The midblastula cell cycle transition and the character of mesoderm in u.v.-induced nonaxial Xenopus development

Development ◽  
1987 ◽  
Vol 99 (2) ◽  
pp. 197-210
Author(s):  
J. Cooke ◽  
J.C. Smith
Keyword(s):  
Cell Cycle ◽  
Cell Number ◽  
Lateral Plate ◽  
Cell Stage ◽  
Mesodermal Cell ◽  
Larval Stages ◽  
Zygotic Transcription ◽  
Positional System ◽  
Three Stages ◽  
Probable Nature

Xenopus embryos (UV embryos) resulting from u.v. (254 nm) irradiation to the vegetal egg hemisphere and thus developing little or no axial pattern (UV5-Scharf & Gerhart, 1983), have been compared histologically with synchronous normal siblings at each of three stages. In addition, the relative amounts of blood-forming tissue produced in normal and in UV embryos have been studied by Western blotting total protein from larval stages and by immunofluorescence on sections. The observations on midblastulae (around 5000 cells) were aimed at detecting any systematic retardation, due to u.v., of the slowing of the cell cycle that normally commences at the 2-4000 cell stage and makes possible zygotic transcription and the preparation for gastrulation. No such retardation was apparent. Observations on postgastrular stages gave an assessment of the size and character of the population of mesoderm founder cells, in relation to the control, for embryos visibly undergoing entirely nonaxial development. Little deficit in total mesodermal cell number was found, though the entire mesoderm adopted the histological character proper to only some 40% of that in the normal pattern i.e. trunk lateral plate. Blood-forming capacity appears to be enhanced out of all proportion to the size of the mesoderm as a whole. The results are discussed in terms of the probable nature of the primary positional system for axial pattern and the later mechanisms of mesodermal patterning.

Cell number in relation to primary pattern formation in the embryo of Xenopus laevis

Development ◽  
1979 ◽  
Vol 53 (1) ◽  
pp. 269-289
Author(s):  
Jonathan Cooke
Keyword(s):  
Cell Cycle ◽  
Pattern Formation ◽  
Cell Cycle Control ◽  
Cell Number ◽  
The Body ◽  
Morphological Evidence ◽  
Tail Bud ◽  
Cell Cycle Time ◽  
Mesodermal Cell ◽  
Body Pattern

Morphological evidence is presented that definitive mesoderm formation in Xenopus is best understood as extending to the end of the neurula phase of development. A process of recruitment of cells from the deep neurectoderm layers into mesodermal position and behaviour, strictly comparable with that already agreed to occur around the internal blastoporal ‘lip’ during gastrula stages, can be shown to continue at the posterior end of the presumptive body pattern up to stage 20 (earliest tail bud). Spatial patterns of incidence of mitosis are described for the fifteen hours of development between the late gastrula and stage 20–22. These are related to the onset of new cell behaviours and overt cyto-differentiations characterizing the dorsal axial pattern,which occur in cranio-caudal and then medio-lateral spatial sequence as development proceeds. A relatively abrupt cessation of mitosis, among hitherto asynchronously cycling cells,precedes the other changes at each level in the presumptive axial pattern. The widespread incidence of cells still in DNA synthesis, anterior to the last mitoses in the posterior-to-anteriordevelopmental sequence of axial tissue, strongly suggests that cells of notochord and somites in their prolonged, non-cycling phase are G2-arrested, and thus tetraploid. This is discussed in relation to what is known of cell-cycle control in other situations. Best estimates for cell-cycle time in the still-dividing, posterior mesoderm of the neurula lie between 10 and 15 h. The supposition of continuing recruitment from neurectoderm can resolve an apparent discrepancy whereby total mesodermal cell number nevertheless contrives to double over a period of approximately 12 h during neurulation when most of the cells are leaving the cycle. Because of pre-existing evidence that cells maintain their relative positions (despite distortion)during the movements that form the mesodermal mantle, the patterns presented in this paper can be understood in two ways: as a temporal sequence of developmental events undergone by individual, posteriorly recruited cells as they achieve their final positions in the body pattern, or alternatively as a succession of wavefronts with respect to changes of cellstate, passing obliquely across the presumptive body pattern in antero-posterior direction. These concepts are discussed briefly in relation to recent ideas about pattern formation in growing systems.

