Rtt109 Prevents Hyper-Amplification of Ribosomal RNA Genes through Histone Modification in Budding Yeast (original) (raw)
Figure 2
Hyper-amplification of rDNA is induced by deletion of RTT109.
A) Chromosome XII with rDNA in the wild type and rtt109 mutant; DNA was analyzed by pulsed-field gel electrophoresis (CHEF), and stained with EtBr (left) and hybridized with rDNA (right). Lanes 1 and 2 are the wild type strains and lanes 3–7 are the rtt109 mutant in the W303 genetic background. Chromosome XII indicated with the asterisk is longer after hyper-amplification. B) Quantitation of rDNA copy number in the wild type (lane 1) and rtt109 mutant (lanes 2 and 3) by Southern hybridization. The upper panel shows the rDNA band after _Bgl_II digestion. MCM2 is the internal control used for normalization. The lower panel indicates quantification of the signal intesity of each band detected by Southern blotting with rDNA and MCM2 probes, respectively. Values are means of three independent experiments and bars are S.D. values. C, D) RTT109 restored wild type levels of rDNA copies in the rtt109 mutant. Chromosome XII was analyzed by CHEF analysis followed by Southern hybridization with an rDNA probe in the rtt109 mutant at ∼24 generations (C) and at ∼90 generations (D) after introduction of the empty vector (C: lanes 1–5, D: lanes 1–4, 9–12) or plasmid-encoded RTT109 (C: lanes 6–10, D: 5–8, 13–16). Mutants were constructed in the W303 and S288c genetic backgrounds.