Matsuyuki Shirota

ATTED-II in 2014: Evaluation of Gene Coexpression in Agriculturally Important Plants

ATTED-II (http://atted.jp) is a database of coexpressed genes that was originally developed to identify functionally related genes in Arabidopsis and rice. Herein, we describe an updated version of ATTED-II, which expands this resource to include additional agriculturally important plants. To improve the quality of the coexpression data for Arabidopsis and rice, we included more gene expression data from microarray and RNA sequencing studies. The RNA sequencing-based coexpression data now cover 94% of the Arabidopsis protein-encoding genes, representing a substantial increase from previously available microarray-based coexpression data (76% coverage). We also generated coexpression data for four dicots (soybean, poplar, grape and alfalfa) and one monocot (maize). As both the quantity and quality of expression data for the non-model species are generally poorer than for the model species, we verified coexpression data associated with these new species using multiple methods. First, the overall performance of the coexpression data was evaluated using gene ontology annotations and the coincidence of a genomic feature. Secondly, the reliability of each guide gene was determined by comparing coexpressed gene lists between platforms. With the expanded and newly evaluated coexpression data, ATTED-II represents an important resource for identifying functionally related genes in agriculturally important plants.

Blind prediction of interfacial water positions in CAPRI.

We report the first assessment of blind predictions of water positions at protein–protein interfaces, performed as part of the critical assessment of predicted interactions (CAPRI) community‐wide experiment. Groups submitting docking predictions for the complex of the DNase domain of colicin E2 and Im2 immunity protein (CAPRI Target 47), were invited to predict the positions of interfacial water molecules using the method of their choice. The predictions—20 groups submitted a total of 195 models—were assessed by measuring the recall fraction of water‐mediated protein contacts. Of the 176 high‐ or medium‐quality docking models—a very good docking performance per se—only 44% had a recall fraction above 0.3, and a mere 6% above 0.5. The actual water positions were in general predicted to an accuracy level no better than 1.5 Å, and even in good models about half of the contacts represented false positives. This notwithstanding, three hotspot interface water positions were quite well predicted, and so was one of the water positions that is believed to stabilize the loop that confers specificity in these complexes. Overall the best interface water predictions was achieved by groups that also produced high‐quality docking models, indicating that accurate modelling of the protein portion is a determinant factor. The use of established molecular mechanics force fields, coupled to sampling and optimization procedures also seemed to confer an advantage. Insights gained from this analysis should help improve the prediction of protein–water interactions and their role in stabilizing protein complexes. Proteins 2014; 82:620–632.

Lack of Transcription Triggers H3K27me3 Accumulation in the Gene Body.

Trimethylated H3K27 (H3K27me3) is associated with transcriptional repression, and its abundance in chromatin is frequently altered in cancer. However, it has remained unclear how genomic regions modified by H3K27me3 are specified and formed. We previously showed that downregulation of transcription by oncogenic Ras signaling precedes upregulation of H3K27me3 level. Here, we show that lack of transcription as a result of deletion of the transcription start site of a gene is sufficient to increase H3K27me3 content in the gene body. We further found that histone deacetylation mediates Ras-induced gene silencing and subsequent H3K27me3 accumulation. The H3K27me3 level increased gradually after Ras activation, requiring at least 35xa0days to achieve saturation. Such maximal accumulation of H3K27me3 was reversed by forced induction of transcription with the dCas9-activator system. Thus, our results indicate that changes in H3K27me3 level, especially in the body of a subset of genes, are triggered by changes in transcriptional activity itself.