Induction of the prospective neural crest of Xenopus

Development ◽  
1995 ◽  
Vol 121 (3) ◽  
pp. 767-777 ◽  
Author(s):  
R. Mayor ◽  
R. Morgan ◽  
M.G. Sargent
Keyword(s):  
Neural Crest ◽  
Marginal Zone ◽  
Ventral Side ◽  
Neural Plate ◽  
Lateral Plate ◽  
Cell Stage ◽  
Shape Characteristic ◽  
Zygotic Transcription ◽  
Neural Plate Border ◽  
Trunk Neural Crest

The earliest sign of the prospective neural crest of Xenopus is the expression of the ectodermal component of Xsna (the Xenopus homologue of snail) in a low arc on the dorsal aspect of stage 11 embryos, which subsequently assumes the horseshoe shape characteristic of the neural folds as the convergence-extension movements shape the neural plate. A related zinc-finger gene called Slug (Xslu) is expressed specifically in this tissue (i.e. the prospective crest) when the convergence extension movements are completed. Subsequently, Xslu is found in pre- and post-migratory cranial and trunk neural crest and also in lateral plate mesoderm after stage 17. Both Xslu and Xsna are induced by mesoderm from the dorsal or lateral marginal zone but not from the ventral marginal zone. From stage 10.5, explants of the prospective neural crest, which is underlain with tissue, are able to express Xslu. However expression of Xsna is not apparently specified until stage 12 and further contact with the inducer is required to raise the level of expression to that seen later in development. Xslu is specified at a later time. Embryos injected with noggin mRNA at the 1-cell stage or with plasmids driving noggin expression after the start of zygotic transcription express Xslu in a ring surrounding the embryo on the ventroposterior side. We suggest this indicates (a) that noggin interacts with another signal that is present throughout the ventral side of the embryo and (b) that Xslu is unable to express in the neural plate either because of the absence of a co-inducer or by a positive prohibition of expression. The ventral co-inducer, in the presence of overexpressed noggin, seems to generate an anterior/posterior pattern in the ventral part of the embryo comparable to that seen in neural crest of normal embryos. We suggest that the prospective neural crest is induced in normal embryos in the ectoderm that overlies the junction of the domains that express noggin and Xwnt-8. In support of this, we show animal cap explants from blastulae and gastrulae, treated with bFGF and noggin express Xslu but not NCAM although the mesoderm marker Xbra is also expressed. Explants treated with noggin alone express NCAM only. An indication that induction of the neural plate border is regulated independently of the neural plate is obtained from experiments using ultraviolet irradiation in the precleavage period. At certain doses, the cranial crest domains are not separated into lateral masses and there is a reduction in the size of the neural plate.

The system specifying body position in the early development of Xenopus, and its response to early perturbations.

Development ◽  
1985 ◽  
Vol 89 (Supplement) ◽  
pp. 69-87
Author(s):  
Jonathan Cooke
Keyword(s):  
Early Development ◽  
Body Position ◽  
Bilateral Symmetry ◽  
The Body ◽  
Spatial Profile ◽  
Cell Stage ◽  
Larval Stages ◽  
Positional System ◽  
Set Up ◽  
Pattern Specification

Evidence is presented that the system setting up preliminary specifications for contributions to the axial body plan, across vegetal regions of the Xenopus embryo, acts in a widespread way at early stages. Mechanisms that regulate the spatial profile of this primary positional variable, and thus ensure the constancy and harmony of the body plans normally achieved, have lost this integrative ability by the 4-cell stage one hour after the plasm shifts that precede first cleavage and symmetrize the egg. Abnormal, partial or distorted profiles of the positional system across whole eggs or isolates, recorded by these times, are retained to give correspondingly partial or imbalanced mes/endodermal pattern at tailbud larval stages. There is evidence that subsequent ‘back-up’ positional interactions, which can heal gross positional discontinuities in isolated presumptive lateral half-eggs and so restore bilateral symmetry, also do this at the price of loss of complete pattern specification. This is probably because of an asymmetrical principle whereby relatively activated (dorsoanterior specified) material can raise the level of originally posterior material on contact, whereas the reverse interaction cannot occur. The observations are discussed in relation to apparently different behaviour in certain other amphibian embryos, and to our knowledge of other positional interactions, normal and also experimentally provoked, such as those that set up the germ layers.

scholarly journals Calcium-free vitrification reduces cryoprotectant-induced zona pellucida hardening and increases fertilization rates in mouse oocytes

Reproduction ◽  
2006 ◽  
Vol 131 (1) ◽  
pp. 53-61 ◽  
Author(s):  
Mark G Larman ◽  
Courtney B Sheehan ◽  
David K Gardner
Keyword(s):  
Ethylene Glycol ◽  
Zona Pellucida ◽  
Cell Number ◽  
Blastocyst Stage ◽  
Initial Increase ◽  
Cell Stage ◽  
Free Treatment ◽  
Zona Hardening ◽  
Free Media ◽  
Oocyte Freezing

