[Visualization of ribosomal genes transcription in SPEV culture cells using bromouridine triphosphate]

We applied a sensitive and specific method for detection of run-on rDNA transcription in cultured mammalian cells. This technique is based on the capability of RNA polymerase I to maintain transcriptional activity following cell fixation with methanol, and on the use of BrUTP as a precursor of rRNA synthesis. The results obtained have shown that in cultured pig cells (PK cells) the ribosomal genes are transcribed during interphase to become repressed at the end of mitotic prophase. The rDNAs are not transcribed at the prometaphase, metaphase and anaphase stages. The ribosomal genes become derepressed at early telophase. At early telophase, the number of BrUTP-incorporated sites is equal to that of the nucleolus organizing regions (NORs), but it is augmented during telophase progression. A similar dynamics of ribosomal gene reactivation is also revealed following spatial separation of NO-chromosomes between individual micronuclei caused by hypotonic chock. This indicates that the spatial integration of chromosomal NORs is not a prerequisite for ribosomal gene reactivation at mitosis.