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. 2023 Dec 19;52(4):1975–1987. doi: 10.1093/nar/gkad1206

Figure 2.

Figure 2.

Purified pre-60S assembly intermediates upon Dbp10 depletion and from dbp10 K187R mutant cells. AID-HA-DBP10 NOP7-FTpA and NSA1-FTpA strains were transformed with plasmids harbouring wild-type DBP10, the dbp10 K187R mutant, or empty vector (–). Subsequently, Nop7 (A) and Nsa1 (B) particles were affinity purified from cells in presence of AID-HA-Dbp10 (untreated) or upon its auxin-induced proteasomal degradation (+auxin). Final eluates were analysed by SDS-PAGE and Coomassie staining (left panels) or Western blotting using indicated antibodies (right panels). Depicted Coomassie stained protein bands were identified by mass spectrometry. Bait proteins are marked with an asterisk. Protein bands with increased or decreased intensity in the K187R mutant samples are indicated in red and blue, respectively. (C–E) Final Nop7 and Nsa1 eluates purified from DBP10 wild-type or K187R mutant cells were analysed by timsTOF mass spectrometry in four biological replicates. Median-normalised label-free quantification (LFQ) values of the co-enriched proteins were log2-transformed and visualized as volcano plots (C and D) or box plots focusing on selected assembly factors from the Nop7 affinity purifications (E). Proteins that were increased or decreased in the K187R mutant vs DBP10 wild-type samples are indicated in red and blue, respectively.