Transfer RNA and the origins of diversified life.

ABSTRACT The evolution of the transfer RNA (tRNA) molecule is controversial but embeds the history of protein biosynthesis, the genetic code, and the origins of diversified life. A new phylogenetic method based on RNA structure that we developed provides new lines of evidence to support the genome tag hypothesis and confirms that the 'top half' of tRNA is more ancient than the 'bottom half' Timelines of amino acid charging function generated from constraint analyses showed that selenocysteine, tyrosine, serine, and leucine specificities were ancient, while those related to asparagine, methionine, and arginine were more recent. The timelines also uncovered an early role of the second and then first codon bases, identified codons for alanine and proline as the most ancient, and revealed important evolutionary take-overs related to the loss of the long variable arm of tRNA. Furthermore, organismal timelines showed Archaea was the oldest superkingdom, followed by viruses, and superkingdoms Eukarya and Bacteria in that order, supporting conclusions from recent phylogenomic studies of protein architecture. Strikingly, results showed that the origin of viruses was not only ancient but was linked to Archaea, supporting the notion that the arehaeal lineage is the most ancient on earth and its origin predated diversification of tRNA function and specificity. Keywords: tRNA, secondary structure, Archaea, Bacteria, Eukarya, viruses, genetic code, amino acids charging, selenocysteine, phylogenetics Introduction The origins and evolution of the genetic code and proteins and the emergence of the diversified world embody fundamental biological questions that are of great importance to our understanding of life. Unfortunately, concrete answers to these questions are lacking, are incomplete, or are highly controversial. This is partially due to our inability to uncover significant evolutionary signal in the molecules of life that have been examined to date. For example, ribosomal RNA (rRNA) molecules have been used to reconstruct a tree of life but the rooting of the tree that defines Bacteria as the most ancient lineage contrasts with evidence derived from genomics and remains controversial. Transfer RNAs (tRNAs) are very ancient nucleic acid molecules and promising molecular fossil candidates. They are central to one of the most fundamental biological functions, the ability to synthesize proteins (translation). Translation is a property shared by all cellular organisms (generally missing in viruses) and its conservation suggests it is ancient and embeds the history of the living world. tRNA molecules are generally short and are composed of runs of ~73-95 nucleotides that generally adopt a typical cloverleaf-shaped secondary structure and fold into a L-like conformation in three dimensional (3D) space (Figure 1A). This cloverleaf structure is made up of two structural and functional domains. The acceptor (Acc) and the pseudouridine (T[PSI]C) arms comprise the "top half" domain, while the "bottom half" domain is composed of the dihydrouridine (DHU) and the anticodon (AC) arms. A variable (extra) arm spans the T[PSI]C and AC arms. tRNA plays vital roles in protein synthesis by: (i) transporting (charging) an amino acid onto its Acc arm thanks to the activity of specific aminoacyl-tRNA synthetases (aaRSs), and (ii) using a triplet (anticodon) nucleotide sequence located in the AC loop to translate information in a complementary (codon) sequence of messenger RNA (mRNA) that encodes amino acid charging and defines a "genetic code" (Figure 1B). When nested into the rRNA subunits of the ribosomal ensemble, the codons are deciphered along the mRNA and charged amino acids are enzymatically transformed into growing polypeptide chains, ultimately generating functional proteins. Because of their central role in protein synthesis, tRNA and several other macromolecules (e.g., aaRSs and rRNAs) have been considered critical references for evolutionary analysis. …