Recent evidence for evolution of the genetic code.

The genetic code, formerly thought to be frozen, is now known to be in a state of evolution. This was first shown in 1979 by Barrell et al. (G. Barrell, A. T. Bankier, and J. Drouin, Nature [London] 282:189-194, 1979), who found that the universal codons AUA (isoleucine) and UGA (stop) coded for methionine and tryptophan, respectively, in human mitochondria. Subsequent studies have shown that UGA codes for tryptophan in Mycoplasma spp. and in all nonplant mitochondria that have been examined. Universal stop codons UAA and UAG code for glutamine in ciliated protozoa (except Euplotes octacarinatus) and in a green alga, Acetabularia. E. octacarinatus uses UAA for stop and UGA for cysteine. Candida species, which are yeasts, use CUG (leucine) for serine. Other departures from the universal code, all in nonplant mitochondria, are CUN (leucine) for threonine (in yeasts), AAA (lysine) for asparagine (in platyhelminths and echinoderms), UAA (stop) for tyrosine (in planaria), and AGR (arginine) for serine (in several animal orders) and for stop (in vertebrates). We propose that the changes are typically preceded by loss of a codon from all coding sequences in an organism or organelle, often as a result of directional mutation pressure, accompanied by loss of the tRNA that translates the codon. The codon reappears later by conversion of another codon and emergence of a tRNA that translates the reappeared codon with a different assignment. Changes in release factors also contribute to these revised assignments. We also discuss the use of UGA (stop) as a selenocysteine codon and the early history of the code.     

Sequence evidence for an altered genetic code in the Neospora caninum plastid

The plastid DNA of Neospora caninum encodes a homologue of the rpoB gene, which is believed to encode a subunit of a bacterial or chloroplast-like RNA polymerase. The predicted protein product of the N. caninum rpoB gene has three in-frame UGA codons which appear to encode tryptophan residues rather than act as stop codons. Based on the nucleotide sequence of a portion of the ssrRNA gene of the N. caninum plastid, a model for suppression of UGA termination in this plastid is presented.     

Aminoacyl-tRNA synthetases, the genetic code, and the evolutionary process.

The aminoacyl-tRNA synthetases (AARSs) and their relationship to the genetic code are examined from the evolutionary perspective. Despite a loose correlation between codon assignments and AARS evolutionary relationships, the code is far too highly structured to have been ordered merely through the evolutionary wanderings of these enzymes. Nevertheless, the AARSs are very informative about the evolutionary process. Examination of the phylogenetic trees for each of the AARSs reveals the following. (i) Their evolutionary relationships mostly conform to established organismal phylogeny: a strong distinction exists between bacterial- and archaeal-type AARSs. (ii) Although the evolutionary profiles of the individual AARSs might be expected to be similar in general respects, they are not. It is argued that these differences in profiles reflect the stages in the evolutionary process when the taxonomic distributions of the individual AARSs became fixed, not the nature of the individual enzymes. (iii) Horizontal transfer of AARS genes between Bacteria and Archaea is asymmetric: transfer of archaeal AARSs to the Bacteria is more prevalent than the reverse, which is seen only for the "gemini group. " (iv) The most far-ranging transfers of AARS genes have tended to occur in the distant evolutionary past, before or during formation of the primary organismal domains. These findings are also used to refine the theory that at the evolutionary stage represented by the root of the universal phylogenetic tree, cells were far more primitive than their modern counterparts and thus exchanged genetic material in far less restricted ways, in effect evolving in a communal sense.     

Reliability in the genetic code.

