Katsuhiko Mineta

Predicted expansion of the claudin multigene family

ZO‐1 and Claudin‐26 colocalize by fluorescence microscopy (View interaction)

Improved genome assembly and evidence-based global gene model set for the chordate Ciona intestinalis: new insight into intron and operon populations

BackgroundThe draft genome sequence of the ascidian Ciona intestinalis, along with associated gene models, has been a valuable research resource. However, recently accumulated expressed sequence tag (EST)/cDNA data have revealed numerous inconsistencies with the gene models due in part to intrinsic limitations in gene prediction programs and in part to the fragmented nature of the assembly.ResultsWe have prepared a less-fragmented assembly on the basis of scaffold-joining guided by paired-end EST and bacterial artificial chromosome (BAC) sequences, and BAC chromosomal in situ hybridization data. The new assembly (115.2 Mb) is similar in length to the initial assembly (116.7 Mb) but contains 1,272 (approximately 50%) fewer scaffolds. The largest scaffold in the new assembly incorporates 95 initial-assembly scaffolds. In conjunction with the new assembly, we have prepared a greatly improved global gene model set strictly correlated with the extensive currently available EST data. The total gene number (15,254) is similar to that of the initial set (15,582), but the new set includes 3,330 models at genomic sites where none were present in the initial set, and 1,779 models that represent fusions of multiple previously incomplete models. In approximately half, 5-ends were precisely mapped using 5-full-length ESTs, an important refinement even in otherwise unchanged models.ConclusionUsing these new resources, we identify a population of non-canonical (non-GT-AG) introns and also find that approximately 20% of Ciona genes reside in operons and that operons contain a high proportion of single-exon genes. Thus, the present dataset provides an opportunity to analyze the Ciona genome much more precisely than ever.

Ancient genome‐wide admixture extends beyond the current hybrid zone between Macaca fascicularis and M. mulatta

Macaca fascicularis and Macaca mulatta are two of the most commonly used laboratory macaques, yet their genetic differences at a genome‐wide level remain unclear. We analysed the multilocus DNA sequence data of 54 autosomal loci obtained from M. fascicularis samples from three different geographic origins and M. mulatta samples of Burmese origin. M. fascicularis shows high nucleotide diversity, four to five times higher than humans, and a strong geographic population structure between Indonesian‐Malaysian and Philippine macaques. The pattern of divergence and polymorphism between M. fascicularis and M. mulatta shows a footprint of genetic exchange not only within their current hybrid zone but also across a wider range for more than 1 million years. However, genetic admixture may not be a random event in the genome. Whereas randomly selected genic and intergenic regions have the same evolutionary dynamics between the species, some cytochrome oxidase P450 (CYP) genes (major chemical metabolizing genes and potential target genes for local adaptation) have a significantly larger species divergence than other genes. By surveying CYP3A5 gene sequences of more than a hundred macaques, we identified three nonsynonymous single nucleotide polymorphisms that were highly differentiated between the macaques. The mosaic pattern of species divergence in the genomes may be a consequence of genetic differentiation under ecological adaptation and may be a salient feature in the genomes of nascent species under parapatry.

Changes in mRNA Stability Associated with Cold Stress in Arabidopsis Cells

Control of mRNA half-life is a powerful strategy to adjust individual mRNA levels to various stress conditions, because the mRNA degradation rate controls not only the steady-state mRNA level but also the transition speed of mRNA levels. Here, we analyzed mRNA half-life changes in response to cold stress in Arabidopsis cells using genome-wide analysis, in which mRNA half-life measurements and transcriptome analysis were combined. Half-lives of average transcripts were determined to be elongated under cold conditions. Taking this general shift into account, we identified more than a thousand transcripts that were classified as relatively stabilized or relatively destabilized. The relatively stabilized class was predominantly observed in functional categories that included various regulators involved in transcriptional, post-transcriptional and post-translational processes. On the other hand, the relatively destabilized class was enriched in categories related to stress and hormonal response proteins, supporting the idea that rapid decay of mRNA is advantageous for swift responses to stress. In addition, pentatricopeptide repeat, cyclin-like F-box and Myb transcription factor protein families were significantly over-represented in the relatively destabilized class. The global analysis presented here demonstrates not only the importance of mRNA turnover control in the cold stress response but also several structural characteristics that might be important in the control of mRNA stability.

