Yoshio Tateno’s research while affiliated with Daegu Gyeongbuk Institute of Science and Technology and other places

Orthologs are widely used for phylogenetic analysis of species; however, identifying genuine orthologs among distantly related species is challenging, because genes obtained through horizontal gene transfer (HGT) and out-paralogs derived from gene duplication before speciation are often present among the predicted orthologs. We developed a program, “Ortholog-Finder,” to obtain ortholog datasets for performing phylogenetic analysis by using all ORF data of species. The program includes 5 processes for minimizing the effects of HGT and out-paralogs in phylogeny construction: (1) HGT filtering: Genes derived from HGT could be detected and deleted from the initial sequence dataset by examining their base compositions. (2) Out-paralog filtering: Out-paralogs are detected and deleted from the dataset based on sequence similarity. (3) Classification of phylogenetic trees: Phylogenetic trees generated for ortholog candidates are classified as monophyletic or polyphyletic trees. (4) Tree splitting: Polyphyletic trees are bisected to obtain monophyletic trees and remove HGT genes and out-paralogs. (5) Threshold changing: Out-paralogs are further excluded from the dataset based on the difference in the similarity scores of genuine orthologs and out-paralogs. We examined how out-paralogs and HGTs affected phylogenetic trees constructed for species based on ortholog datasets obtained by Ortholog-Finder with the use of simulation data, and we determined the effects of confounding factors. We then used Ortholog-Finder in phylogeny construction for 12 gram-positive bacteria from 2 phyla and validated each node of the constructed tree by comparison with individually constructed ortholog trees.

The chicken domestication process represents a typical model of artificial selection, and gives significant insight into the general understanding of the influence of artificial selection on recognizable phenotypes. Two Japanese domesticated chicken varieties, the fighting cock (Shamo) and the long-crowing chicken (Naganakidori), have been selectively bred for dramatically different phenotypes. The former has been selected exclusively for aggressiveness and the latter for long crowing with an obedient sitting posture. To understand the particular mechanism behind these genetic changes during domestication, we investigated the degree of genetic differentiation in the aforementioned chickens, focusing on dopamine receptor D2, D3, and D4 genes. We studied other ornamental chickens such as Chabo chickens as a reference for comparison. When genetic differentiation was measured by an index of nucleotide differentiation (NST) newly devised in this study, we found that the NST value of DRD4 for Shamo (0.072) was distinctively larger than those of the other genes among the three populations, suggesting that aggressiveness has been selected for in Shamo by collecting a variety of single nucleotide polymorphisms. In addition, we found that in DRD4 in Naganakidori, there is a deletion variant of one proline at the 24th residue in the repeat of nine prolines of exon 1. We thus conclude that artificial selection has operated on these different kinds of genetic variation in the DRD4 genes of Shamo and Naganakidori so strongly that the two domesticated varieties have differentiated to obtain their present opposite features in a relatively short period of time.

To study the male and female lineages of East Asian and European humans, we have sequenced 25 short tandem repeat markers on 453 Y-chromosomes and collected sequences of 72 complete mitochondrial genomes to construct independent phylogenetic trees for male and female lineages. The results indicate that East Asian individuals fall into two clades, one that includes East Asian individuals only and a second that contains East Asian and European individuals. Surprisingly, the European individuals did not form an independent clade, but branched within in the East Asians. We then estimated the divergence time of the root of the European clade as ∼41,000 years ago. These data indicate that, contrary to traditional views, Europeans diverged from East Asians around that time. We also address the origin of the Ainu lineage in northern Japan.

