Nucleolar development and allocation of key nucleolar proteins require de novo transcription in bovine embryos
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The goal of the present study was to investigate whether key nucleolar proteins involved in ribosomal RNA (rRNA) transcription and processing are transcribed de novo or from maternally inherited messenger RNAs (mRNA) in bovine embryos, and to which extent de novo transcription of these proteins mRNA is required for the development of functional nucleoli during the major activation of the embryonic genome. Immunofluorescence for localization of key nucleolar proteins, autoradiography for detection of transcriptional activity, and transmission electron microscopy were applied to in vitro produc ed bovine embryos cultured from the 2-cell stage with or without (control groups) a-amanitin, which blocks the RNA plymerases II and III transcription and, thus the synthesis of mRNA. In the control groups, weak autoradiographic labelling was initially observed in the periphery of few nuclei at the 4-cell and the early 8-cell stage, and the entire nucleoplasm as well as nucleolus precursor bodies (NBBs) were prominently labelled in all late 8-cell stages. The NPBs displayed initial transformation into fibrillo-granular nucleoli. In the a-amanitin group, lack of autoradiographic labelling was seen at all developmental stages and disintegrated NPBs stage were found at the late 8-cell. Our immunofluorescence data indicate that RNA polymerase I, UBF, topoisomerase I and fibrillarin are transcribed de novo whereas nucleolin and nucleophosmin are maternally inherited as demonstrated by a-amanitin inhibition. However, localization of these two proteins to the nucleolar compartments was negatively affected by the a-aminitin treatment. Consequentl, functional nucleoli were not established.
|Journal||Molecular Reproduction and Development|
|Number of pages||8|
|Publication status||Published - 2007|
- LIFE - embryo, maternal-embryonic transition, nucleolus, gene expression, alpha-amanitin