Structure and function of primitive immunoglobulin superfamily neural cell adhesion molecules: a lesson from studies on planarian

Precise wiring and proper remodeling of the neural network are essential for its normal function. The freshwater planarian is an attractive animal in which to study the formation and maintenance of the neural network due to its high regenerative capability and developmental plasticity. Although a recent study revealed that homologs of netrin and its receptors are required for regeneration and maintenance of the planarian central nervous system (CNS), the roles of cell adhesion in the formation and maintenance of the planarian neural network remain poorly understood. In the present study, we found primitive immunoglobulin superfamily cell adhesion molecules (IgCAMs) in a planarian that are homologous to vertebrate neural IgCAMs. We identified planarian orthologs of NCAM, L1CAM, contactin and DSCAM, and designated them DjCAM, DjLCAM, DjCTCAM and DjDSCAM, respectively. We further confirmed that they function as cell adhesion molecules using cell aggregation assays. DjCAM and DjDSCAM were found to be differentially expressed in the CNS. Functional analyses using RNA interference revealed that DjCAM is partly involved in axon formation, and that DjDSCAM plays crucial roles in neuronal cell migration, axon outgrowth, fasciculation and projection.

CIPRO 2.5: Ciona intestinalis protein database, a unique integrated repository of large-scale omics data, bioinformatic analyses and curated annotation, with user rating and reviewing functionality

The Ciona intestinalis protein database (CIPRO) is an integrated protein database for the tunicate species C. intestinalis. The database is unique in two respects: first, because of its phylogenetic position, Ciona is suitable model for understanding vertebrate evolution; and second, the database includes original large-scale transcriptomic and proteomic data. Ciona intestinalis has also been a favorite of developmental biologists. Therefore, large amounts of data exist on its development and morphology, along with a recent genome sequence and gene expression data. The CIPRO database is aimed at collecting those published data as well as providing unique information from unpublished experimental data, such as 3D expression profiling, 2D-PAGE and mass spectrometry-based large-scale analyses at various developmental stages, curated annotation data and various bioinformatic data, to facilitate research in diverse areas, including developmental, comparative and evolutionary biology. For medical and evolutionary research, homologs in humans and major model organisms are intentionally included. The current database is based on a recently developed KH model containing 36u2009034 unique sequences, but for higher usability it covers 89u2009683 all known and predicted proteins from all gene models for this species. Of these sequences, more than 10u2009000 proteins have been manually annotated. Furthermore, to establish a community-supported protein database, these annotations are open to evaluation by users through the CIPRO website. CIPRO 2.5 is freely accessible at http://cipro.ibio.jp/2.5.

Specific expression of Gsta4 in mouse cochlear melanocytes: a novel role for hearing and melanocyte differentiation

Mammalian pigment cells produce melanin as the main pigment. Melanocytes, one of the two types of mammalian pigment cells, differentiate from the neural crest and migrate to a variety of organs during development. Melanocytes exist not only in the skin but also in other sites such as the cochlea where they are essential for hearing. Mitfmi‐bw is one of the known recessive alleles of the mouse microphthalmia‐associated transcription factor (Mitf) locus, which is essential for the development of pigment cells. Homozygous Mitfmi‐bw/Mitfmi‐bw mice have a completely white coat with black eyes and are deaf due to the lack of melanocytes. By comparing gene expression profiles in cochleae of wild‐type and Mitfmi‐bw/Mitfmi‐bw mice, we now demonstrate the specific expression of glutathione S‐transferase alpha 4 (Gsta4) in the stria vascularis. Gsta4 encodes one of the cytosolic glutathione S‐transferases (GSTs) which participate in detoxification processes of many tissues. This gene is specifically expressed in intermediate cells of the stria vascularis, suggesting a novel function for cochlear melanocytes. Moreover, among mammalian pigment cells, expression of Gsta4 was restricted to cochlear melanocytes, suggesting that melanocytes in various tissues differentiate from one another depending on their location.