Lactobacillus gasseri ATCC33323(T) expresses four enzymes showing phospho-β-galactosidase activity (LacG1, LacG2, Pbg1 and Pbg2). We previously reported the purification and characterization of two phospho-β-galactosidases (Pbg1 and Pbg2) from Lactobacillus gasseri JCM1031 cultured in lactose medium. Here we aimed to characterize LacG1 and LacG2, and classify the four enzymes into 'phospho-β-galactosidase' or 'phospho-β-glucosidase.' LacG1 and recombinant LacG2 (rLacG2), from Lb. gasseri ATCC33323(T), were purified to homogeneity using column chromatography. Kinetic experiments were performed using sugar substrates, o-nitrophenyl-β-D-galactopyranoside 6-phosphate (ONPGal-6P) and o-nitrophenyl-β-D-glucopyranoside 6-phosphate (ONPGlc-6P), synthesized in our laboratory. LacG1 and rLacG2 exhibited high k(cat)/K(m) values for ONPGal-6P as compared with Pbg1 and Pbg2. The V(max) values for ONPGal-6P were higher than phospho-β-galactosidases previously purified and characterized from several lactic acid bacteria. A phylogenetic tree analysis showed that LacG1 and LacG2 belong to the phospho-β-galactosidase cluster and Pbg1 and Pbg2 belong to the phospho-β-glucosidase cluster. Our data suggest two phospho-β-galactosidase, LacG1 and LacG2, are the primary enzymes for lactose utilization in Lb. gasseri ATCC33323(T). We propose a reclassification of Pbg1 and Pbg2 as phospho-β-glucosidase.

Unlabelled: Horizontal gene transfer (HGT) is a common event in prokaryotic evolution. Therefore, it is very important to consider HGT in the study of molecular evolution of prokaryotes. This is true also for conducting computer simulations of their molecular phylogeny because HGT is known to be a serious disturbing factor for estimating their correct phylogeny. To the best of our knowledge, no existing computer program has generated a phylogenetic tree with HGT from an original phylogenetic tree. We developed a program called HGT-Gen that generates a phylogenetic tree with HGT on the basis of an original phylogenetic tree of a protein or gene. HGT-Gen converts an operational taxonomic unit or a clade from one place to another in a given phylogenetic tree. We have also devised an algorithm to compute the average length between any pair of branches in the tree. It defines and computes the relative evolutionary time to normalize evolutionary time for each lineage. The algorithm can generate an HGT between a pair of donor and acceptor lineages at the same evolutionary time. HGT-Gen is used with a sequence-generating program to evaluate the influence of HGT on the molecular phylogeny of prokaryotes in a computer simulation study. Availability: The database is available for free at http://www.grl.shizuoka.ac.jp/˜thoriike/HGT-Gen.html.

Evolutionary conserved and novel tick-specific R. microplus miRNAs expressed during various life stages and selected organs. Rhipicephalus microplus miRNAs with known counterpart in D. melanogaster, I. scapularis or other relevant species are shown with reference miRBase IDs. The number of short reads overlapping each miRNA locus, the corresponding percentage fraction of the total number of reads overlapping all miRNA loci for each sample and the normalized reads per million (RPM) counts for each miRNA in each sample are shown. N.A. = Not available in miRBase and/or not cloned previously.

Real Time PCR quantification of selected R. microplus miRNAs. Cattle tick miRNAs identically conserved in D. melanogaster were selected and specific Drosophila miRNA TaqMan probes (Applied Biosystems) were used to amplify these in eggs, larvae and female tick samples.

Evolutionary analysis of R. microplus miRNAs. Evidence supporting R. microplus miRNAs conservation since the A) Nephrozoan, B) Protostomian, C) Arthropoda and D) Ixodidae ancestors are presented as well as E) miRNA nucleotide variants unique to R. microplus based on currently available datasets (miRBase release 17). Ancestral sequences were determined using a Maximum Likelihood method [49,50] under the Junkes-Cantor model [63] using a tree topology similar to that shown in Figure 5A (modified from [21]). Evolutionary analyses were conducted in MEGA5 package [51]. Key miRNAs supporting each ancestral state were selected. Base-substitutions compared to predict ancestral sequences are highlighted in red fonts. Green fonts denote bases for which an ancestral state could not be determined. R. microplus miRNAs are denoted by bold fonts.

Comparison of changes in tick miRNA expression between larvae and frustrated larvae samples. Statistically significant changes in R. microplus miRNA expression between libraries was calculated using three statistical tests including Pairwise Audic & Claverie test (AC), Pairwise Chi sq. test (Chi2 × 2) and Multiple Chi sq. test (Chi) as previously described by Romualdi et al. (2003). Bonferroni correction was applied and significant thresholds of 9.62E-05 for AC, Chi2 × 2 and Chi was defined. Bold fonts denote values that are statistically significant. RPM = Reads per million; N.A. = Not applicable.