Visualization of the dynamic behavior of ribosomal RNA gene repeats in living yeast cells

Abstract

The ribosomal RNA (rRNA) gene forms an extremely large repeat (rDNA) in the chromosome. In budding yeast, Saccharomyces cerevisiae, the rDNA is located on chromosome XII and occupies approximately 60% (1.5Mb) of the chromosome and 10% of the total genome. The rDNA is composed of approximately 150 copies and produces rRNA that accounts for approximately 80% of the total RNA. Owing to its repetitive structure and high transcription activity, rDNA is recombinogenic. Thus, the repeat is a distinctive and unique region in the genome. To investigate the dynamic nature of the repeat during the cell division cycle, we developed yeast strains in which every rDNA repeat unit has lacO or tetO arrays that associate with LacI-GFP or TetR-mRFP proteins, respectively. In these strains, the entire repeat can be visualized in a living cell without the need for any special treatment. Analyses of these strains showed that the rDNA actively moved and changed shape at the boundary between the nucleolus and the nucleoplasm. At the G2/M phase, the associated sister chromatids of the rDNA formed a ring structure in the nucleolus that gave the appearance of 'rotating'. We also found that the length of the rDNA repeat affected the time taken for segregation during the M phase of cell cycle. © 2011 The Authors. Journal compilation © 2011 by the Molecular Biology Society of Japan/Blackwell Publishing Ltd.