The Effects of Ultraviolet Irradiation on Uncleaved Eggs of Xenopus Laevis

1960 ◽  
Vol s3-101 (55) ◽  
pp. 299-311
Author(s):  
J. B. GURDON
Keyword(s):  
Xenopus Laevis ◽  
Ultraviolet Irradiation ◽  
Sperm Nucleus ◽  
Nuclear Transplantation ◽  
Vitelline Membrane ◽  
Nuclear Marker ◽  
Small Doses ◽  
Transplantation Experiments ◽  
Better Than

The effects are described of ultraviolet (u.v.) irradiation upon the eggs of Xenopus laevis. The results obtained apply to fertilized eggs and also to unfertilized eggs into which blastula nuclei have been transplanted. Eggs were irradiated up to 20 min after laying, for periods varying from 15 to 50 sec. The egg nucleus is completely inactivated by small doses of u.v. If fertilized eggs are used this gives rise to haploids, which the use of a nuclear marker has shown to be androgenetic. After irradiation the egg nucleus descends towards the centre of the egg, and comes to lie adjacent to the transplanted or sperm nucleus. At the first mitosis, however, it does not fuse but remains as a pycnotic clump in the centre of the spindle. Soon after this it disappears, without disintegrating into visible fragments. The transplanted or sperm nucleus appears to be unaffected by the irradiation and death of the egg nucleus. The egg cytoplasm does not appear to be damaged, even after doses of u.v. which are considerably more than sufficient to kill the egg nucleus. The main reasons for this belief are that haploids obtained by other means develop no better than those obtained by u.v. An increase of u.v. treatment from 30 to 80 sec results in no increase in abnormalities sustained. The jelly is broken down and the vitelline membrane weakened. This enables the egg to be penetrated by a micropipette without causing damage or preventing healing. This investigation was undertaken to facilitate the analysis of nuclear transplantation experiments in Xenopus. The increased penetrability of eggs is of technical value for this purpose. The interpretation of these experiments is greatly facilitated by the knowledge that the egg cytoplasm need not be damaged by the u.v. and that the egg nucleus is completely inactivated so as not to interfere with the development of the egg.

Ploidy, pigment patterns and species specific antigenicity in interspecific nuclear transplantations in newts

Development ◽  
1967 ◽  
Vol 17 (2) ◽  
pp. 319-330
Author(s):  
F. Sládeček ◽  
A. Romanovský
Keyword(s):  
Xenopus Laevis ◽  
Functional Activity ◽  
Nuclear Transplantation ◽  
Skin Grafts ◽  
Species Specific ◽  
Transplantation Antigens ◽  
Transplantation Experiments ◽  
Pigment Patterns

It was shown by Simnett (1964) that in Xenopus laevis skin grafts in adult frogs between members of the same nuclear clone were tolerated in the same way as autografts, but in skin grafts made between individuals belonging to different nuclear clones a homograft rejection occurred. The nucleus is therefore responsible for the synthesis of specific transplantation antigens. It seemed to us useful to investigate the species-specific antigenicity of animals derived from eggs transplanted with foreign nuclei in correlation with their ploidy and with the development of their species-specific pigment patterns, as a proof of functional activity of transplanted nuclei. For this purpose we used two species of Triturus, T. vulgaris and T. alpestris, because of earlier studies carried out in our laboratory on the pigmentation of their hybrids (Romanovský & Ŝtefanová, 960; Mazáková-Štefanová, 1965) and on their species-specific antigenicity (Romanovský, 1962 a, b), in spite of the known difficulties and limitations of nuclear transplantation experiments in these species (Lehman, 1955; Sládeček & Mazáková-Štefanová, 1964, 1965).

scholarly journals The Developmental Capacity of Nuclei Taken from Differentiating Endoderm Cells of Xenopus laevis

Development ◽  
1960 ◽  
Vol 8 (4) ◽  
pp. 505-526
Author(s):  
J. B. Gurdon
Keyword(s):  
Xenopus Laevis ◽  
Rana Pipiens ◽  
Nuclear Transplantation ◽  
Blastula Stage ◽  
Developmental Capacity ◽  
Nuclear Changes ◽  
Nuclear Differentiation ◽  
Development Proceeds ◽  
Transplantation Experiments ◽  
Different Parts

An important question concerning embryonic differentiation is whether the nuclei of somatic cells in different parts of an embryo come to differ genetically from each other during development. It has become possible to investigate this matter since King & Briggs (1955) have shown that nuclear transplantation is a satisfactory technique for testing the developmental potentialities of embryonic nuclei. These authors (1957, 1960) have used Rana pipiens for transplantation experiments with endoderm nuclei, and have found that these nuclei become progressively limited in their developmental capacity after the late blastula stage. This paper describes some similar experiments carried out with endoderm nuclei of Xenopus laevis. The general conclusion that nuclei change as development proceeds is confirmed; there are, however, considerable differences between Rana and Xenopus in the rate and time of onset of nuclear changes. These differences make it easier to understand the significance of nuclear differentiation during embryonic development.