Base sequences of φ × 174 and MS2 viruses genomes and of some mRNAs (Coat protein fd virus, Rabbit B. Globin, Rat Growth Hormone and Human Chorionic, Somatomammotropin) show a preferential use of some amino-acid codons. Based on this observation the reliability of three non-degenerate codes are analyzed. All of them display higher reliability than the standing genetic code and specially one formed by a set of non-directly related codons.The absence of these type of codes in Nature is discussed in terms of a balance between reliability and mutability of the genetic information, able to preserve species and maintain evolution     

The genetic code, an instantaneous absolutely optimal code

The genetic code is an instantaneous code i.e., each codon is deciphered out of ambiguities without knowing other symbols than the constituting nucleotides. Moreover entropy of the genetic source of information has a maximal value.     

The origin of the genetic code.

A new approach to the origin of the genetic code is proposed based on some regularities in the nucleotide distribution pattern of the code. The relative amounts of various amino acids in primitive proteins were possibly different from those in organisms living today. The primordial ratio was supposed to shift to the modern one guided by the action of primitive nucleotides. Each primitive tRNA had a discriminator site and, distinguished from it, an anticodon site. It also postulated that primordially each amino acid could correspond to a wide variety of codons. During the course of the evolutionary change, a selective mechanism worked among the protobionts so that less frequent nucleotides became associated with more abundant amino acids in the primordial conditions,thus finally leading to the present codon catalogue.     

Optimization and the genetic code

The present paper will focus on the relation between the structure of the table of the genetic code and the evolution of primitive organisms: it will be shown that the organization of the code table according to an optimization principle based on the notion of resistance to errors can provide a criterium for selection. The ordered aspect of the genetic code table makes this result a plausible starting point for studies of the origin and evolution of the genetic code: these could include, besides a more refined optimization principle at the logical level, some effects more directly related to the physico-chemical context, and the construction of realistic models incorporating both aspects.     

Hemoglobin and the genetic code

Summary One-half of the twenty amino acids of the genetic code are just one mutational step away from the chain-terminator codons UAA, UAG, and UGA. It is postulated that somatic mutation to terminator is a hazard to which the organism has had to respond by adjusting certain proteins in the direction of fewer mutable residues. This view is supported by calculations based on the primary structure of five of the human hemoglobin chains. Each chain is scored for mutability to terminator in accord with the numbers and kinds of amino acids present. Among the adult chains, the most essential one, the alpha, has lowest mutability. The beta and delta follow, and in order of the presumed harm to the organism of a shortage of chain copies. Ante-natal chains tend to have higher mutabilities, supporting the view that cumulative mutational change in DNA can do little harm if the gene ceases to transcribe early in life. Two other predictions based on the supposition of effective selection against mutability to terminator are also met: chain length of polypeptides is negatively correlated with their scores for mutability to terminator, and examination of the recently determined sequence of beta messenger RNA shows preferential use of codons that are not readily mutable to terminator.Supported in part by the National Institutes of Health, Grant HL-16005     

Flexizymes for genetic code reprogramming

Genetic code reprogramming is a method for the reassignment of arbitrary codons from proteinogenic amino acids to nonproteinogenic ones; thus, specific sequences of nonstandard peptides can be ribosomally expressed according to their mRNA templates. Here we describe a protocol that facilitates genetic code reprogramming using flexizymes integrated with a custom-made in vitro translation apparatus, referred to as the flexible in vitro translation (FIT) system. Flexizymes are flexible tRNA acylation ribozymes that enable the preparation of a diverse array of nonproteinogenic acyl-tRNAs. These acyl-tRNAs read vacant codons created in the FIT system, yielding the desired nonstandard peptides with diverse exotic structures, such as N-methyl amino acids, D-amino acids and physiologically stable macrocyclic scaffolds. The facility of the protocol allows a wide variety of applications in the synthesis of new classes of nonstandard peptides with biological functions. Preparation of flexizymes and tRNA used for genetic code reprogramming, optimization of flexizyme reaction conditions and expression of nonstandard peptides using the FIT system can be completed by one person in approximately 1 week. However, once the flexizymes and tRNAs are in hand and reaction conditions are fixed, synthesis of acyl-tRNAs and peptide expression is generally completed in 1 d, and alteration of a peptide sequence can be achieved by simply changing the corresponding mRNA template.     

