Goro Terai

Genome sequencing and analysis of Aspergillus oryzae

The genome of Aspergillus oryzae, a fungus important for the production of traditional fermented foods and beverages in Japan, has been sequenced. The ability to secrete large amounts of proteins and the development of a transformation system have facilitated the use of A. oryzae in modern biotechnology. Although both A. oryzae and Aspergillus flavus belong to the section Flavi of the subgenus Circumdati of Aspergillus, A. oryzae, unlike A. flavus, does not produce aflatoxin, and its long history of use in the food industry has proved its safety. Here we show that the 37-megabase (Mb) genome of A. oryzae contains 12,074 genes and is expanded by 7–9 Mb in comparison with the genomes of Aspergillus nidulans and Aspergillus fumigatus. Comparison of the three aspergilli species revealed the presence of syntenic blocks and A. oryzae-specific blocks (lacking synteny with A. nidulans and A. fumigatus) in a mosaic manner throughout the genome of A. oryzae. The blocks of A. oryzae-specific sequence are enriched for genes involved in metabolism, particularly those for the synthesis of secondary metabolites. Specific expansion of genes for secretory hydrolytic enzymes, amino acid metabolism and amino acid/sugar uptake transporters supports the idea that A. oryzae is an ideal microorganism for fermentation.

fRNAdb: a platform for mining/annotating functional RNA candidates from non-coding RNA sequences

There are abundance of transcripts that code for no particular protein and that remain functionally uncharacterized. Some of these transcripts may have novel functions while others might be junk transcripts. Unfortunately, the experimental validation of such transcripts to find functional non-coding RNA candidates is very costly. Therefore, our primary interest is to computationally mine candidate functional transcripts from a pool of uncharacterized transcripts. We introduce fRNAdb: a novel database service that hosts a large collection of non-coding transcripts including annotated/non-annotated sequences from the H-inv database, NONCODE and RNAdb. A set of computational analyses have been performed on the included sequences. These analyses include RNA secondary structure motif discovery, EST support evaluation, cis-regulatory element search, protein homology search, etc. fRNAdb provides an efficient interface to help users filter out particular transcripts under their own criteria to sort out functional RNA candidates. fRNAdb is available at

The Functional RNA Database 3.0: databases to support mining and annotation of functional RNAs

We developed a pair of databases that support two important tasks: annotation of anonymous RNA transcripts and discovery of novel non-coding RNAs. The database combo is called the Functional RNA Database and consists of two databases: a rewrite of the original version of the Functional RNA Database (fRNAdb) and the latest version of the UCSC GenomeBrowser for Functional RNA. The former is a sequence database equipped with a powerful search function and hosts a large collection of known/predicted non-coding RNA sequences acquired from existing databases as well as novel/predicted sequences reported by researchers of the Functional RNA Project. The latter is a UCSC Genome Browser mirror with large additional custom tracks specifically associated with non-coding elements. It also includes several functional enhancements such as a presentation of a common secondary structure prediction at any given genomic window ⩽500 bp. Our GenomeBrowser supports user authentication and user-specific tracks. The current version of the fRNAdb is a complete rewrite of the former version, hosting a larger number of sequences and with a much friendlier interface. The current version of UCSC GenomeBrowser for Functional RNA features a larger number of tracks and richer features than the former version. The databases are available at http://www.ncrna.org/.

DBTBS: a database of Bacillus subtilis promoters and transcription factors

With the completion of the determination of its entire genome sequence, one of the next major targets of Bacillus subtilis genomics is to clarify the whole gene regulatory network. To this end, the results of systematic experiments should be compared with the rich source of individual experimental results accumulated so far. Thus, we constructed a database of the upstream regulatory information of B.subtilis (DBTBS). The current version was constructed by surveying 291 references and contains information on 90 binding factors and 403 promoters. For each promoter, all of its known cis-elements are listed according to their positions, while these cis-elements are aligned to illustrate their consensus sequence for each transcription factor. All probable transcription factors coded in the genome were classified with the Pfam motifs. Using this database, we compared the character of B.subtilis promoters with that of Escherichia coli promoters. Our database is accessible at http://elmo.ims.u-tokyo.ac.jp/dbtbs/.

CapR: revealing structural specificities of RNA-binding protein target recognition using CLIP-seq data

RNA-binding proteins (RBPs) bind to their target RNA molecules by recognizing specific RNA sequences and structural contexts. The development of CLIP-seq and related protocols has made it possible to exhaustively identify RNA fragments that bind to RBPs. However, no efficient bioinformatics method exists to reveal the structural specificities of RBP–RNA interactions using these data. We present CapR, an efficient algorithm that calculates the probability that each RNA base position is located within each secondary structural context. Using CapR, we demonstrate that several RBPs bind to their target RNA molecules under specific structural contexts. CapR is available at https://sites.google.com/site/fukunagatsu/software/capr.