Despite the success of embryo cyropreservation, routine oocyte freezing has proved elusive with only around 200 children born since the first reported birth in 1986. The reason for the poor efficiency is unclear, but evidence of zona pellucida hardening following oocyte freezing indicates that current protocols affect oocyte physiology. Here we report that two cryoprotectants commonly used in vitrification procedures, dimethyl sulfoxide (DMSO) and ethylene glycol, cause a large transient increase in intracellular calcium concentration in mouse metaphase II (MII) oocytes comparable to the initial increase triggered at fertilization. Removal of extracellular calcium from the medium failed to affect the response exacted by DMSO challenge, but significantly reduced the ethylene glycol-induced calcium increase. These results suggest that the source of the DMSO-induced calcium increase is solely from the internal calcium pool, as opposed to ethylene glycol that causes an influx of calcium across the plasma membrane from the external medium. By carrying out vitrification in calcium-free media, it was found that zona hardening is significantly reduced and subsequent fertilization and development to the two-cell stage significantly increased. Furthermore, such calcium-free treatment appears not to affect the embryo adversely, as shown by development rates to the blastocyst stage and cell number/allocation. Since zona hardening is one of the early activation events normally triggered by the sperm-induced calcium increases observed at fertilization, it is possible that other processes are negatively affected by the calcium rise caused by cryoprotectants used during oocyte freezing, which might explain the current poor efficiency of this technique.

scholarly journals Mutator Foci Are Regulated by Developmental Stage, RNA, and the Germline Cell Cycle in Caenorhabditis elegans

2020 ◽  
Vol 10 (10) ◽  
pp. 3719-3728 ◽  
Author(s):  
Celja J. Uebel ◽  
Dana Agbede ◽  
Dylan C. Wallis ◽  
Carolyn M. Phillips
Keyword(s):  
Cell Cycle ◽  
Caenorhabditis Elegans ◽  
Epigenetic Inheritance ◽  
Germline Cell ◽  
Late Pachytene ◽  
Larval Stages ◽  
Transcriptional Inhibition ◽  
Male Germline ◽  
Early Embryos ◽  
Adult Hermaphrodite

RNA interference is a crucial gene regulatory mechanism in Caenorhabditis elegans. Phase-separated perinuclear germline compartments called Mutator foci are a key element of RNAi, ensuring robust gene silencing and transgenerational epigenetic inheritance. Despite their importance, Mutator foci regulation is not well understood, and observations of Mutator foci have been largely limited to adult hermaphrodite germlines. Here we reveal that punctate Mutator foci arise in the progenitor germ cells of early embryos and persist throughout all larval stages. They are additionally present throughout the male germline and in the cytoplasm of post-meiotic spermatids, suggestive of a role in paternal epigenetic inheritance. In the adult germline, transcriptional inhibition results in a pachytene-specific loss of Mutator foci, indicating that Mutator foci are partially reliant on RNA for their stability. Finally, we demonstrate that Mutator foci intensity is modulated by the stage of the germline cell cycle and specifically, that Mutator foci are brightest and most robust in the mitotic cells, transition zone, and late pachytene of adult germlines. Thus, our data defines several new factors that modulate Mutator foci morphology which may ultimately have implications for efficacy of RNAi in certain cell stages or environments.

scholarly journals Characterization of p96h2bk: immunoreaction with an anti-Erk(extracellular-signal-regulated kinase) peptide antibody and activity in Xenopus oocytes and eggs

1998 ◽  
Vol 335 (1) ◽  
pp. 43-50
Author(s):  
Dong-Hua CHEN ◽  
Chin-Tin CHEN ◽  
Yong ZHANG ◽  
Mei-Ann LIU ◽  
Roberto CAMPOS-GONZALEZ ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Xenopus Oocytes ◽  
Protein Kinase Activity ◽  
Cell Stage ◽  
Extracellular Signal Regulated Kinase ◽  
Oncogenic Ras ◽  
Cycle Arrest ◽  
Extracellular Signal ◽  
M Phase

