Nucleolar substructures of rabbit cleaving embryos: An immunocytochemical study

Abstract

The structure‐function relationships of the nucleolar substructures were studied in preimplantation rabbit embryos, where nucleologenesis is extending over the first four cell cycles and may not be synchronous in each blastomere. Immunocytochemical methods using light and electron microscopy were applied for protein and RNA localization as well as nick translation and terminal deoxynucleotidyl transferase techniques for DNA detection. DNA was gradually associated with the periphery of the compact nucleolar precursor bodies (NPBs) but was never found inside NPBs at the four‐cell stage. In 16‐cell embryos, some NPBs displayed a reticulated periphery forming the branching network of the dense fibrillar component (DFC) surrounding the “residual body” (remnant of NPB) in the process of activation. At the 32‐cell stage, fully reticulated nucleoli were observed in each blastomere. DNA was then associated with the DFC of reticulated nucleoli. RNA was first detected at the 16 cell‐stage in close contact with the DFC as well as inside the “residual body” which was not immunolabeled with the DNA antibodies used. When observed by light microscopy, fibrillarin, nucleolin, and protein B23 displayed a changing distribution pattern during nucleologenesis. At early stages (up to the 16‐cell stage), small dot‐ and spot‐like structures were distributed within the whole nuclei. In 16‐cell embryos, these proteins started to accumulate in an irregular thin layer around the NPBs in the process of activation. The reorganization process described may be in relation with the redistribution of chromatin and nuclear/nucleolar matrix components during the activation of rDNA transcription localized in the NPB shell. In conclusion, nucleologenesis is only achieved at the fourth cell cycle in the cleaving rabbit embryo at the corresponding time when the first detectable nucleolus‐associated RNA is detectable. Our results show a good correlation between the establishment of structure and function. Mol. Reprod. Dev. 48:34–44, 1997. © 1997 Wiley‐Liss, Inc.

Impairments in Embryonic Genome Activation in Rhesus Monkey Somatic Cell Nuclear Transfer Embryos

“…The onset of fibrillarin expression mirrored that of tritiated uridine incorporation into the nucleolus, first appearing in sixto eight-cell stage macaque embryos (Schramm and Bavister, 1999). It was also reported that fibrillarin acts as a genome activation marker in mice (Baran et al, 1995;Cuadros-Fernandez and Esponda, 1996), rabbits (Baran et al, 1997;PintoCorreia et al, 1995), and cows (Schram and Paprocki, 2000). In the present study, transcripts of fibrillarin, nucleolin, nucleophosmin, PAF53, and UBF were detected in all blastocysts.…”

DNA topoisomerase II distribution in mouse preimplantation embryos

“…This was con®rmed by co-localization with ®brillarin, a 34 kDa component of the ®brillar center and dense ®brillar component of the nucleolus (Ochs et al, 1985). We found that ®brillarin staining in the nucleolar periphery of 2-to 8-cell mouse embryos is similar to previous studies that show this staining pattern in mouse, rabbit, porcine, bovine, and monkey embryos, and actively growing somatic cell lines (Jimenez-Garcia et al, 1989;Ochs et al, 1994;Baran et al, 1995;Baran et al, 1996;Cuadros-Ferna  ndez and Esponda, 1996;Baran et al, 1997;Schramm and Bavister, 1999;Hyttel et al, 2000;Laurincik et al, 2000). In addition, the lack of MER1 antibody staining in nucleoli of mouse embryos (except pronucleate embryos) indicates topo IIb was absent from nucleoli of preimplantation embryos up to the blastocyst stage.…”

The nature of the ‘nucleolus precursor body’ in early preimplantation embryos: a review of fine-structure cytochemical, immunocytochemical and autoradiographic data related to nucleolar function

“…There is also general agreement that the core of the mouse-type NPB does not contain DNA either before or during the onset of rRNA transcription and nucleolonema differentiation (Faken & Odartchenko, 1980;Baran et al, 1993;Biggiogera et al, 1994;Kopečný et al, 1995). The absence of DNA was also confirmed recently in the rabbit NPB by the anti-DNA labelling and terminal deoxyinucleotidyl transferase techniques (Baran et al, 1997). The appearance of the nucleolonema and hence of rRNA transcription only at the surface of the NPB was also supported by immunoelectron microscope detection of typical nucleolar proteins exclusively at the NPB periphery (Baran et al, 1995;Cuadros-Fernandez & Esponda, 1996).…”