Effect of trmA on tRNA expression in Pseudomonas aeruginosa

Goal: to study the differences in P. aeruginosa tRNA expression resulted from absence of trmA, a tRNA m5U modifying enzyme. Results: In exponential-phase wild-type and trmA mutant cells, seven tRNAs comprised almost 50% of the total tRNA abundance (Gly-GCC, Arg-ACG, Ala-GGC, Asp-GTC, Gln-TTG, Leu-CAG, and Tyr-GTA), whereas Arg-CCT had the lowest expression (0.04-0.08%) of the tRNA pool. In the trmA mutant compared to the wild type, 3 genes were up-regulated, and 3 genes were down-regulated in the trmA mutant. Conclusion: This observation shows that the m5U modification affect certain tRNA expression in the tRNA pool in P. aeruginosa, even though it presents in all tRNA species. To investigate the effect of the loss of TrmA on the abundance of tRNAs, we constructed the trmA mutant strain. Small RNAs were extracted from exponential phase cells of both the wild-type and trmA mutant strains with three biological replicates. cDNA libraries were constructed following the protocol of Absolute Quantification RNA Sequencing (AQRNA-seq) and sequenced on an Illumina MiSeq platform. Differential expression analysis was performed on the AQRNA-seq data to identify tRNAs that were significantly up- or down-regulated in response to the loss of TrmA.