Comprehensive phosphoproteome analysis unravels the core signaling network that initiates the earliest synapse pathology in preclinical Alzheimer's disease brain.

Using a high-end mass spectrometry, we screened phosphoproteins and phosphopeptides in four types of Alzheimer's disease (AD) mouse models and human AD postmortem brains. We identified commonly changed phosphoproteins in multiple models and also determined phosphoproteins related to initiation of amyloid beta (Aß) deposition in the mouse brain. After confirming these proteins were also changed in and human AD brains, we put the proteins on experimentally verified protein-protein interaction databases. Surprisingly, most of the core phosphoproteins were directly connected, and they formed a functional network linked to synaptic spine formation. The change of the core network started at a preclinical stage even before histological Aß deposition. Systems biology analyses suggested that phosphorylation of myristoylated alanine-rich C-kinase substrate (MARCKS) by overactivated kinases including protein kinases C and calmodulin-dependent kinases initiates synapse pathology. Two-photon microscopic observation revealed recovery of abnormal spine formation in the AD model mice by targeting a core protein MARCKS or by inhibiting candidate kinases, supporting our hypothesis formulated based on phosphoproteome analysis.

XiP: a computational environment to create, extend and share workflows.

UNLABELLED: XiP (eXtensible integrative Pipeline) is a flexible, editable and modular environment with a user-friendly interface that does not require previous advanced programming skills to run, construct and edit workflows. XiP allows the construction of workflows by linking components written in both R and Java, the analysis of high-throughput data in grid engine systems and also the development of customized pipelines that can be encapsulated in a package and distributed. XiP already comes with several ready-to-use pipeline flows for the most common genomic and transcriptomic analysis and ∼300 computational components. AVAILABILITY: XiP is open source, freely available under the Lesser General Public License (LGPL) and can be downloaded from http://xip.hgc.jp.

IRView: a database and viewer for protein interacting regions.

UNLABELLED: Protein-protein interactions (PPIs) are mediated through specific regions on proteins. Some proteins have two or more protein interacting regions (IRs) and some IRs are competitively used for interactions with different proteins. IRView currently contains data for 3417 IRs in human and mouse proteins. The data were obtained from different sources and combined with annotated region data from InterPro. Information on non-synonymous single nucleotide polymorphism sites and variable regions owing to alternative mRNA splicing is also included. The IRView web interface displays all IR data, including user-uploaded data, on reference sequences so that the positional relationship between IRs can be easily understood. IRView should be useful for analyzing underlying relationships between the proteins behind the PPI networks. AVAILABILITY: IRView is publicly available on the web at http://ir.hgc.jp/

CSO validator: improving manual curation workflow for biological pathways.

SUMMARY: Manual curation and validation of large-scale biological pathways are required to obtain high-quality pathway databases. In a typical curation process, model validation and model update based on appropriate feedback are repeated and requires considerable cooperation of scientists. We have developed a CSO (Cell System Ontology) validator to reduce the repetition and time during the curation process. This tool assists in quickly obtaining agreement among curators and domain experts and in providing a consistent and accurate pathway database. AVAILABILITY: The tool is available on http://csovalidator.csml.org. CONTACT: masao@hgc.jp.

MIRACH: efficient model checker for quantitative biological pathway models.

UNLABELLED: Model checking is playing an increasingly important role in systems biology as larger and more complex biological pathways are being modeled. In this article we report the release of an efficient model checker MIRACH 1.0, which supports any model written in popular formats such as CSML and SBML. MIRACH is integrated with a Petri-net-based simulation engine, enabling efficient online (on-the-fly) checking. In our experiment, by using Levchenko et al. model, we reveal that timesaving gains by using MIRACH easily surpass 400% compared with its offline-based counterpart. AVAILABILITY AND IMPLEMENTATION: MIRACH 1.0 was developed using Java and thus executable on any platform installed with JDK 6.0 (not JRE 6.0) or later. MIRACH 1.0, along with its source codes, documentation and examples are available at http://sourceforge.net/projects/mirach/ under the LGPLv3 license.

Systems biology model repository for macrophage pathway simulation.

