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. 2012 Apr 13;31(11):2579–2589. doi: 10.1038/emboj.2012.85

Figure 4.

Figure 4

Proteasome activity is required for 25S NRD. (A–D) Stability of nonfunctional rRNA in a proteasome-inhibited cell. (A–C) The 18-nt-tagged 25S rRNA was detected by northern blotting. (A) The cim3-1 ts strain was grown at the permissive temperature. (B) Proteasomes were depleted in the RPT2 tet-off strain with Dox treatment for 12 h. (C, D) MG132 was administered at the indicated concentrations to the erg6Δ strain containing the combination of plasmids indicated, to overexpress ubiquitin. (D) The other 16-nt-tagged 18S rRNAs, with or without A1492C mutation, were detected by northern blotting. (E, F) Subcellular localization of 18-nt-tagged 25S rRNAs in various mutant strains. After transcriptional shut-off, the cells were harvested at the indicated time points. (E) The tagged 25S rRNAs were visualized with an in-situ hybridization technique. Scale bar, 4 μm. (F) Cleared lysates were resolved on a 10–40% sucrose gradient and the amounts of tagged rRNA were visualized by northern blotting. All dots of every lane come from the same membrane. Figure source data can be found with the Supplementary data.