Detailed analysis of pronucleus development in bovine zygotes in vivo: Ultrastructure and cell cycle chronology

Laurinčík, Jozef; Kopečný, V.; Hyttel, Poul

doi:10.1002/(sici)1098-2795(199601)43:1<62::aid-mrd8>3.3.co;2-9

Cited by 6 publications

(5 citation statements)

References 28 publications

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“…The chromatin-like cap of the latter clusters observed in the electron microscope could correspond to the rRNA gene clusters and associated heterochromatin. In this light, the affinity between the NPBs and these granule clusters as previously described [11,12] seems logical. At the ultrastructural level, however, the granule clusters are observed already from the first cell cycle, whereas the rRNA hybridization signal in the present experiment did not appear until the third cell cycle.…”

Section: Discussionmentioning

confidence: 58%

“…It has, however, not been possible to reveal whether this RNA synthesis also includes the rRNA genes. In this context, ultrastructural studies have been of only limited value, since the functional significance of the minor morphological changes of the nucleolus precursor bodies (NPBs) that are observed over the first, second, and third cell cycles of the bovine embryo is unclear [5,6,11,12].…”

Section: Introductionmentioning

confidence: 93%

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Ribosomal Ribonucleic Acid Is Transcribed at the 4-Cell Stage in In Vitro-Produced Bovine Embryos1

Viuff

Hyttel

Avery

et al. 1998

Biology of Reproduction

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Ribosomal RNA, rRNA genes, and silver-staining nucleolar proteins were visualized in in vitro-produced bovine embryos from the 2-cell stage to the blastocyst using a sequential fluorescent in situ hybridization (FISH) and a silver-staining procedure. At FISH, the rRNA was differentiated from the signal of the rRNA genes through comparison of RNase- and non-RNase-treated embryos. Both RNase- and non-RNase-treated 2-cell embryos revealed up to 10 small clusters of fluorescein isothiocynate (FITC) labeling in interphase nuclei. The RNase-treated 4-cell embryos displayed the same FITC pattern as the 2-cell embryos. In the non-RNase-treated 4-cell embryos, in contrast, the clusters were larger and included numerous small spots. In 2-cell as well as 4-cell embryos, almost all FITC-labeled clusters colocalized with silver-stained spots. In the RNase-treated 8- to 16-cell embryos, up to 10 clusters of FITC labeling were organized as one or more large spots surrounding a central faint but homogeneously labeled area. The non-RNase-treated 8- to 16-cell embryos displayed similar complexes, but the central areas consisted of small labeled spots. In 8- to 16-cell embryos, all FITC-labeled clusters were again colocalized with silver-stained areas. In the blastocysts, 1-6 big clusters of FITC labeling colocalized with silver staining. In the RNase-treated blastocysts, the FITC labeling was typically located at the edges of the silver-stained areas, whereas in the non-RNase-treated blastocysts, the FITC labeling totally covered the silver-stained areas. In conclusion, there is a close association between the rRNA genes and silver-staining nucleolar proteins in in vitro-produced bovine embryos from the second cell cycle, i.e., the 2-cell stage; the first rRNA is apparently transcribed during the third cell cycle, and during the fourth cell cycle the molecular composition of functional nucleoli is established.

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Section: Discussionmentioning

confidence: 58%

Section: Introductionmentioning

confidence: 93%

Ribosomal Ribonucleic Acid Is Transcribed at the 4-Cell Stage in In Vitro-Produced Bovine Embryos1

Viuff

Hyttel

Avery

et al. 1998

Biology of Reproduction

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“…On other locations in the pronuclei, clusters of electron-dense granules resembling interchromatin granules appear. During the S and G2 phases of the first cell cycle, the fibrillar spheres and the GCs become spatially associated, and at least in in vivo-derived bovine embryos the fibrillar spheres develop a central vacuole [41], which is less common in in vitro-derived zygotes [22].…”

Section: Discussionmentioning

confidence: 99%

Immunolocalization of Nucleolar Proteins During Bovine Oocyte Growth, Meiotic Maturation, and Fertilization1

Fair

Hyttel²,

Lonergan

et al. 2001

Biology of Reproduction

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During the growth phase of the bovine oocyte transcripts, polypeptides and ribosomes are accumulated in the oocyte to drive and sustain future meiotic maturation, fertilization, and early embryonic development. The oocyte also furnishes the early embryo with the components required to establish a functional transcriptionally active nucleolus at the time of maternal embryonic transition. The aim of the present study was to describe the behavior of key components of the nucleolus. The temporal localization of nucleolar proteins fibrillarin, nucleophosmin, nucleolin, RNA polymerase I (RNA pol I), topoisomerase I, upstream binding factor (UBF), and coilin 5P10 was investigated in growing and fully grown immature bovine oocytes during in vitro maturation and during the first postfertilization cell cycle using whole-mount immunocytochemistry and confocal microscopy. During the oocyte growth phase, fibrillarin, nucleophosmin, nucleolin, RNA pol I, and UBF were localized to the oocyte nucleolus. On completion of the growth phase, nucleolin and nucleophosmin appeared to migrate to the periphery of the nucleolus and into the nucleoplasm, and the proportion of oocytes displaying RNA pol I localization had decreased. Topoisomerase I was not detected at any stage. Fibrillarin appeared to be localized to large foci within the nucleolus and/or nucleoplasm. Nucleophosmin and nucleolin labeling was characterized by a homogeneous signal over the nucleolus. RNA pol I and UBF were characterized by the localization of the antibodies to individual or clustered foci in the nucleolus and/or nucleoplasm. Following oocyte nucleus breakdown (ONBD), the proteins appeared to disperse into the cytoplasm. All proteins were undetectable during meiotic maturation and were not relocalized until 5-10 h postinsemination (hpi). UBF was localized to the fertilizing sperm head of most zygotes at 5 hpi. By 10 hpi, all proteins were detected in most oocytes displaying two pronuclei. Nucleolar protein localization was exclusive to or more abundant in one pronucleus up to 20 hpi; thereafter, the pattern was more evenly distributed. Fibrillarin, nucleophosmin, nucleolin, UBF, and Pol I are present in the nuclei of growing and fully grown bovine oocytes until ONBD. They reappear at the late telophase stage of meiosis II and continue to be present up to the first mitotic division of embryo development.