SUMMARY: The Macrophage Pathway Knowledgebase (MACPAK) is a computational system that allows biomedical researchers to query and study the dynamic behaviors of macrophage molecular pathways. It integrates the knowledge of 230 reviews that were carefully checked by specialists for their accuracy and then converted to 230 dynamic mathematical pathway models. MACPAK comprises a total of 24 009 entities and 12 774 processes and is described in the Cell System Markup Language (CSML), an XML format that runs on the Cell Illustrator platform and can be visualized with a customized Cytoscape for further analysis. AVAILABILITY: MACPAK can be accessed via an interactive web site at http://macpak.csml.org. The CSML pathway models are available under the Creative Commons license.

Cross-Border Transmissions of the Delta Substrain AY.29 During Tokyo Olympic and Paralympic Games.

Tokyo Olympic and Paralympic Games, postponed for the COVID-19 pandemic, were finally held in the summer of 2021. Just before the games, the Alpha variant was being replaced with the more contagious Delta variant. AY.4 substrain AY.29, which harbors two additional characteristic mutations of 5239C > T (NSP3 Y840Y) and 5514T > C (NSP3 V932A), emerged in Japan and became dominant in Tokyo by the time of the Olympic Games. Variants of SARS-CoV-2 genomes were performed to extract AY.29 Delta substrain samples with 5239C > T and 5514T > C. Phylogenetic analysis was performed to illustrate how AY.29 strains evolved and were introduced into countries abroad. Simultaneously, ancestral searches were performed for the overseas AY.29 samples to identify their origins in Japan using the maximum variant approach. As of January 10, 2022, 118 samples were identified in 20 countries. Phylogenetic analysis and ancestral searches identified 55 distinct introductions into those countries. The United States had 50 samples with 10 distinct introductions, and the United Kingdom had 13 distinct strains introduced in 18 samples. Other countries or regions with multiple introductions were Canada, Germany, South Korea, Hong Kong, Thailand, and the Philippines. Among the 20 countries, most European and North American countries have vaccination rates over 50% and sufficient genomic surveillances are conducted; transmissions seem contained. However, propagation to unvaccinated regions might have caused unfathomable damages. Since samples in those unvaccinated countries are also undersampled with a longer lead time for data sharing, it will take longer to grasp the whole picture. More rigorous departure screenings for the participants from the unvaccinated countries might have been necessary.

DA 1.0: parameter estimation of biological pathways using data assimilation approach.

SUMMARY: Data assimilation (DA) is a computational approach that estimates unknown parameters in a pathway model using time-course information. Particle filtering, the underlying method used, is a well-established statistical method that approximates the joint posterior distributions of parameters by using sequentially generated Monte Carlo samples. In this article, we report the release of Java-based software (DA 1.0) with an intuitive and user-friendly interface to allow users to carry out parameters estimation using DA. AVAILABILITY AND IMPLEMENTATION: DA 1.0 was developed using Java and thus would be executable on any platform installed with JDK 6.0 (not JRE 6.0) or later. DA 1.0 is freely available for academic users and can be launched or downloaded from http://da.csml.org.

Cell illustrator 4.0: a computational platform for systems biology.

Cell Illustrator is a software platform for Systems Biology that uses the concept of Petri net for modeling and simulating biopathways. It is intended for biological scientists working at bench. The latest version of Cell Illustrator 4.0 uses Java Web Start technology and is enhanced with new capabilities, including: automatic graph grid layout algorithms using ontology information; tools using Cell System Markup Language (CSML) 3.0 and Cell System Ontology 3.0; parameter search module; high-performance simulation module; CSML database management system; conversion from CSML model to programming languages (FORTRAN, C, C++, Java, Python and Perl); import from SBML, CellML, and BioPAX; and, export to SVG and HTML. Cell Illustrator employs an extension of hybrid Petri net in an object-oriented style so that biopathway models can include objects such as DNA sequence, molecular density, 3D localization information, transcription with frame-shift, translation with codon table, as well as biochemical reactions.

Host-parasite relations of bacteria and phages can be unveiled by oligostickiness, a measure of relaxed sequence similarity.