Validation of multiple single nucleotide variation calls by additional exome analysis with a semiconductor sequencer to supplement data of whole-genome sequencing of a human population

BackgroundValidation of single nucleotide variations in whole-genome sequencing is critical for studying disease-related variations in large populations. A combination of different types of next-generation sequencers for analyzing individual genomes may be an efficient means of validating multiple single nucleotide variations calls simultaneously.ResultsHere, we analyzed 12 independent Japanese genomes using two next-generation sequencing platforms: the Illumina HiSeq 2500 platform for whole-genome sequencing (average depth 32.4×), and the Ion Proton semiconductor sequencer for whole exome sequencing (average depth 109×). Single nucleotide polymorphism (SNP) calls based on the Illumina Human Omni 2.5-8 SNP chip data were used as the reference. We compared the variant calls for the 12 samples, and found that the concordance between the two next-generation sequencing platforms varied between 83% and 97%.ConclusionsOur results show the versatility and usefulness of the combination of exome sequencing with whole-genome sequencing in studies of human population genetics and demonstrate that combining data from multiple sequencing platforms is an efficient approach to validate and supplement SNP calls.

Ion concentration-dependent ion conduction mechanism of a voltage-sensitive potassium channel.

Voltage-sensitive potassium ion channels are essential for life, but the molecular basis of their ion conduction is not well understood. In particular, the impact of ion concentration on ion conduction has not been fully studied. We performed several micro-second molecular dynamics simulations of the pore domain of the Kv1.2 potassium channel in KCl solution at four different ion concentrations, and scrutinized each of the conduction events, based on graphical representations of the simulation trajectories. As a result, we observed that the conduction mechanism switched with different ion concentrations: at high ion concentrations, potassium conduction occurred by Hodgkin and Keynes knock-on mechanism, where the association of an incoming ion with the channel is tightly coupled with the dissociation of an outgoing ion, in a one-step manner. On the other hand, at low ion concentrations, ions mainly permeated by a two-step association/dissociation mechanism, in which the association and dissociation of ions were not coupled, and occurred in two distinct steps. We also found that this switch was triggered by the facilitated association of an ion from the intracellular side within the channel pore and by the delayed dissociation of the outermost ion, as the ion concentration increased.

SAHG, a comprehensive database of predicted structures of all human proteins

Most proteins from higher organisms are known to be multi-domain proteins and contain substantial numbers of intrinsically disordered (ID) regions. To analyse such protein sequences, those from human for instance, we developed a special protein-structure-prediction pipeline and accumulated the products in the Structure Atlas of Human Genome (SAHG) database at http://bird.cbrc.jp/sahg. With the pipeline, human proteins were examined by local alignment methods (BLAST, PSI-BLAST and Smith–Waterman profile–profile alignment), global–local alignment methods (FORTE) and prediction tools for ID regions (POODLE-S) and homology modeling (MODELLER). Conformational changes of protein models upon ligand-binding were predicted by simultaneous modeling using templates of apo and holo forms. When there were no suitable templates for holo forms and the apo models were accurate, we prepared holo models using prediction methods for ligand-binding (eF-seek) and conformational change (the elastic network model and the linear response theory). Models are displayed as animated images. As of July 2010, SAHG contains 42u2009581 protein-domain models in approximately 24u2009900 unique human protein sequences from the RefSeq database. Annotation of models with functional information and links to other databases such as EzCatDB, InterPro or HPRD are also provided to facilitate understanding the protein structure-function relationships.

Isolated inclusion body myopathy caused by a multisystem proteinopathy–linked hnRNPA1 mutation