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“…In the present work we were able to show EB-PTA stained granules within the vacuolised NPB which indicate RNP dynamics in NPBs. Intensely contrasted granules of the same diameter were seen in bovine pronuclei NPBs (Laurincˇík et al, 1996;Fig. 5) and were also observed in goat embryo NPBs (unpublished results).…”

Section: Nucleolus Precursor Body (Npb)mentioning

confidence: 54%

“…As in other mammalian embryos, the nucleolus develops in the bovine preimplantation embryo stepwise during cleavage, sterically related to its precursor, the NPB, by a defined morphological process (Camous et al, 1986;King et al, 1988;Kopecˇný et al, 1989a, b;Laurincˇík et al, 1996Laurincˇík et al, , 1998Kanka et al, 1999). It is interesting to note that this process is reversible, as shown by nuclear transfer experiments (Kanka et al, 1999), and the status of the NPB is thus evidently under cytoplasmic control.…”

Section: Nucleolus Precursor Body (Npb)mentioning

confidence: 92%

Fine-structural cytochemical and immunocytochemical observations on nuclear bodies in the bovine 2-cell embryo

Kopečný

Biggiogera

Pivko

et al. 2000

Zygote

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Nuclear bodies occuring during the 2-cell stage of bovine embryos (obtained either by in vitro fertilisation of in vitro matured ovarian oocytes, or collection after fertilisation and cleavage in vivo) were studied using ultrastructural cytochemistry and immunocytochemistry to determine whether their occurrence may be linked with the onset of embryonic transcription. In addition, the species-specific ultrastructural features of the interchromatin structures of the 2-cell bovine embryo were displayed. Three different types of nuclear bodies were distinguished: (i) nucleolus precursor bodies (NPBs), (ii) loose bodies (LBs) and (iii) dense bodies (DBs). In order to determine their possible functional significance, we considered parallels between these three nuclear entities and interchromatin compartments reported in other cells. As detected by their preferential ribonucleoprotein staining, all types of nuclear bodies contained ribonucleoproteins. In contrast to the other types of nuclear bodies studied, NPBs contained argyrophilic proteins but in no case they did show morphological features of functional nucleoli. Both compact and vacuolated forms of NPBs were seen in both in vivo and in vitro embryos, sometimes simultaneously in the same nucleus. LBs and DBs reacted with antibodies to Sm antigen, indicating the presence of a group of nucleoplasmic, non-nucleolar small nuclear ribonucleoproteins (snRNPs). The immunoreactivity for Sm antigen was more intense and homogeneous in DBs than in LBs. DBs were seen in both categories of embryo. A possible kinship of DBs with the sphere organelle known from oocytes of different animal species or the prominent spherical inclusions of the early mouse embryo nuclei is suggested. The last type of intranuclear body, the LBs, showed a composite structure. Their granular component, occurring in clusters and displaying immunoreactivity for Sm antigen, was similar to interchromatin granules and was therefore named IG-like granules. Another component forming the LBs showed a much finer structure and a lower immunoreactivity with anti-Sm antibodies. We suggest that this amorphous component may be related to the IG-associated zone. All three types of intranuclear bodies were often seen close together, suggesting their possible mutual functional relationship. From these and other observations we conclude that the intranuclear bodies in 2-cell bovine embryos correspond, with the exception of the NPB, to similar structures/compartments supposed to accumulate inactive spliceosomal components in certain phases of somatic cell nucleus functions. Accordingly, the occurrence of such nuclear bodies does not represent cytological evidence for RNA synthesis. In contrast to this, an important morphological feature revealing the status of the bovine 2-cell embryo is the vacuol-isation of the NPB.

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Detailed analysis of pronucleus development in bovine zygotes in vivo: Ultrastructure and cell cycle chronology

Cited by 6 publications

References 28 publications

Ribosomal Ribonucleic Acid Is Transcribed at the 4-Cell Stage in In Vitro-Produced Bovine Embryos1

Ribosomal Ribonucleic Acid Is Transcribed at the 4-Cell Stage in In Vitro-Produced Bovine Embryos1

Immunolocalization of Nucleolar Proteins During Bovine Oocyte Growth, Meiotic Maturation, and Fertilization1

Fine-structural cytochemical and immunocytochemical observations on nuclear bodies in the bovine 2-cell embryo

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