Formation of nuclear microarchitecture in the preimplantation bovine embryo at the onset of transcription: Implications for biotechnology

Abstract

Fine-structural and topochemical methods have revealed a highly ordered sequence of nuclear events in blastomeres of the early bovine embryo at the time when the embryo gains the capacity for the transcription of RNA. At the 8-cell stage, the DNA, in earlier stages localized at the periphery of the nucleus, is equally distributed throughout the nucleoplasm. At the same time, both nucleolar and extranucleolar RNA synthesis is resumed. The time sequence of this process is reflected by the ultrastructural morphology of nucleologenesis, during which the nucleolus precursor body (NPB) is transformed into a functional nucleolus as a result of the stepwise penetration by DNA and by changes in the extranucleolar chromatin. The most remarkable feature of differentiation is the chromatin organization in distinct blocks on the borderline of which the synthesis and maturation of extranucleolar RNA takes place. This is demonstrated by the localization of perichromatin fibrils and by detection of a class of small nuclear ribonucleoproteins (snRNPs) known to be involved in RNA splicing in that specific region. The conspicuous morphological characteristics of bovine embryonic nucleologenesis can be used as landmarks for the evaluation of the degree of nuclear differentiation under various experimental conditions. In embryos derived from different biotechnological approaches deviations from the normal pattern of differentiation in the nuclear microarchitecture were detected: (I) The condensed chromatin in 8-cell stages formed more prominent blocks in nuclei from embryos obtained by in vitro procedures than in those grown in vivo. (II) In embryos produced in vitro from oocytes isolated from different size categories of antral follicles the degree of extranucleolar chromatin condensation often did not reach that of ‘normal’ embryos. This finding was connected with a less intense RNA synthesis as detected by autoradiography. The NPB development was altered since its association with the perinuclear chromatin was less developed. (III) In embryos reconstructed after fusion of a blastomere with an enucleated oocyte, the development of the NPB structure as well as the RNA synthesis were arrested for three consecutive cleavages. (IV) The morphological features and in particular the autoradiographic data can be interpreted as the visualization of a consecutive switch-on of genes probably inversely related to the totipotency of a given nucleus and can thus be helpful in the assessment of nuclear transfer efficiency. Collectively, these findings suggest that embryos resulting from various biotechnological procedures can display distinct ultrastructural deviations at the level of nuclear function that could contribute to their lower developmental capacity.