We have shown previously that oncogenic Ras induces cell cycle arrest in activated Xenopus egg extracts [Pan, Chen and Lin (1994) J. Biol. Chem. 269, 5968–5975]. The cell cycle arrest correlates with the stimulation of a protein kinase activity that phosphorylates histone H2b in vitro (designated p96h2bk) [Chen and Pan (1994) J. Biol. Chem. 269, 28034–28043]. We report here that p96h2bk is likely to be p96ram, a protein of approx. 96 kDa that immunoreacts with a monoclonal antibody (Mk-1) raised against a synthetic peptide derived from a sequence highly conserved in Erk1/Erk2 (where Erk is extracellular-signal-regulated kinase). This is supported by two lines of evidence. First, activation/inactivation of p96h2bk correlates with upward/downward bandshifts of p96ram in polyacrylamide gels. Secondly, both p96h2bk and p96ram can be immunoprecipitated by antibody Mk-1. We also studied the activity of p96h2bk/p96ram in Xenopus oocytes and eggs. p96h2bk/p96ram was inactive in stage 6 oocytes, was active in unfertilized eggs, and became inactive again in eggs after fertilization. Since stage 6 oocytes are at G2-phase of the cell cycle, unfertilized eggs arrest at M-phase and eggs exit M-phase arrest after fertilization, the results thus indicate that p96h2bk/p96ram activity is cell cycle dependent. Moreover, microinjection of oncogenic Ras into fertilized eggs at the one-cell stage arrests the embryos at the two-cell stage, and this induced arrest is correlated with an inappropriate activation of p96h2bk/p96ram. The data are consistent with the concept that inappropriate activation of p96h2bk/p96ram plays a role in the cell cycle arrest induced by oncogenic Ras.

Expression of the cell cycle control gene, cdc25, is constitutive in the segmental founder cells but is cell-cycle-regulated in the micromeres of leech embryos

Development ◽  
1995 ◽  
Vol 121 (9) ◽  
pp. 3035-3043 ◽  
Author(s):  
S.T. Bissen
Keyword(s):  
Cell Cycle ◽  
Cell Division ◽  
Cell Cycle Control ◽  
Control Gene ◽  
Cdc25 Phosphatase ◽  
Cell Cycles ◽  
Cell Divisions ◽  
Zygotic Transcription ◽  
Founder Cells ◽  
Drosophila Embryos

The identifiable cells of leech embryos exhibit characteristic differences in the timing of cell division. To elucidate the mechanisms underlying these cell-specific differences in cell cycle timing, the leech cdc25 gene was isolated because Cdc25 phosphatase regulates the asynchronous cell divisions of postblastoderm Drosophila embryos. Examination of the distribution of cdc25 RNA and the zygotic expression of cdc25 in identified cells of leech embryos revealed lineage-dependent mechanisms of regulation. The early blastomeres, macromeres and teloblasts have steady levels of maternal cdc25 RNA throughout their cell cycles. The levels of cdc25 RNA remain constant throughout the cell cycles of the segmental founder cells, but the majority of these transcripts are zygotically produced. Cdc25 RNA levels fluctuate during the cell cycles of the micromeres. The levels peak during early G2, due to a burst of zygotic transcription, and then decline as the cell cycles progress. These data suggest that cells of different lineages employ different strategies of cell cycle control.

Maternal beta-catenin establishes a ‘dorsal signal’ in early Xenopus embryos

Development ◽  
1996 ◽  
Vol 122 (10) ◽  
pp. 2987-2996 ◽  
Author(s):  
C. Wylie ◽  
M. Kofron ◽  
C. Payne ◽  
R. Anderson ◽  
M. Hosobuchi ◽  
...  
Keyword(s):  
Glycogen Synthase ◽  
Axis Formation ◽  
Beta Catenin ◽  
Cell Stage ◽  
Midblastula Transition ◽  
Xenopus Embryos ◽  
Ability To Act ◽  
Zygotic Transcription ◽  
Spatial Terms ◽  
Marginal Zones

In previous work, we demonstrated that maternally encoded beta-catenin, the vertebrate homolog of armadillo, is required for formation of dorsal axial structures in early Xenopus embryos (Heasman, J., Crawford, A., Goldstone, K., Garner-Hamrick, P., Gumbiner, B., Kintner, C., Yoshida-Noro, C. and Wylie, C. (1994). Cell 79, 791–803). Here we investigated, firstly, the role(s) of beta-catenin in spatial terms, in different regions of the embryo, by injecting beta-catenin mRNA into individual blastomeres of beta-catenin-depleted embryos at the 32 cell stage. The results indicate that beta-catenin can rescue the dorsal axial structures in a non-cell-autonomous way and without changing the fates of the injected cells. This suggests that cells overexpressing beta-catenin send a ‘dorsal signal’ to other cells. This was confirmed by showing that beta-catenin overexpressing animal caps did not cause wild-type caps to form mesoderm, but did cause isolated beta-catenin-deficient marginal zones to form dorsal mesoderm. Furthermore beta-catenin-deficient vegetal masses treated with overexpressing caps regained their ability to act as Nieuwkoop Centers. Secondly, we studied the temporal activity of beta-catenin. We showed that zygotic transcription of beta-catenin starts after the midblastula transition (MBT), but does not rescue dorsal axial structures. We further demonstrated that the vegetal mass does not release a dorsal signal until after the onset of transcription, at the midblastula stage, suggesting that maternal beta-catenin protein is required at or before this time. Thirdly we investigated where, in relationship to other gene products known to be active in axis formation, beta-catenin is placed. We find that BVg1, bFGF, tBR (the truncated form of BMP2/4R), siamois and noggin activities are all downstream of beta-catenin, as shown by the fact that injection of their mRNAs rescues the effect of depleting maternally encoded beta-catenin. Interference with the action of glycogen synthase kinase (GSK), a vertebrate homolog of the Drosophila gene product, zeste white 3 kinase, does not rescue the effect, suggesting that it is upstream.

