Visualization of the dynamic behavior of ribosomal RNA gene repeats in living yeast cells
Visualization of the dynamic behavior of ribosomal RNA gene repeats in living yeast cells
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DOI：
10.1111/j.1365-2443.2011.01506.x
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发表时间：
2011-05-01
期刊：
影响因子：
2.1
通讯作者：
Kobayashi, Takehiko
中科院分区：
文献类型：
--
作者：
Miyazaki, Takaaki;Kobayashi, Takehiko
核糖体RNA(RRNA)基因在染色体上形成一个极大的重复(RDNA)。在芽生酵母中，rDNA位于第12号染色体上，约占染色体的60%(1.5Mb)和总基因组的10%。RDNA由大约150个拷贝组成，产生的rRNA约占总RNA的80%。由于rDNA的重复结构和高转录活性，rDNA是重组的。因此，重复序列是基因组中一个独特而独特的区域。为了研究细胞分裂周期中重复序列的动态性质，我们开发了酵母菌株，其中每个rDNA重复单元都具有分别与LacI-GFP或TetR-MRFP蛋白相关联的LACO或Teto阵列。在这些菌株中，整个重复序列可以在活细胞中可视化，而不需要任何特殊处理。对这些菌株的分析表明，rDNA在核仁和核质之间的交界处活跃移动并改变形状。在G2/M期，rDNA的姐妹染色单体在核仁中形成环状结构，呈现出旋转的外观。我们还发现，在细胞周期的M期，rDNA重复的长度影响分离所需的时间。
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% (1.5 Mb) 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.