in die Nucleinsäuren von Furchungsstadien der Eier des Seeigels Paracentrotus Lividus

1967 ◽  
Vol 22 (11) ◽  
pp. 1176-1182 ◽  
Author(s):  
G. Czihak ◽  
H. G. Wittmann ◽  
I. Hindennach
Keyword(s):  
Sea Urchin ◽  
Rna Synthesis ◽  
Fission Products ◽  
Paracentrotus Lividus ◽  
The Other ◽  
Soluble Form ◽  
Blastula Stage ◽  
High Turnover ◽  
Cell Stage ◽  
Sea Urchin Egg

1. Uridine taken up during the 16th-cell stage of the sea urchin egg is stored in acid soluble form.2. During the interphase of this stage uridine is used for an RNA-synthesis in the micromeres, the smallest blastomeres of the egg, which have a prolonged interphase. There seems to occur no RNA-synthesis in the other blastomeres of this stage or the synthesis in these cells is considerably smaller. The RNA of the micromeres has a high turnover, it is neither tRNA nor rRNA but probably mRNA.3. 2½ hours after the 16th-cell stage — this is in the young blastula stage — about 5½ hours after fertilization, at least half of the micromere-RNA is already decomposed, the fission products of which having been used together with the stored uridine to synthesize rRNA and (after methylation of the uridine) DNA.

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