Objective: To identify the genetic cause of isolated inclusion body myopathy (IBM) with autosomal dominant inheritance in 2 families. Methods: Genetic investigations were performed using whole-exome and Sanger sequencing of the heterogeneous nuclear ribonucleoprotein A1 gene (hnRNPA1). The clinical and pathologic features of patients in the 2 families were evaluated with neurologic examinations, muscle imaging, and muscle biopsy. Results: We identified a missense p.D314N mutation in hnRNPA1, which is also known to cause familial amyotrophic lateral sclerosis, in 2 families with IBM. The affected individuals developed muscle weakness in their 40s, which slowly progressed toward a limb-girdle pattern. Further evaluation of the affected individuals revealed no apparent motor neuron dysfunction, cognitive impairment, or bone abnormality. The muscle pathology was compatible with IBM, lacking apparent neurogenic change and inflammation. Multiple immunohistochemical analyses revealed the cytoplasmic aggregation of hnRNPA1 in close association with autophagosomes and myonuclei. Furthermore, the aberrant accumulation was characterized by coaggregation with ubiquitin, sequestome-1/p62, valosin-containing protein/p97, and a variety of RNA-binding proteins (RBPs). Conclusions: The present study expands the clinical phenotype of hnRNPA1-linked multisystem proteinopathy. Mutations in hnRNPA1, and possibly hnRNPA2B1, will be responsible for isolated IBM with a pure muscular phenotype. Although the mechanisms underlying the selective skeletal muscle involvement remain to be elucidated, the immunohistochemical results suggest a broad sequestration of RBPs by the mutated hnRNPA1.

jMorp: Japanese Multi Omics Reference Panel

Abstract We developed jMorp, a new database containing metabolome and proteome data for plasma obtained from >5000 healthy Japanese volunteers from the Tohoku Medical Megabank Cohort Study, which is available at https://jmorp.megabank.tohoku.ac.jp. Metabolome data were measured by proton nuclear magnetic resonance (NMR) and liquid chromatography–mass spectrometry (LC–MS), while proteome data were obtained by nanoLC–MS. We released the concentration distributions of 37 metabolites identified by NMR, distributions of peak intensities of 257 characterized metabolites by LC–MS, and observed frequencies of 256 abundant proteins. Additionally, correlation networks for the metabolites can be observed using an interactive network viewer. Compared with some existing databases, jMorp has some unique features: (i) Metabolome data were obtained using a single protocol in a single institute, ensuring that measurement biases were significantly minimized; (ii) The database contains large-scale data for healthy volunteers with various health records and genome data and (iii) Correlations between metabolites can be easily observed using the graphical viewer. Metabolites data are becoming important intermediate markers for evaluating the health states of humans, and thus jMorp is an outstanding resource for a wide range of researchers, particularly those in the fields of medical science, applied molecular biology, and biochemistry.

Genomic analysis identifies masqueraders of full-term cerebral palsy

Cerebral palsy is a common, heterogeneous neurodevelopmental disorder that causes movement and postural disabilities. Recent studies have suggested genetic diseases can be misdiagnosed as cerebral palsy. We hypothesized that two simple criteria, that is, full‐term births and nonspecific brain MRI findings, are keys to extracting masqueraders among cerebral palsy cases due to the following: (1) preterm infants are susceptible to multiple environmental factors and therefore demonstrate an increased risk of cerebral palsy and (2) brain MRI assessment is essential for excluding environmental causes and other particular disorders.

Analyses of the general rule on residue pair frequencies in local amino acid sequences of soluble, ordered proteins

The amino acid sequences of soluble, ordered proteins with stable structures have evolved due to biological and physical requirements, thus distinguishing them from random sequences. Previous analyses have focused on extracting the features that frequently appear in protein substructures, such as α‐helix and β‐sheet, but the universal features of protein sequences have not been addressed. To clarify the differences between native protein sequences and random sequences, we analyzed 7368 soluble, ordered protein sequences, by inspecting the observed and expected occurrences of 400 amino acid pairs in local proximity, up to 10 residues along the sequence in comparison with their expected occurrence in random sequence. We found the trend that the hydrophobic residue pairs and the polar residue pairs are significantly decreased, whereas the pairs between a hydrophobic residue and a polar residue are increased. This trend was universally observed regardless of the secondary structure content but was not observed in protein sequences that include intrinsically disordered regions, indicating that it can be a general rule of protein foldability. The possible benefits of this rule are discussed from the viewpoints of protein aggregation and disorder, which are both caused by low‐complexity regions of hydrophobic or polar residues.