Root of the universal tree of life based on ancient aminoacyl-tRNA synthetase gene duplications.

Universal trees based on sequences of single gene homologs cannot be rooted. Iwabe et al. [Iwabe, N., Kuma, K.-I., Hasegawa, M., Osawa, S. & Miyata, T. (1989) Proc. Natl. Acad. Sci. USA 86, 9355-9359] circumvented this problem by using ancient gene duplications that predated the last common ancestor of all living things. Their separate, reciprocally rooted gene trees for elongation factors and ATPase subunits showed Bacteria (eubacteria) as branching first from the universal tree with Archaea (archaebacteria) and Eucarya (eukaryotes) as sister groups. Given its topical importance to evolutionary biology and concerns about the appropriateness of the ATPase data set, an evaluation of the universal tree root using other ancient gene duplications is essential. In this study, we derive a rooting for the universal tree using aminoacyl-tRNA synthetase genes, an extensive multigene family whose divergence likely preceded that of prokaryotes and eukaryotes. An approximately 1600-bp conserved region was sequenced from the isoleucyl-tRNA synthetases of several species representing deep evolutionary branches of eukaryotes (Nosema locustae), Bacteria (Aquifex pyrophilus and Thermotoga maritima) and Archaea (Pyrococcus furiosus and Sulfolobus acidocaldarius). In addition, a new valyl-tRNA synthetase was characterized from the protist Trichomonas vaginalis. Different phylogenetic methods were used to generate trees of isoleucyl-tRNA synthetases rooted by valyl- and leucyl-tRNA synthetases. All isoleucyl-tRNA synthetase trees showed Archaea and Eucarya as sister groups, providing strong confirmation for the universal tree rooting reported by Iwabe et al. As well, there was strong support for the monophyly (sensu Hennig) of Archaea. The valyl-tRNA synthetase gene from Tr. vaginalis clustered with other eukaryotic ValRS genes, which may have been transferred from the mitochondrial genome to the nuclear genome, suggesting that this amitochondrial trichomonad once harbored an endosymbiotic bacterium.

Bootstrap confidence levels for phylogenetic trees

Evolutionary trees are often estimated from DNA or RNA sequence data. How much confidence should we have in the estimated trees? In 1985, Felsenstein [Felsenstein, J. (1985) Evolution 39, 783–791] suggested the use of the bootstrap to answer this question. Felsenstein’s method, which in concept is a straightforward application of the bootstrap, is widely used, but has been criticized as biased in the genetics literature. This paper concerns the use of the bootstrap in the tree problem. We show that Felsenstein’s method is not biased, but that it can be corrected to better agree with standard ideas of confidence levels and hypothesis testing. These corrections can be made by using the more elaborate bootstrap method presented here, at the expense of considerably more computation.

Matchings and phylogenetic trees

This paper presents a natural coordinate system for phylogenetic trees using a correspondence with the set of perfect matchings in the complete graph. This correspondence produces a distance between phylogenetic trees, and a way of enumerating all trees in a minimal step order. It is useful in randomized algorithms because it enables moves on the space of trees that make random optimization strategies “mix” quickly. It also promises a generalization to intermediary trees when data are not decisive as to their choice of tree, and a new way of constructing Bayesian priors on tree space.

A dual-specificity aminoacyl-tRNA synthetase in the deep-rooted eukaryote Giardia lamblia

Cysteinyl-tRNA (Cys-tRNA) is essential for protein synthesis. In most organisms the enzyme responsible for the formation of Cys-tRNA is cysteinyl-tRNA synthetase (CysRS). The only known exceptions are the euryarchaea Methanococcus jannaschii and Methanobacterium thermoautotrophicum, which do not encode a CysRS. Deviating from the accepted concept of one aminoacyl-tRNA synthetase per amino acid, these organisms employ prolyl-tRNA synthetase as the enzyme that carries out Cys-tRNA formation. To date this dual-specificity prolyl-cysteinyl-tRNA synthetase (ProCysRS) is only known to exist in archaea. Analysis of the preliminary genomic sequence of the primitive eukaryote Giardia lamblia indicated the presence of an archaeal prolyl-tRNA synthetase (ProRS). Its proS gene was cloned and the gene product overexpressed in Escherichia coli. By using G. lamblia, M. jannaschii, or E. coli tRNA as substrate, this ProRS was able to form Cys-tRNA and Pro-tRNA in vitro. Cys-AMP formation, but not Pro-AMP synthesis, was tRNA-dependent. The in vitro data were confirmed in vivo, as the cloned G. lamblia proS gene was able to complement a temperature-sensitive E. coli cysS strain. Inhibition studies of CysRS activity with proline analogs (thiaproline and 5′-O-[N-(l-prolyl)-sulfamoyl]adenosine) in a Giardia S-100 extract predicted that the organism also contains a canonical CysRS. This prediction was confirmed by cloning and analysis of the corresponding cysS gene. Like a number of archaea, Giardia contains two enzymes, ProCysRS and CysRS, for Cys-tRNA formation. In contrast, the purified Saccharomyces cerevisiae and E. coli ProRS enzymes were unable to form Cys-tRNA under these conditions. Thus, the dual specificity is restricted to the archaeal genre of ProRS. G. lamblia's archaeal-type prolyl- and alanyl-tRNA synthetases refine our understanding of the evolution and interaction of archaeal and eukaryal translation systems.