A Description of the Technique for Nuclear Transplantation in Xenopus laevis

Development ◽  
1960 ◽  
Vol 8 (4) ◽  
pp. 437-444
Author(s):  
T. R. Elsdale ◽  
J. B. Gurdon ◽  
M. Fischberg
Keyword(s):  
Xenopus Laevis ◽  
Rana Pipiens ◽  
Nuclear Transplantation ◽  
Laboratory Conditions ◽  
Technical Details ◽  
Comparison Of The Results ◽  
Transplantation Experiments

A Method by which nuclei can be successfully transplanted into Amphibian eggs was first worked out by Briggs & King (1952) for the eggs of Rana pipiens. Xenopus laevis is an atypical Anuran since its eggs can be obtained throughout the year, and the resulting embryos can be reared to maturity within 12 months under laboratory conditions. Because of these advantages we have used Xenopus for nuclear transplantation experiments. Though the principle of Briggs & King's technique has been followed, differences between the eggs of Rana and Xenopus have made it necessary to modify their technique before it can be satisfactorily applied to the eggs of Xenopus. The purpose of this publication is first to give technical details of these modifications, and secondly to discuss the extent to which they might affect a direct comparison of the results of transplantations in Rana and Xenopus.

Cyclic cytoplasmic activity of non-nucleate egg fragments of Xenopus controls the morphology of injected sperms

1983 ◽  
Vol 63 (1) ◽  
pp. 69-76
Author(s):  
M. Sakai ◽  
A. Shinagawa
Keyword(s):  
Xenopus Laevis ◽  
Sperm Nucleus ◽  
Cyclic Activity ◽  
Triton X

Triton X-100-treated sperms were injected into non-nucleate egg fragments of Xenopus laevis to determine whether the structure of the injected sperm nucleus is affected by the cyclic activity of the cytoplasm. Swollen vesicular nuclei were very frequently observed when the sperms were injected and incubated during the ‘rounding-up’ phase of the recipient fragment, whereas no such structures were found when they were incubated during the ‘relaxing’ phase.

Insulin receptors on Xenopus laevis oocytes: effects of injection of ob/ob mouse liver mRNA

1991 ◽  
Vol 100 (1) ◽  
pp. 167-171
Author(s):  
D.A. Diss ◽  
B.D. Greenstein
Keyword(s):  
Xenopus Laevis ◽  
Binding Sites ◽  
Insulin Receptors ◽  
Insulin Binding ◽  
Cell Types ◽  
Follicle Cells ◽  
Vitelline Membrane ◽  
Xenopus Laevis Oocytes ◽  
Endogenous Insulin ◽  
Order Of Magnitude

We describe here conditions for the detection of insulin binding sites on Xenopus laevis oocytes. The binding of 125I-labelled insulin displayed sigmoidal behaviour, which is characteristic of the binding relationship between insulin and its receptor. Resolution of the resulting curvilinear Scatchard plot into two components revealed KD values of 8.86 × 10(−10) +/− 1.9 × 10(−10) and 5.32 × 10(−9) +/− 2.4 × 10(−9) M and n values of 9.7 × 10(7) +/− 0.4 × 10(7) and 3.3 × 10(8) +/− 0.5 × 10(8) binding sites per oocyte, respectively. The possibility cannot be excluded, however, that receptors for IGF-1 were also being detected. Also described are conditions for the rapid and efficient removal of all tissues surrounding the oocyte, including the vitelline membrane. We could not detect any specific 125I-labelled insulin binding to oocytes that had their follicle cells or vitelline membrane removed and this was not due to the enzymic treatment used in the process. Microinjection of oocytes without follicular layers did not result in the appearance of any detectable insulin binding sites, which were, however, observed if oocytes were first stripped of the vitelline membrane. We suggest that oocytes may possess endogenous insulin receptors on their surface in numbers of the same order of magnitude as those present on somatic cells. The removal of tissues surrounding the oocyte should facilitate studies aimed at determining functional interactions of the various cell types during oocyte development and for studying insulin receptors on the oocyte-follicular cell complex.