Ustiloxins, fungal cyclic peptides, are ribosomally synthesized in Ustilaginoidea virens

MOTIVATION Ustiloxins A and B are toxic cyclic tetrapeptides, Tyr-Val/Ala-Ile-Gly (Y-V/A-I-G), that were originally identified from Ustilaginoidea virens, a pathogenic fungus affecting rice plants. Contrary to our report that ustiloxin B is ribosomally synthesized in Aspergillus flavus, a recent report suggested that ustiloxins are synthesized by a non-ribosomal peptide synthetase in U.virens. Thus, we analyzed the U.virens genome, to identify the responsible gene cluster. RESULTS The biosynthetic gene cluster was identified from the genome of U.virens based on homologies to the ribosomal peptide biosynthetic gene cluster for ustiloxin B identified from A.flavus. It contains a gene encoding precursor protein having five Tyr-Val-Ile-Gly and three Tyr-Ala-Ile-Gly motifs for ustiloxins A and B, respectively, strongly indicating that ustiloxins A and B from U.virens are ribosomally synthesized. AVAILABILITY AND IMPLEMENTATION Accession codes of the U.virens and A.flavus gene clusters in NCBI are BR001221 and BR001206, respectively. Supplementary data are available at Bioinformatics online.

Phylogeny of Organisms Investigated by the Base-Pair Changes in the Stem Regions of Small and Large Ribosomal Subunit RNAs

Abstract. In order to obtain the evolutionary distance data that are as purely additive as possible, we have developed a novel method for evaluating the evolutionary distances from the base-pair changes in stem regions of ribosomal RNAs (rRNAs). The application of this method to small-subunit (SSU) and large-subunit (LSU) rRNAs provides the distance data, with which both the unweighted pair group method of analysis and the neighbor-joining method give almost the same tree topology of most organisms except for some Protoctista, thermophilic bacteria, parasitic organisms, and endosymbionts. Although the evolutionary distances calculated with LSU rRNAs are somewhat longer than those with SSU rRNAs, the difference, probably due to a slight difference in functional constraint, is substantially decreased when the distances are converted into the divergence times of organisms by the measure of the time scale estimated in each type of rRNAs. The divergence times of main branches agree fairly well with the geological record of organisms, at least after the appearance of oxygen-releasing photosynthesis, although the divergence times of Eukaryota, Archaebacteria, and Eubacteria are somewhat overestimated in comparison with the geological record of Earth formation. This result is explained by considering that the mutation rate is determined by the accumulation of misrepairs for DNA damage caused by radiation and that the effect of radiation had been stronger before the oxygen molecules became abundant in the atmosphere of the Earth.

Prediction of co-regulated genes in Bacillus subtilis on the basis of upstream elements conserved across three closely related species

BackgroundIdentification of co-regulated genes is essential for elucidating transcriptional regulatory networks and the function of uncharacterized genes. Although co-regulated genes should have at least one common sequence element, it is generally difficult to identify these genes from the presence of this element because it is very easily obscured by noise. To overcome this problem, we used conserved information from three closely related species: Bacillus subtilis,B. halodurans and B. stearothermophilus.ResultsEven though such species have a limited number of clearly orthologous genes, we obtained 1,884 phylogenetically conserved elements from the upstream intergenic regions of 1,568 B. subtilis genes. Similarity between these elements was used to cluster these genes. No other a priori knowledge on genes and elements was used. We could identify some genes known or suggested to be regulated by a common transcription factor as well as genes regulated by a common attenuation effector.ConclusionsWe confirmed that our method generates relatively few false positives in clusters with higher scores and that general elements such as -35/-10 boxes and Shine-Dalgarno sequence are not major obstacles. Moreover, we identified some plausible additional members of groups of known co-regulated genes. Thus, our approach is promising for exploring potentially co-regulated genes.

Predicting rules on organization of cis-regulatory elements, taking the order of elements into account

MOTIVATION In eukaryotes, rules regarding organization of cis-regulatory elements are complex. They sometimes govern multiple kinds of elements and positional restrictions on elements. RESULTS We propose a method for detecting rules, by which the order of elements is restricted. The order restriction is expressed as element patterns. We extract all the element patterns that occur in promoter regions of at least the specified number of genes. Then, we find significant patterns based on the expression similarity of genes with promoter regions containing each of the extracted patterns. When we applied our method to Saccharomyces cerevisiae, we detected significant patterns overlooked by previous methods, thus demonstrating the utility of our method for analyses of eukaryotic gene regulation. We also suggest that several types of element organization exist: (i) those in which only the order of elements is important, (ii) order and distance both are important and (iii) only the combination of elements is important. AVAILABILITY The program for extracting element patterns is available upon request.

Discovery of short pseudogenes derived from messenger RNAs

More than 40% of the human genome is generated by retrotransposition, a series of in vivo processes involving reverse transcription of RNA molecules and integration of the transcripts into the genomic sequence. The mechanism of retrotransposition, however, is not fully understood, and additional genomic elements generated by retrotransposition may remain to be discovered. Here, we report that the human genome contains many previously unidentified short pseudogenes generated by retrotransposition of mRNAs. Genomic elements generated by non-long terminal repeat retrotransposition have specific sequence signatures: a poly-A tract that is immediately downstream and a pair of duplicated sequences, called target site duplications (TSDs), at either end. Using a new computer program, TSDscan, that can accurately detect pseudogenes based on the presence of the poly-A tract and TSDs, we found 654 short (≤300 bp), previously unknown pseudogenes derived from mRNAs. Comprehensive analyses of the pseudogenes that we identified and their parent mRNAs revealed that the pseudogene length depends on the parent mRNA length: long mRNAs generate more short pseudogenes than do short mRNAs. To explain this phenomenon, we hypothesize that most long mRNAs are truncated before they are reverse transcribed. Truncated mRNAs would be rapidly degraded during reverse transcription, resulting in the generation of short pseudogenes.