Tetraploidy and early development: effects on developmental timing and embryonic metabolism

Development ◽  
1981 ◽  
Vol 66 (1) ◽  
pp. 91-108
Author(s):  
Martin A. Eglitis ◽  
Lynn M. Wiley
Keyword(s):  
Cell Surface ◽  
Surface Antigen ◽  
Gene Dosage ◽  
Genetic Material ◽  
Cell Number ◽  
Embryonic Cell ◽  
Cell Surface Antigen ◽  
Metabolic Effects ◽  
Cell Stage ◽  
Preimplantation Mouse

The effect of balanced gene dosage changes on the timing of cavitation and on the timing of appearance of a stage-specific embryonic cell surface antigen was studied in preimplantation mouse embryos. Gene dosage was increased by creating tetraploid embryos at the 4-cell stage, either by blastomere fusion with polyethylene glycol (PEG) or by incubation in cytochalasin B (cytB) to block cell division. Removal of the zona pellucida with Pronase from diploid embryos caused a 7 h delay in cavitation. Further manipulations, either with PEG or cytB to induce tetraploidy, did not produce a statistically significant additional delay in cavitation timing. Likewise, PEG-induced tetraploidy did not affect the timing of appearance or disappearance of the embryonic cell surface antigen as compared with diploid control embryos. In analysing the metabolic effects of tetraploidy, we found that in tetraploid embryos with cell number equivalent to intact diploid embryos, MDH activity did not double with the doubling of the genome, being only 50% greater than diploid levels in cytB-induced tetraploid embryos and only 20% greater than diploid levels in PEG-induced tetraploid embryos. However, in tetraploid embryos with one-half normal cell number, enzyme activity was equal to that in whole diploid embryos, suggesting that in such embryos, MDH activity increased in parallel with increases in gene dosage. Further studies showed that levels of RNA synthesis in PEGinduced etraploid embryos also did not increase in parallel with the doubling of the genome. Rather, these results suggested that in tetraploid embryos, compensation was made for at least part of the excess genetic material.

DNA replication and compaction in the cleaving embryo of the mouse

Development ◽  
1985 ◽  
Vol 89 (1) ◽  
pp. 133-148
Author(s):  
Roger K. W. Smith ◽  
Martin H. Johnson
Keyword(s):  
Dna Replication ◽  
Dna Polymerase ◽  
Mouse Embryo ◽  
Cell Number ◽  
Reversible Inhibitor ◽  
Cell Stage ◽  
Dna Polymerase Alpha ◽  
Surface Polarization ◽  
Continuous Presence ◽  
Almost All

The effects of aphidicolin, a reversible inhibitor of DNA polymerase alpha, both on replication and on development of the mouse embryo from the 2- and 4-cell stages to the compacted late 8-cell stage have been assessed. The continuous presence of aphidicolin from G1 of the 4-cell stage resulted in inhibition of DNA replication and prevention of division from 4 to 8 cells, but was without effect on the timing or incidence of cell flattening, surface polarization and cytoplasmic polarization. The continuous presence of aphidicolin from G1 of the 2-cell stage resulted in inhibition of DNA replication, division, and polarization. Some slight intercellular flattening in a few embryos did occur. If addition of aphidicolin was delayed by 10 h to early in G2 of the 2-cell stage, further rounds of replication were blocked and some embryos failed to cleave to 4-cells. Nevertheless, almost all embryos showed evidence of flattening and polarization regardless of cell number. In contrast, if aphidicolin was added in G1 of the 2-cell stage and removed after 10 h, the cells showed delayed DNA replication, little evidence of division, and no cell flattening or polarization. We conclude that DNA replication at the 2-cell stage may be essential for the components of compaction studied, but that DNA replication at the 4- and 8-cell stages is not.

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