Exploring functionally related enzymes using radially distributed properties of active sites around the reacting points of bound ligands

BackgroundStructural genomics approaches, particularly those solving the 3D structures of many proteins with unknown functions, have increased the desire for structure-based function predictions. However, prediction of enzyme function is difficult because one member of a superfamily may catalyze a different reaction than other members, whereas members of different superfamilies can catalyze the same reaction. In addition, conformational changes, mutations or the absence of a particular catalytic residue can prevent inference of the mechanism by which catalytic residues stabilize and promote the elementary reaction. A major hurdle for alignment-based methods for prediction of function is the absence (despite its importance) of a measure of similarity of the physicochemical properties of catalytic sites. To solve this problem, the physicochemical features radially distributed around catalytic sites should be considered in addition to structural and sequence similarities.ResultsWe showed that radial distribution functions (RDFs), which are associated with the local structural and physicochemical properties of catalytic active sites, are capable of clustering oxidoreductases and transferases by function. The catalytic sites of these enzymes were also characterized using the RDFs. The RDFs provided a measure of the similarity among the catalytic sites, detecting conformational changes caused by mutation of catalytic residues. Furthermore, the RDFs reinforced the classification of enzyme functions based on conventional sequence and structural alignments.ConclusionsOur results demonstrate that the application of RDFs provides advantages in the functional classification of enzymes by providing information about catalytic sites.

CIPRO 2.5: Ciona intestinalis Protein Database - a unique integrated repository of large-scale omics data, bioinformatic analyses, and curated annotation, with ability for user rating and comments

CIPRO database (http://cipro.ibio.jp/2.5) is an integrated protein database for a tunicate species Ciona intestinalis that is part of the Urochordata. Although CIPRO provides proteomic and transcriptomic data on a single species, the animal is considered unique in the evolutionary tree, representing a possible origin of the vertebrates and is a good model for understanding chordate evolution, including human evolution. Furthermore, C. intestinalis has been one of the favorites of developmental biologists; therefore, a lot of amount of accumulated knowledge on its development, morphology, in addition to the recent genome sequence and gene expression data exists. The CIPRO database aims to collect published data and to present unique information, including the unpublished transcriptomic and proteomic data and human curated annotation, for the use of researchers in biology and bioinformatics. The current database contains 89,673 unique sequences covering all the proteins from all the gene models on this species; the number was reduced to 70,493 by similarity clustering. Of these sequences, more than 5,000 proteins are manually annotated based on the large-scale transcriptomic, proteomic and bioinformatic data. Those annotations can be subjected to be qualification by rating, curation, and comments by named and anonymous users through the web site of CIPRO database. Unique features of CIPRO database include: n n(i) Original experimental data Unpublished experimental data, including 2D-PAGE with the identified protein spots by protein mass fingerprint (PMF) MS analysis, expressions or localizations of protein and RNA across developmental stages and tissues, altogether summarized in a single chart for the comparison among status and methods. RNA expressions are observed by microarray and EST. Each protein is linked to an independent Ascidian Proteome Database summarizing large-scale MS-based proteomic analyses. n n(ii) Whole Ciona intestinalis proteome database Proteins across gene models are presented: all protein models derived from published gene models are incorporated, including Kyoto model (KG), KH (successor of KG model), PROCITS, JGIs versions 1 and 2, and Ensembl (version 58.2) are incorporated. Identical sequences across gene models are shown. n n(iii) Original comprehensive user-friendly interfaces Bioinformatic analyses and prediction results are summarized in pictures for grasp at a glance: homology search, cytolocalization, secondary structure prediction combined with modification sites, such include phosphorylation and three-dimensional structures. n n(iv) Comparative analysis data for disease association Comparison with human genome: map location of human homologues is graphically shown with associated disease information. Comparative data for other model organisms are also included. n n(v) Community-wide curation capability opened to users To facilitate progressive improvement of annotation by visited users, users can place additional annotation for the protein name and/or comments, which will be subjected to rating by the followed data viewers. To aid curation by wide community, information for literature and essence of matched motif patterns and other related protein information are shown with the links. n n(vi) Useful search facilities Various search methods are provided including blast homology, free text, partial sequence, protein mass fragment, and cross item searches.