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. 2018;15(4-5):518-527.
doi: 10.1080/15476286.2017.1349049. Epub 2017 Aug 29.

Functionality of tRNAs encoded in a mobile genetic element from an acidophilic bacterium

Pamela Alamos  1 , Mario Tello  2 , Paula Bustamante  1 , Fernanda Gutiérrez  1 , Amir Shmaryahu  3 , Juan Maldonado  1 , Gloria Levicán  4 , Omar Orellana  1
Affiliations

Functionality of tRNAs encoded in a mobile genetic element from an acidophilic bacterium

Pamela Alamos et al. RNA Biol. 2018.

Abstract

The genome of the acidophilic, bioleaching bacterium Acidithiobacillus ferrooxidans, strain ATCC 23270, contains 95 predicted tRNA genes. Thirty-six of these genes (all 20 species) are clustered within an actively excising integrative-conjugative element (ICEAfe1). We speculated that these tRNA genes might have a role in adapting the bacterial tRNA pool to the codon usage of ICEAfe1 genes. To answer this question, we performed theoretical calculations of the global tRNA adaptation index to the entire A. ferrooxidans genome with and without the ICEAfe1 encoded tRNA genes. Based on these calculations, we observed that tRNAs encoded in ICEAfe1 negatively contribute to adapt the tRNA pool to the codon use in A. ferrooxidans. Although some of the tRNAs encoded in ICEAfe1 are functional in aminoacylation or protein synthesis, we found that they are expressed at low levels. These findings, along with the identification of a tRNA-like RNA encoded in the same cluster, led us to speculate that tRNA genes encoded in the mobile genetic element ICEAfe1 might have acquired mutations that would result in either inactivation or the acquisition of new functions.

Keywords: Transfer RNA; acidophilic bacterium; horizontal gene transfer; mobile genetic elements; phylogeny; translation.

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Figures

Figure 1.
Figure 1.
Cladogram of genes encoding for bacterial tRNAs from acidophilic and other microorganisms: Phylogenetic reconstruction of bacterial genes encoding for tRNAs that charge lysine, arginine, serine and alanine. The colored symbols represent the tRNAs present in ICEAfe1 from A. ferrooxidans ATCC 23270 (red circle) or chromosomal tRNAs from, Acidithiobacillus sp GGI221, A. ferrooxidans strains ATCC 53993, 23270, YQH1, Hel18, BY0502; A. ferrivorans strains SS3, CF27, DLC5, PQ33, YL15; A. thiooxidans strains 19377, A01, Licanantay, DXSW, BY02, ZBY, A02, DMC, GD13, JYC17; A. caldus SM-1, MTH04, DX, ZBY, ZJ, S1 (green rhombus) and chromosomal tRNAs from Leptospirillum ferriphilum ML-04, L. ferrooxidans C2–3 (blue rhombus). tRNAs from other bacteria were obtained from tRNA database. To improve the visualization, clades including tRNAs not related with acidithiobacillus genera were collapsed and are shown in gray lines. Clades that contain mainly tRNAs from Acidithiobacillus genus were collapsed, named as groups and signaled with green lines when they contain tRNA(s) that belong to ICEAfe1.
Figure 2.
Figure 2.
tRNA adaptation index (tAI) of genes from Acidithiobacillus ferrooxidans strain ATCC 23270 (A)and B. Gene frequency vs. tAI value for wild-type (wt) or virtual mutant (vm, tRNA genes from ICEAfe1 were excluded from calculations). (C) and D,. gene frequency vs. log-ratio of the tAI values for the wild-type and virtual mutant, as in (A) and B, except that the calculations were performed based on the actual tRNA levels obtained from RNA-seq data (Experiment 1). Total genes represent genes from chromosome and ICEAfe1. ICE genes are genes from ICEAfe1 only.
Figure 3.
Figure 3.
Expression level of tRNAs from A. ferrooxidans. Number of reads for each tRNA from RNA-seq, ordered as they are organized in the genome. Data of reads for each tRNA are in S2 Table 1. Horizontal bar points out the tRNAs encoded within ICEAfe1.
Figure 4.
Figure 4.
Functional analysis of tRNASer UGA. a (Upper panel). Schematic representation of the genes (arrows) serTi, encoding the tRNASer UGA 75-containing intron (light gray), and serT, encoding the canonical tRNASer UGA 55 (ICEAfe1, white bar; chromosome, black line. Not at scale), from A. ferrooxidans strain ATCC 23270. b (Lower panel) Northern blot analysis of periodate-treated total tRNA from A. ferrooxidans before or after (OH-/OH+) incubation with Tris-acetate, pH 9, followed by electrophoresis under denaturing conditions, revealed with specific probes for tRNASer UG A 75, tRNA Ser UGA 55 or tRNAGln UUG (chromosomal tRNA used as control). A tRNA preparation from strain ATCC 53993 that lacks ICEAfe1 and therefore contains only the chromosomally-encoded version of tRNASer UGA was also used as a control.
Figure 5.
Figure 5.
Predicted structure of the RNA encoded by tRNA-OTH. Schematic representation of the predicted RNA based on tRNAscan (A), ARAGORN (B) and MFold (C). Note that RNAs predicted in A and B are contained in C.
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Grants and funding

This work was supported by Fondo Nacional de Ciencias y Tecnología, Chile under grants1110203 and 1150834 to O.O. (http://www.fondecyt.cl); and the Fundación de Innovación Agraria, Chile under grants PYT20120056 and USA 1555 to MT. (http://www.fia.cl). A graduate fellowship from Programa de Mejoramiento de la Educación Superior, Chile was granted to P.A. A graduate fellowship from USACH was granted to J.M.