Experimental evidence for asymmetrical shielding of nucleosomal DNA by histones.

Electron spin resonance study of Mn (II) binding to chromatin and derivatives, including core particles, shows that Mn (II) is a good probe for testing the overall electrostatic balance of the nucleoproteic complex as well as DNA accessibility. Experimental results are in good agreement with a recent model proposed (Mirzabekov A. D. and Rich A. (1979) Proc. Natl. Acad. Sci. USA 76, 1118-1121), for the core particle, in which an asymmetrical shielding of DNA by the protein core is assumed. Furthermore, it was found that the histone H1 hinders a number of charges on the linker DNA in a proportion equal to the net positive charge of the histone itself. This result is interpreted as due to a tighter interaction between the linker DNA and the core histones in the presence of histone H1.     

Evolution of mitochondrial genomes and the genetic code.

Mitochondrial genomes are clearly marked by a strong tendency towards reductive evolution. This tendency has been facilitated by the transfer of most of the essential genes for mitochondrial propogation and function to the nuclear genome. The most extreme examples of genomic simplification are seen in animal mitochondria, where there also are the greatest tendencies to codon reassignment. The reassignment of codons to amino acids different from those designated in the so called universal code is seen in part as an expression of the reduction of the number of genes used by these genomes to code for tRNA species. The driving force for the reductive evolution of mitochondrial genomes is identified with two population genetic effects which may also be operating on populations of parasites.     

Streptomycin-induced, third-position misreading of the genetic code

Streptomycin was used to increase the frequency of errors in protein synthesis in vivo. In the system under study two misreading errors were observed. Both involved the erroneous insertion of lysine at asparagine codons, because of misreading of a pyrimidine as a purine at the 3' position of the codon. Streptomycin increased the errors at the two codons AAU and AAC to the same extent, thereby maintaining the error ratio found for basal level mistranslation.     

tRNA genes and the genetic code

The genetic code describes translational assignments between codons and amino acids. tRNAs and aminoacyl-tRNA synthetases (aaRSs) are those molecules by means of which these assignments are established. Any aaRS recognizes its tRNAs according to some of their nucleotides called identity elements (IEs). Let a 1Mut-similarity Sim (1Mut) be the average similarity between such tRNA genes whose codons differ by one point mutation. We showed that: (1) a global maximum of Sim (1Mut) is reached at the standard genetic code 27 times for 4 sets of IEs of tRNA genes of eukaryotic species, while it is so only 5 times for similarities Sim (C&R) between all tRNA genes whose codons lie in the same column or row of the code. Therefore, point mutations of anticodons were tested by nature to recruit tRNAs from one isoaccepting group to another, (2) because plain similarities Sim (all) between tRNA genes of species within any of the three domains of life are higher than between tRNA genes of species belonging to different domains, tRNA genes retained information about early evolution of cells, (3) we searched the order of tRNAs in which they were most probably assigned to their codons and amino acids. The beginning Ala, (Val), Pro, Ile, Lys, Arg, Trp, Met, Asp, Cys, (Ser) of our resulting chronology lies under a plateau on a graph of Sim (1Mut,IE)(univ.ancestors) plotted over this chronology for a set S(IE) of all IEs of tRNA genes, whose universal ancestors were separately computed for each codon. This plateau has remained preserved along the whole line of evolution of the code and is consistent with observations of Ribas de Pouplana and Schimmel [2001. Aminoacy1-tRNA synthetases: potential markers of genetic code development. Trends Biochem. Sci. 26, 591-598] that specific pairs of aaRSs-one from each of their two classes-can be docked simultaneously onto the acceptor stem of tRNA and hence an interaction existed between their ancestors using a reduced code, (4) sharpness of a local maximum of Sim (1Mut) at the standard code is almost 100% along our chronologies.