MOTIVATION: The recent metagenome analysis has been producing a large number of host-unassigned viruses. Although assigning viruses to their hosts is basically important not only for virology but also for prevention of epidemic, it has been a laborious and difficult task to date. The only effective method for this purpose has been to find them in a same microscopic view. Now, we tried a computational approach based on genome sequences of bacteria and phages, introducing a physicochemical parameter, SOSS (set of oligostickiness similarity score) derived from oligostickiness, a measure of binding affinity of oligonucleotides to template DNA. RESULTS: We could confirm host-parasite relationships of bacteria and their phages by SOSS analysis: all phages tested (25 species) had a remarkably higher SOSS value with its host than with unrelated bacteria. Interestingly, according to SOSS values, lysogenic phages such as lambda phage (host: Escherichia coli) or SPP1 (host: Bacillus subtilis) have distinctively higher similarity with its host than its non-lysogenic (excretive or virulent) ones such as fd and T4 (host: E.coli) or phages gamma and PZA (host: B.subtilis). This finding is very promising for assigning host-unknown viruses to its host. We also investigated the relationship in codon usage frequency or G+C content of genomes to interpret the phenomenon revealed by SOSS analysis, obtaining evidences which support the hypothesis that higher SOSS values resulted from the cohabitation in the same environment which may cause the common biased mutation. Thus, lysogenic phages which stay inside longer resemble the host.

Cell Illustrator 4.0: a computational platform for systems biology.

Cell Illustrator is a software platform for Systems Biology that uses the concept of Petri net for modeling and simulating biopathways. It is intended for biological scientists working at bench. The latest version of Cell Illustrator 4.0 uses Java Web Start technology and is enhanced with new capabilities, including: automatic graph grid layout algorithms using ontology information; tools using Cell System Markup Language (CSML) 3.0 and Cell System Ontology 3.0; parameter search module; high-performance simulation module; CSML database management system; conversion from CSML model to programming languages (FORTRAN, C, C++, Java, Python and Perl); import from SBML, CellML, and BioPAX; and, export to SVG and HTML. Cell Illustrator employs an extension of hybrid Petri net in an object-oriented style so that biopathway models can include objects such as DNA sequence, molecular density, 3D localization information, transcription with frame-shift, translation with codon table, as well as biochemical reactions.

Comparison of gene expression profiles produced by CAGE, illumina microarray and real time RT-PCR.

Several technologies are currently used for gene expression profiling, such as Real Time RT-PCR, microarray and CAGE (Cap Analysis of Gene Expression). CAGE is a recently developed method for constructing transcriptome maps and it has been successfully applied to analyzing gene expressions in diverse biological studies. The principle of CAGE has been developed to address specific issues such as determination of transcriptional starting sites, the study of promoter regions and identification of new transcripts. Here, we present both quantitative and qualitative comparisons among three major gene expression quantification techniques, namely: CAGE, illumina microarray and Real Time RT-PCR, by showing that the quantitative values of each method are not interchangeable, however, each of them has unique characteristics which render all of them essential and complementary. Understanding the advantages and disadvantages of each technology will be useful in selecting the most appropriate technique for a determined purpose.

Orengedokuto and berberine improve indomethacin-induced small intestinal injury via adenosine.

BACKGROUND: Recent endoscopic technology has revealed that small intestinal injury is a serious threat to patients receiving nonsteroidal anti-inflammatory drugs (NSAIDs). We previously showed that Japanese herbal medicine, Orengedokuto (OGT; Huang-Lian-Jie-Du-Tang in Chinese), protects mice from lethal indomethacin (IND)-induced enteropathy. To elucidate the mechanism of the protective effect of OGT, we performed microarray analyses and high power statistical analyses of microarray data using new bioinformatics tools. METHODS: Female BALB/c mice were subcutaneously injected with IND (20 mg/kg) once a day for 2 days. OGT-treated mice received a diet containing OGT from the first IND injection until the end of the experiment. Gene expression signals of small intestine were obtained with GeneChip. Analyses for overrepresentation of Gene Ontology categories were conducted using MetaGene Profiler (MGP) and the changes were visualized by Cell Illustrator Online (CIO). Furthermore, active ingredients of OGT were investigated. RESULTS: MGP and CIO suggested a critical role for the adenosine system, especially adenosine deaminase (ADA), a key enzyme of adenosine catabolism. Quantitative real time RT-PCR and in situ hybridization showed that OGT decreased the expression of ADA, which possibly resulted in the elevation of the anti-inflammatory nucleoside adenosine. Blockade of the adenosine A2a receptor abrogated the protective effect of OGT. Berberine, a major ingredient of OGT, suppressed ADA expression and reduced the incidence of lethality. CONCLUSIONS: OGT may prevent IND-induced enteropathy by decreasing ADA which results in the elevation of adenosine. Modulation of the adenosine system may be an efficient therapeutic strategy for NSAID-induced enteropathy.

