Modification sites at positions 46 or 47 that are lost in mutant strain of Pseudomonas aeruginosa PA14_17650. (A) LC–MS/MS quantification of acp3U from purified tRNA of wild-type P. aeruginosa versus PA14_17650 mutant for n = 2 replicates. Statistical significance was determined by Student's t-test. (B) Heatmap of average (from three independent samples cultured in CDM) frequency of misincorporation and average termination score at positions 46 and 47 in tRNA of P. aeruginosa from wild-type (left) and PA14_17650 mutant (right). Only tRNAs with read coverage >100 at position 46 are represented. The GtRNAdb names for P. aeruginosa tRNAs are listed on the left, with the anticodon shown in parentheses. The short name for the modification is shown at top right column (Supplemental Table S11). The triangle symbol labeled on the right of the heatmap indicates whether a signature is dependent PA14_17650, wherein the signature is lost in the mutant. The circle marks labeled on the right of the heatmap are color-coded and indicate conservation in Escherichia coli (key listed below the heatmap). The criteria used is based on a 5% cutoff of both misincorporation and termination for either position 46 or 47.
