Asymmetry of nucleotide substitutions in tRNAs indicates common descent of modern organisms from a thermophilic ancestor

   The nature of the last universal common ancestor (LUCA) of all living organisms remains a controversial issue in biology. There is evidence of both thermophilic and mesophilic LUCA origin. The increasing complexity of the cellular apparatus during the evolution from early life forms to modern organisms could have manifested itself in long-term evolutionary changes in the nucleotide composition of genetic sequences. This work is devoted to the identification of such trends in tRNA sequences. The results of an evolutionary analysis of single-nucleotide substitutions in tRNAs of 123 species from three domains – Bacteria, Archaea and Eukaryota – are presented. A universal vector of directed evolutionary change in tRNA sequences has been discovered, in which substitutions of guanine (G) to adenine (A) and cytosine (C) to uracil (U) occur more frequently than the reverse. The most striking asymmetry in the number of substitutions is observed in the following transitions: a) purine-to-purine, where G→A outnumbers A→G, b) pyrimidine-to-pyrimidine, where C→U outnumbers U→C, and c) purine-to-pyrimidine and vice versa, where G→U outnumbers U→G. As a result, tRNAs could lose “strong” three-hydrogen-bond complementary pairs formed by guanine and cytosine and fix “weak” two-hydrogen-bond complementary pairs formed by adenine and uracil. 16 out of 20 tRNA families are susceptible to the detected change in sequence composition, which corresponds to the significance level p = 0.006 according to the one-sided binomial test. The identified pattern indicates a high GC content in the common ancestor of modern tRNAs, supporting the hypothesis that the last universal common ancestor (LUCA) lived in a hotter environment than do most contemporary organisms.

1. Bermudez-Santana C., Attolini C.S.-O., Kirsten T., Engelhardt J., Prohaska S.J., Steigele S., Stadler P.F. Genomic organization of eukaryotic tRNAs. BMC Genomics. 2010;11(1):270. doi: 10.1186/1471-2164-11-270

2. Cantine M.D., Fournier G.P. Environmental adaptation from the origin of life to the last universal common ancestor. Orig Life Evol Biosph. 2017;48(1):35-54. doi: 10.1007/s11084-017-9542-5

3. Di Giulio M. The universal ancestor lived in a thermophilic or hyperthermophilic environment. J Theor Biol. 2000;203(3):203-213. doi: 10.1006/jtbi.2000.1086

4. Dutta A., Chaudhuri K. Analysis of tRNA composition and folding in psychrophilic, mesophilic and thermophilic genomes: indications for thermal adaptation. FEMS Microbiol Lett. 2010;305(2):100-108. doi: 10.1111/j.1574-6968.2010.01922.x

5. Galtier N., Tourasse N., Gouy M. A non hyperthermophilic common ancestor to extant life forms. Science. 1999;283(5399):220-221. doi: 10.1126/science.283.5399.220

6. Jordan I.K., Kondrashov F.A., Adzhubei I.A., Wolf Y.I., Koonin E.V., Kondrashov A.S., Sunyaev S. A universal trend of amino acid gain and loss in protein evolution. Nature. 2005;433(7026):633-638. doi: 10.1038/nature03306

7. Klopfstein S., Vilhelmsen L., Ronquist F. A nonstationary Markov model detects directional evolution in hymenopteran morphology. Syst Biol. 2015;64(6):1089-1103. doi: 10.1093/sysbio/syv052

8. Lehmann E.L. The Fisher, Neyman-Pearson theories of testing hypotheses: one theory or two? In: Rojo J. (Ed.) Selected Works of E.L. Lehmann. Selected Works in Probability and Statistics. Boston, MA: Springer, 2012;201-208. doi :10.1007/978-1-4614-1412-4_19

9. Men Y., Lu G., Wang Y., Lin J., Xie Q. Reconstruction of the rRNA sequences of LUCA, with bioinformatic implication of the local similarities shared by them. Biology. 2022;11(6):837. doi: 10.3390/biology11060837

10. Moody E.R.R., Álvarez-Carretero S., Mahendrarajah T.A., Clark J.W., Betts H.C., Dombrowski N., Szánthó L.L., … Spang A., Pisani D., Williams T.A., Lenton T.M., Donoghue P.C.J. The nature of the last universal common ancestor and its impact on the early Earth system. Nat Ecol Evol. 2024;8(9):1654-1666. doi: 10.1038/s41559-024-02461-1

11. Rickert D.A., Fan L.W.-T., Hahn M.W. Inconsistency of parsimony under the multispecies coalescent. Theor Popul Biol. 2025;166:56-69. doi: 10.1016/j.tpb.2025.09.004

12. Romanova E.V., Bukin Y.S., Mikhailov K.V., Logacheva M.D., Aleoshin V.V., Sherbakov D.Yu. Hidden cases of tRNA gene duplication and remolding in mitochondrial genomes of amphipods. Mol Phylogenet Evol. 2020;144:106710. doi: 10.1016/j.ympev.2019.106710

13. Soucy S.M., Huang J., Gogarten J.P. Horizontal gene transfer: building the web of life. Nat Rev Genet. 2015;16(8):472-482. doi: 10.1038/nrg3962

14. Sprinzl M., Horn C., Brown M., Ioudovitch A., Steinberg S. Compilation of tRNA sequences and sequences of tRNA genes. Nucleic Acids Res. 1998;26(1):148-153. doi: 10.1093/nar/26.1.148

15. Velandia-Huerto C.A., Berkemer S.J., Hoffmann A., Retzlaff N., Romero Marroquín L.C., Hernández-Rosales M., Stadler P.F., Bermúdez-Santana C.I. Orthologs, turn-over, and remolding of tRNAs in primates and fruit flies. BMC Genomics. 2016;17(1):617. doi: 10.1186/s12864-016-2927-4

16. Weiss M.C., Sousa F.L., Mrnjavac N., Neukirchen S., Roettger M., Nelson-Sathi S., Martin W.F. The physiology and habitat of the last universal common ancestor. Nat Microbiol. 2016;1(9):16116. doi: 10.1038/nmicrobiol.2016.116