B12-dependent ribonucleotide reductases from deeply rooted eubacteria are structurally related to the aerobic enzyme from Escherichia coli

The ribonucleotide reductases from three ancient eubacteria, the hyperthermophilic Thermotoga maritima (TM), the radioresistant Deinococcus radiodurans (DR), and the thermophilic photosynthetic Chloroflexus aurantiacus, were found to be coenzyme-B12 (class II) enzymes, similar to the earlier described reductases from the archaebacteria Thermoplasma acidophila and Pyrococcus furiosus. Reduction of CDP by the purified TM and DR enzymes requires adenosylcobalamin and DTT. dATP is a positive allosteric effector, but stimulation of the TM enzyme only occurs close to the temperature optimum of 80–90°C. The TM and DR genes were cloned by PCR from peptide sequence information. The TM gene was sequenced completely and expressed in Escherichia coli. The deduced amino acid sequences of the two eubacterial enzymes are homologous to those of the archaebacteria. They can also be aligned to the sequence of the large protein of the aerobic E. coli ribonucleotide reductase that belongs to a different class (class I), which is not dependent on B12. Structure determinations of the E. coli reductase complexed with substrate and allosteric effectors earlier demonstrated a 10-stranded β/α-barrel in the active site. From the conservation of substrate- and effector-binding residues we propose that the B12-dependent class II enzymes contain a similar barrel.

Visualization of Cavitated Vessels in Winter and Refilled Vessels in Spring in Diffuse-Porous Trees by Cryo-Scanning Electron Microscopy1

Xylem cavitation in winter and recovery from cavitation in the spring were visualized in two species of diffuse-porous trees, Betula platyphylla var. japonica Hara and Salix sachalinensis Fr. Schm., by cryo-scanning electron microscopy after freeze-fixation of living twigs. Water in the vessel lumina of the outer three annual rings of twigs of B. platyphylla var. japonica and of S. sachalinensis gradually disappeared during the period from January to March, an indication that cavitation occurs gradually in these species during the winter. In April, when no leaves had yet expanded, the lumina of most of the vessels of both species were filled with water. Many vessel lumina in twigs of both species were filled with water during the period from the subsequent growth season to the beginning of the next winter. These observations indicate that recovery in spring occurs before the onset of transpiration and that water transport through twigs occurs during the subsequent growing season. We found, moreover, that vessels repeat an annual cycle of winter cavitation and spring recovery from cavitation for several years until irreversible cavitation occurs.

Bootstrap confidence levels for phylogenetic trees.

Evolutionary trees are often estimated from DNA or RNA sequence data. How much confidence should we have in the estimated trees? In 1985, Felsenstein [Felsenstein, J. (1985) Evolution 39, 783-791] suggested the use of the bootstrap to answer this question. Felsenstein's method, which in concept is a straightforward application of the bootstrap, is widely used, but has been criticized as biased in the genetics literature. This paper concerns the use of the bootstrap in the tree problem. We show that Felsenstein's method is not biased, but that it can be corrected to better agree with standard ideas of confidence levels and hypothesis testing. These corrections can be made by using the more elaborate bootstrap method presented here, at the expense of considerably more computation.

Hyperplane arrangements, interval orders, and trees.

A hyperplane arrangement is a finite set of hyperplanes in a real affine space. An especially important arrangement is the braid arrangement, which is the set of all hyperplanes xi - xj = 1, 1 trees. For instance, the number of labeled interval orders that can be obtained from n intervals I1,..., In of generic lengths is counted. There is also discussed an arrangement due to N. Linial whose number of regions is the number of alternating (or intransitive) trees, as defined by Gelfand, Graev, and Postnikov [Gelfand, I. M., Graev, M. I., and Postnikov, A. (1995), preprint]. Finally, a refinement is given, related to counting labeled trees by number of inversions, of a result of Shi [Shi, J.-Y. (1986), Lecture Notes in Mathematics, no. 1179, Springer-Verlag] that a certain deformation of the braid arrangement has (n + 1)n-1 regions.

Calculating the probability of multitaxon evolutionary trees: bootstrappers Gambit.

The reconstruction of multitaxon trees from molecular sequences is confounded by the variety of algorithms and criteria used to evaluate trees, making it difficult to compare the results of different analyses. A global method of multitaxon phylogenetic reconstruction described here, Bootstrappers Gambit, can be used with any four-taxon algorithm, including distance, maximum likelihood, and parsimony methods. It incorporates a Bayesian-Jeffreys'-bootstrap analysis to provide a uniform probability-based criterion for comparing the results from diverse algorithms. To examine the usefulness of the method, the origin of the eukaryotes has been investigated by the analysis of ribosomal small subunit RNA sequences. Three common algorithms (paralinear distances, Jukes-Cantor distances, and Kimura distances) support the eocyte topology, whereas one (maximum parsimony) supports the archaebacterial topology, suggesting that the eocyte prokaryotes are the closest prokaryotic relatives of the eukaryotes.