Figure 2.

Majority of Ψ modifications are present in the 35S and 45S intermediates. The calculation of average mutation rates for the Ψs detections were performed, as explained in the Materials and Methods section of the paper. (A,B) Average mutation rates for specific nucleotides of the 35S and 45S intermediates and native 50S large subunit are shown in green, blue, and red, respectively. The data are the averages of two biological replicates, and the errors are the standard deviation from the averages. (A) Nucleotides U 748, 957, 2461, 2508, 2584, 2608, 2609 isomerizations to Ψ occur before the 35S and 45S intermediates are populated in cells. The extents of the nucleotides U 748, 957, 2461, 2508, 2584, 2608, and 2609 isomerizations to Ψ are similar for the 35S, 45S, and 50S particles. Therefore, these isomerizations occur before the 35S and 45S intermediates are formed. (B) Nucleotides U 1915, 1919, and 1921 are in the process of being isomerized to Ψ in the 35S and 45S intermediates or will be isomerized to Ψ during the later stages of large subunit ribosome assembly. The nucleotides U 1915, 1919, and 1921 are extensively more isomerized to Ψ in the native 50S large subunit than that in the 45S and 35S intermediates. As explained in the manuscript, the average mutation rate for m³Ψ 1919 (1919*) was calculated from the two CMCT- and NaHCO₃-treated samples without subtracting the mutation rate of NaHCO₃ only treated control sample. Thus, no background correction was performed on the m³Ψ 1919 nucleotide data.