ExonMiner: Web service for analysis of GeneChip Exon array data.

BACKGROUND: Some splicing isoform-specific transcriptional regulations are related to disease. Therefore, detection of disease specific splice variations is the first step for finding disease specific transcriptional regulations. Affymetrix Human Exon 1.0 ST Array can measure exon-level expression profiles that are suitable to find differentially expressed exons in genome-wide scale. However, exon array produces massive datasets that are more than we can handle and analyze on personal computer. RESULTS: We have developed ExonMiner that is the first all-in-one web service for analysis of exon array data to detect transcripts that have significantly different splicing patterns in two cells, e.g. normal and cancer cells. ExonMiner can perform the following analyses: (1) data normalization, (2) statistical analysis based on two-way ANOVA, (3) finding transcripts with significantly different splice patterns, (4) efficient visualization based on heatmaps and barplots, and (5) meta-analysis to detect exon level biomarkers. We implemented ExonMiner on a supercomputer system in order to perform genome-wide analysis for more than 300,000 transcripts in exon array data, which has the potential to reveal the aberrant splice variations in cancer cells as exon level biomarkers. CONCLUSION: ExonMiner is well suited for analysis of exon array data and does not require any installation of software except for internet browsers. What all users need to do is to access the ExonMiner URL http://ae.hgc.jp/exonminer. Users can analyze full dataset of exon array data within hours by high-level statistical analysis with sound theoretical basis that finds aberrant splice variants as biomarkers.

Targeting Tyro3 ameliorates a model of PGRN-mutant FTLD-TDP via tau-mediated synaptic pathology.

Mutations in the progranulin (PGRN) gene cause a tau pathology-negative and TDP43 pathology-positive form of frontotemporal lobar degeneration (FTLD-TDP). We generated a knock-in mouse harboring the R504X mutation (PGRN-KI). Phosphoproteomic analysis of this model revealed activation of signaling pathways connecting PKC and MAPK to tau prior to TDP43 aggregation and cognitive impairments, and identified PKCα as the kinase responsible for the early-stage tau phosphorylation at Ser203. Disinhibition of Gas6 binding to Tyro3 due to PGRN reduction results in activation of PKCα via PLCγ, inducing tau phosphorylation at Ser203, mislocalization of tau to dendritic spines, and spine loss. Administration of a PKC inhibitor, B-Raf inhibitor, or knockdown of molecules in the Gas6-Tyro3-tau axis rescues spine loss and cognitive impairment of PGRN-KI mice. Collectively, these results suggest that targeting of early-stage and aggregation-independent tau signaling represents a promising therapeutic strategy for this disease.

AYUMS: an algorithm for completely automatic quantitation based on LC-MS/MS proteome data and its application to the analysis of signal transduction.

BACKGROUND: Comprehensive description of the behavior of cellular components in a quantitative manner is essential for systematic understanding of biological events. Recent LC-MS/MS (tandem mass spectrometry coupled with liquid chromatography) technology, in combination with the SILAC (Stable Isotope Labeling by Amino acids in Cell culture) method, has enabled us to make relative quantitation at the proteome level. The recent report by Blagoev et al. (Nat. Biotechnol., 22, 1139-1145, 2004) indicated that this method was also applicable for the time-course analysis of cellular signaling events. Relative quatitation can easily be performed by calculating the ratio of peak intensities corresponding to differentially labeled peptides in the MS spectrum. As currently available software requires some GUI applications and is time-consuming, it is not suitable for processing large-scale proteome data. RESULTS: To resolve this difficulty, we developed an algorithm that automatically detects the peaks in each spectrum. Using this algorithm, we developed a software tool named AYUMS that automatically identifies the peaks corresponding to differentially labeled peptides, compares these peaks, calculates each of the peak ratios in mixed samples, and integrates them into one data sheet. This software has enabled us to dramatically save time for generation of the final report. CONCLUSION: AYUMS is a useful software tool for comprehensive quantitation of the proteome data generated by LC-MS/MS analysis. This software was developed using Java and runs on Linux, Windows, and Mac OS X. Please contact ayums@ims.u-tokyo.ac.jp if you are interested in the application. The project web page is http://www.csml.org/ayums/.

