NORs and their transcription competence during the cell cycle.
In human cells ribosomal genes are organized as clusters, NORs, situated on the short arms of acrocentric chromosomes. It was found that essential components of the RNA polymerase I transcription machinery, including UBF, can be detected on some NORs, termed "competent" NORs, during mitosis. The competent NORs are believed to be transcriptionally active during interphase. However, since individual NORs were not observed in the cell nucleus, their interphase status remains unclear. To address this problem, we detected the competent NORs by two commonly used methods, UBF immunofluorescence and silver staining, and combined them with FISH for visualization of rDNA and/or specific chromosomes. We found that the numbers of competent NORs on specific chromosomes were largely conserved in the subsequent cell cycles, with certain NOR-bearing homologues displaying a very stable pattern of competence. Importantly, those and only those NORs that were loaded with UBF incorporated bromo-uridine in metaphase after stimulation with roscovitine and in telophase, suggesting that competent and only competent NORs contain ribosomal genes transcriptionally active during interphase. Applying premature chromosome condensation with calyculin A, we visualized individual NORs in interphase cells, and found the same pattern of competence as observed in the mitotic chromosomes.
Smirnov
E
E
Institute of Cellular Biology and Pathology, 1st Faculty of Medicine, Charles University, Prague, Czech Republic. esmir@lfl.cuni.cz
Kalmárová
M
M
Koberna
K
K
Zemanová
Z
Z
Malínský
J
J
Masata
M
M
Cvacková
Z
Z
Michalová
K
K
Raska
I
I
D002453:Cell Cycle / Q000187:drug effects; D002877:Chromosomes, Human / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D007399:Interphase / Q000187:drug effects; D007621:Karyotyping; D008677:Metaphase / Q000187:drug effects; D009697:Nucleolus Organizer Region / Q000187:drug effects / Q000235:genetics*; D035463:Pol1 Transcription Initiation Complex Proteins / Q000378:metabolism; D011687:Purines / Q000494:pharmacology; D000077546:Roscovitine; D016622:Silver Staining; D013692:Telophase / Q000187:drug effects; D014158:Transcription, Genetic / Q000187:drug effects
D035463:Pol1 Transcription Initiation Complex Proteins; D011687:Purines; C063844:transcription factor UBF; D000077546:Roscovitine