Localisation of RNAs and proteins in nucleolar precursor bodies of early mouse embryos

Abstract

Early embryos of all mammalian species contain morphologically distinct but transcriptionally silent nucleoli called the nucleolar precursor bodies (NPBs), which, unlike normal nucleoli, have been poorly studied at the biochemical level. To bridge this gap, here we examined the occurrence of RNA and proteins in early mouse embryos with two fluorochromes – an RNA-binding dye pyronin Y (PY) and the protein-binding dye fluorescein-5′-isothiocyanate (FITC). The staining patterns of zygotic NPBs were then compared with those of nucleolus-like bodies (NLBs) in fully grown surrounded nucleolus (SN)-type oocytes, which are morphologically similar to NPBs. We show that both entities contain proteins, but unlike NLBs, NPBs are significantly impoverished for RNA. Detectable amounts of RNA appear on the NPB surface only after resumption of rDNA transcription and includes pre-rRNAs and 28S rRNA as evidenced by fluorescence in situ hybridisation with specific oligonucleotide probes. Immunocytochemical assays demonstrate that zygotic NPBs contain rRNA processing factors fibrillarin, nucleophosmin and nucleolin, while UBF (the RNA polymerase I transcription factor) and ribosomal proteins RPL26 and RPS10 are not detectable. Based on the results obtained and data in the contemporary literature, we suggest a scheme of NPB assembly and maturation to normal nucleoli that assumes utilisation of maternally derived nucleolar proteins but of nascent rRNAs.