Cell fate simulation model of gustatory neurons with MicroRNAs double-negative feedback loop by hybrid functional Petri net with extension.

Biological regulatory networks have been extensively researched. Recently, the microRNA regulation has been analyzed and its importance has increasingly emerged. We have applied the Hybrid Functional Petri net with extension (HFPNe) model and succeeded in creating model biological pathways, e.g. metabolic pathways, gene regulatory networks, cell signaling networks, and cell-cell interaction models with one of the HFPNe implementations Cell Illustrator. Thus, we have applied HFPNe to model regulatory networks that involve a new key regulator microRNA. As a test case, we selected the cell fate determination model of two gustatory neurons of Caenorhabditis elegans-ASE left (ASEL) and ASE right (ASER). These neurons are morphologically bilaterally symmetric but physically asymmetric in function. Johnston et al. have suggested that their cell fate is determined by the double-negative feedback loop involving the lsy-6 and mir-273 microRNAs. Our simulation model confirms their hypothesis. In addition, other well-known mutants that are related with the double-negative feedback loop are also well-modeled. The new upstream regulator of lsy-6 (lsy-2) that is mentioned in another paper is also integrated into this model for the mechanism of switching between ASEL and ASER without any contradictions. Therefore, the HFPNe-based modeling will be one of the promising modeling methods and simulation architectures that illustrate microRNA regulatory networks.

Simple preparation of green fluorescent protein conjugated with ß-cyclodextrin in a site specific manner.

We have site-directedly linked a green fluorescent protein (GFP) variant and a ß-cyclodextrin (ß-CD) with a simple method to develop a basic complex for sophisticated supramolecules. We have confirmed ß-CD grafting on GFP with several methods including matrix-assisted laser desorption/ionization linear time-of-flight mass spectrometry (MALDI-TOF MS) without protease digestion and characterized the complex as well. In consideration of the resulting properties, the product we plainly and efficiently obtained could have applications related to sensing devices and drug delivery systems.

Molecular features of triple negative breast cancer cells by genome-wide gene expression profiling analysis.

Triple negative breast cancer (TNBC) has a poor outcome due to the lack of beneficial therapeutic targets. To clarify the molecular mechanisms involved in the carcinogenesis of TNBC and to identify target molecules for novel anticancer drugs, we analyzed the gene expression profiles of 30 TNBCs as well as 13 normal epithelial ductal cells that were purified by laser-microbeam microdissection. We identified 301 and 321 transcripts that were significantly upregulated and downregulated in TNBC, respectively. In particular, gene expression profile analyses of normal human vital organs allowed us to identify 104 cancer-specific genes, including those involved in breast carcinogenesis such as NEK2, PBK and MELK. Moreover, gene annotation enrichment analysis revealed prominent gene subsets involved in the cell cycle, especially mitosis. Therefore, we focused on cell cycle regulators, asp (abnormal spindle) homolog, microcephaly-associated (Drosophila) (ASPM) and centromere protein K (CENPK) as novel therapeutic targets for TNBC. Small-interfering RNA-mediated knockdown of their expression significantly attenuated TNBC cell viability due to G1 and G2/M cell cycle arrest. Our data will provide a better understanding of the carcinogenesis of TNBC and could contribute to the development of molecular targets as a treatment for TNBC patients.

PRD: A protein-RNA interaction database.

UNLABELLED: Although protein-RNA interactions (PRIs) are involved in various important cellular processes, compiled data on PRIs are still limited. This contrasts with protein-protein interactions, which have been intensively recorded in public databases and subjected to network level analysis. Here, we introduce PRD, an online database of PRIs, dispersed across several sources, including scientific literature. Currently, over 10,000 interactions have been stored in PRD using PSI-MI 2.5, which is a standard model for describing detailed molecular interactions, with an emphasis on gene level data. Users can browse all recorded interactions and execute flexible keyword searches against the database via a web interface. Our database is not only a reference of PRIs, but will also be a valuable resource for studying characteristics of PRI networks. AVAILABILITY: PRD can be freely accessed at http://pri.hgc.jp/