Factors Responsible for the Abnormal Development of Embryos Obtained by Nuclear Transplantation in Xenopus laevis

Development ◽  
1960 ◽  
Vol 8 (3) ◽  
pp. 327-340
Author(s):  
J. B. Gurdon
Keyword(s):  
Xenopus Laevis ◽  
Embryo Development ◽  
Normal Development ◽  
Nuclear Transplantation ◽  
Abnormal Development ◽  
Donor Nucleus ◽  
Innate Capacity

In Xenopus the embryos derived from nuclear transplantation often develop abnormally. These abnormalities must be due to the limited potentiality for development of either the donor nucleus or the egg cytoplasm; this limited potentiality will in turn be due to technical damage during transplantation or to the innate condition of the nucleus or cytoplasm before the experiment. The extent to which these technical and innate factors are responsible for abnormalities of transplant-embryo development has been analysed by considering the effect of each factor in turn. Nuclei from early donor stages have been used, since these nuclei are believed to be undifferentiated (see p. 338) and therefore to have the innate capacity for entirely normal development. The effects of other factors have been investigated by experiments in which each factor is varied in different ways. Any correlation between variations in one factor and the resulting proportion of abnormal transplant-embryos is then recorded.

scholarly journals Roles of LAP2 Proteins in Nuclear Assembly and DNA Replication: Truncated LAP2β Proteins Alter Lamina Assembly, Envelope Formation, Nuclear Size, and DNA Replication Efficiency in Xenopus laevis Extracts

1999 ◽  
Vol 144 (6) ◽  
pp. 1083-1096 ◽  
Author(s):  
Tracey Michele Gant ◽  
Crafford A. Harris ◽  
Katherine L. Wilson
Keyword(s):  
Dna Replication ◽  
Xenopus Laevis ◽  
Chromatin Binding ◽  
Binding Region ◽  
Nuclear Assembly ◽  
Downstream Effector ◽  
Common Domain ◽  
Splicing Isoforms ◽  
High Concentrations ◽  
Better Than

Humans express three major splicing isoforms of LAP2, a lamin- and chromatin-binding nuclear protein. LAP2β and γ are integral membrane proteins, whereas α is intranuclear. When truncated recombinant human LAP2β proteins were added to cell-free Xenopus laevis nuclear assembly reactions at high concentrations, a domain common to all LAP2 isoforms (residues 1–187) inhibited membrane binding to chromatin, whereas the chromatin- and lamin-binding region (residues 1–408) inhibited chromatin expansion. At lower concentrations of the common domain, membranes attached to chromatin with a unique scalloped morphology, but these nuclei neither accumulated lamins nor replicated. At lower concentrations of the chromatin- and lamin-binding region, nuclear envelopes and lamins assembled, but nuclei failed to enlarge and replicated on average 2.5-fold better than controls. This enhancement was not due to rereplication, as shown by density substitution experiments, suggesting the hypothesis that LAP2β is a downstream effector of lamina assembly in promoting replication competence. Overall, our findings suggest that LAP2 proteins mediate membrane–chromatin attachment and lamina assembly, and may promote replication by influencing chromatin structure.

Nuclear Transplantation in Xenopus laevis

Nature ◽  
1958 ◽  
Vol 181 (4606) ◽  
pp. 424-424 ◽  
Author(s):  
M. FISCHBERG ◽  
J. B. GURDON ◽  
T. R. ELSDALE
Keyword(s):  
Xenopus Laevis ◽  
Nuclear Transplantation

scholarly journals Direct somatic lineage conversion

2015 ◽  
Vol 370 (1680) ◽  
pp. 20140368 ◽  
Author(s):  
Koji Tanabe ◽  
Daniel Haag ◽  
Marius Wernig
Keyword(s):  
Transcription Factors ◽  
Cell Types ◽  
Nuclear Transplantation ◽  
Specific Cell ◽  
Direct Reprogramming ◽  
Cell Type ◽  
Mesodermal Cell ◽  
Related Cell ◽  
Transplantation Experiments ◽  
Cell Specialization

The predominant view of embryonic development and cell differentiation has been that rigid and even irreversible epigenetic marks are laid down along the path of cell specialization ensuring the proper silencing of unrelated lineage programmes. This model made the prediction that specialized cell types are stable and cannot be redirected into other lineages. Accordingly, early attempts to change the identity of somatic cells had little success and was limited to conversions between closely related cell types. Nuclear transplantation experiments demonstrated, however, that specialized cells even from adult mammals can be reprogrammed into a totipotent state. The discovery that a small combination of transcription factors can reprogramme cells to pluripotency without the need of oocytes further supported the view that these epigenetic barriers can be overcome much easier than assumed, but the extent of this flexibility was still unclear. When we showed that a differentiated mesodermal cell can be directly converted to a differentiated ectodermal cell without a pluripotent intermediate, it was suggested that in principle any cell type could be converted into any other cell type. Indeed, the work of several groups in recent years has provided many more examples of direct somatic lineage conversions. Today, the question is not anymore whether a specific cell type can be generated by direct reprogramming but how it can be induced.

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