Activation of nucleolar and extranucleolar RNA synthesis and changes in the ribosomal content of human embryos developing in vitro

RNA synthetic activity of human 2-16-cell embryos developing in vitro was studied by [3H]uridine light-microscope autoradiography. Parallelly cut thin sections were examined in the electron microscope. The first extranucleolar RNA synthesis was detected in 4-cell embryos, but nucleoli were never labelled until the 3rd cleavage (6-8-cell embryos). In 6-cell embryos the nucleolar labelling was mostly confined to a narrow peripheral zone. In later cleavage stages most of the blastomeres showed intensive labelling of nucleoli and extranucleolar chromatin. However, rather low levels of extranucleolar RNA synthesis and the absence of nucleolar activity were often seen even in blastomeres of fully compacted morulae. The activation of nucleolar RNA synthesis entailed a noticeable increase in the number of ribosomes (estimated by electron microscope morphometry) that followed a marked drop during the period between the 2-cell and 8-cell stages. The results indicate that the concentration of ribosomes in the preovulatory oocyte is a major factor of its developmental potential.     

Nucleic acid synthesis in preimplantation mouse embryos

Summary Mouse embryos were collected at the 2-cell stage, cultured in vitro in the presence of³H deoxyuridine or uridine for 6 or 4 h and autoradiographed.Deoxyuridine is actively incorporated into the DNA of cleaving mouse embryos indicating the existence of thymidylate synthetase activity at least at the 4-cell stage and presumably already before this.RNAase treatment of embryos squashed on slides shows a weak but obvious incorporation of uridine into DNA of cleaving mouse embryos, from the 4-cell stage onwards; this incorporation is totally inhibited by hydroxyurea. The reduction of ribonucleotides to deoxyribonucleotides is a metabolic pathway already required for cleavage, as shown by hydroxyurea experiments.The second polar pody, known to incorporate thymidine, is unable to incorporate either deoxyuridine or uridine.     

Localization of nucleic acids in the nucleoli of oocytes and early embryos of mouse and hamster: An autoradiographic study

Mouse preovulatory oocytes, zygotes, parthenogenetically activated pronuclear oocytes, and early embryos, as well as hamster zygotes, were analyzed, by autoradiography, for the distribution of either “maternal” or newly synthesized RNAs. Early mouse embryos were also examined for the distribution of newly replicated DNA. Special attention was attributed to NLBs in oocytes or to NPBs in early embryos. In mouse oocytes, [5-³H]uridine radioactivity accumulated (after a 2-hr pulse) in vitro, in addition to other nuclear compartments, in the central compact material of the NLBs. There was no cytoplasmic labeling. In all parthenogenetic pronuclear embryos developed from similarly labeled oocytes, this label was distinctly detectable in the central compact material of the NPBs; less intensive labeling was seen in the nucleoplasm and cytoplasm. On the contrary, the central compact part of the mouse NPB did not show labeling in DNA after a continuous culture with [6-³H]thymidine. In mouse and hamster pronuclear zygotes, convincing evidence was obtained for a lack of any newly synthesized nucleic acids in the compact material of NPBs using 4- to 10-hr culture with [8-³H]adenosine. Based on these data, it was shown that the NLBs of oocytes or NPBs of early embryos probably contain RNAs synthesized during the last stages of antral follicle oocyte differentiation. This unique pathway of RNAs in the oocyte—embryo system may explain the specific morphology of both oocyte and early embryo “nucleoli.” © 1995 Wiley-Liss, Inc.