クラスタリング指標における新しい評価基準のアイデア

第14回情報メディア学会研究大会で発表したポスター（2015年6月27日 同志社大学 今出川キャンパス 良心館

A genome-wide gain-of-function analysis of rice genes using the FOX-hunting system

Funding Information: Acknowledgements This work was supported by a grant from the Ministry of Agriculture, Forestry and Fisheries of Japan (Green Technology Project EF-1004). We are grateful to Dr. Takuji Sasaki for his encouragement throughout the project and his excellent advice on the improvement of this manuscript, and to Dr. Shoshi Kikuchi for providing useful information on rice FL-cDNAs. We thank Professors Kokichi Hinata, Atsushi Hirai, Hiroshi Kamada and Masashi Ugaki for their encouragement, critical comments and helpful suggestions, and Drs. Hisato Okuizumi and Hiroyuki Kawahigashi for their administrative support throughout the project. We also thank Mayumi Akagawa, Hiroko Abe, Keiko Mori, Etsuko Sugai, Yumiko Nakane, Ken-ichi Watanabe, Mayumi Takeya, and Kana Miyata for their technical assistance; the members of the Technical Support Section of the National Institute of Agrobiological Sciences for their help in the care of the FOX-rice plants; Haruko Onodera and Kazuko Ono for their technical assistance and advice on rice transformation; Inplanta Innovations Inc. for their technical help on the construction of theThe latest report has estimated the number of rice genes to be ∼32 000. To elucidate the functions of a large population of rice genes and to search efficiently for agriculturally useful genes, we have been taking advantage of the Full-length cDNA Over-eXpresser (FOX) gene-hunting system. This system is very useful for analyzing various gain-of-function phenotypes from large populations of transgenic plants overexpressing cDNAs of interest and others with unknown or important functions. We collected the plasmid DNAs of 13 980 independent full-length cDNA (FL-cDNA) clones to produce a FOX library by placing individual cDNAs under the control of the maize Ubiquitin-1 promoter. The FOX library was transformed into rice by Agrobacterium-mediated high-speed transformation. So far, we have generated approximately 12 000 FOX-rice lines. Genomic PCR analysis indicated that the average number of FL-cDNAs introduced into individual lines was 1.04. Sequencing analysis of the PCR fragments carrying FL-cDNAs from 8615 FOX-rice lines identified FL-cDNAs in 8225 lines, and a database search classified the cDNAs into 5462 independent ones. Approximately 16.6% of FOX-rice lines examined showed altered growth or morphological characteristics. Three super-dwarf mutants overexpressed a novel gibberellin 2-oxidase gene, confirming the importance of this system. We also show here the other morphological alterations caused by individual FL-cDNA expression. These dominant phenotypes should be valuable indicators for gene discovery and functional analysis.publishersversionPeer reviewe

A Novel Method of Characterizing Genetic Sequences: Genome Space with Biological Distance and Applications

Most existing methods for phylogenetic analysis involve developing an evolutionary model and then using some type of computational algorithm to perform multiple sequence alignment. There are two problems with this approach: (1) different evolutionary models can lead to different results, and (2) the computation time required for multiple alignments makes it impossible to analyse the phylogeny of a whole genome. This motivates us to create a new approach to characterize genetic sequences.To each DNA sequence, we associate a natural vector based on the distributions of nucleotides. This produces a one-to-one correspondence between the DNA sequence and its natural vector. We define the distance between two DNA sequences to be the distance between their associated natural vectors. This creates a genome space with a biological distance which makes global comparison of genomes with same topology possible. We use our proposed method to analyze the genomes of the new influenza A (H1N1) virus, human rhinoviruses (HRV) and mammalian mitochondrial. The result shows that a triple-reassortant swine virus circulating in North America and the Eurasian swine virus belong to the lineage of the influenza A (H1N1) virus. For the HRV and mammalian mitochondrial genomes, the results coincide with biologists' analyses.Our approach provides a powerful new tool for analyzing and annotating genomes and their phylogenetic relationships. Whole or partial genomes can be handled more easily and more quickly than using multiple alignment methods. Once a genome space has been constructed, it can be stored in a database. There is no need to reconstruct the genome space for subsequent applications, whereas in multiple alignment methods, realignment is needed to add new sequences. Furthermore, one can make a global comparison of all genomes simultaneously, which no other existing method can achieve

モンゴルザイライヤギ 3シュウダン ノ トウキタイジュウソンモウリツ ト カシミヤモウセイサンリョウ

モンゴル国における優良ヤギ品種の確立を目的としてカシミヤ毛生産関連遺伝子のクローニングを試みている。その研究対象のリソース・ファミリーとしてBayandelger(BD),Gobi Gurvan Saikhan(GGS)およびZalaajinst White(ZW)の3集団をモンゴル国内に維持している。今回,本ヤギ集団における体重の加齢および季節的変動,並びにカシミヤ毛生産量からそれらの品種特性を調査した結果,以下の成績が得られた。1. GGS, ZW, BD集団の同一個体の冬季間における体重の損耗率を比較した結果,4歳齢の春にはGGSとZW集団には有意差は認められなかったが,この2集団とBD集団間に有意差が認められた(p<0.05～0.01)。体重損耗率はBD集団で最も低く23.70%で,次いでGGS集団の24.90%であった。最も体重の重かったZW集団は冬季間の損耗が36.06%で3集団の中で最も大きかった。2. GGS, ZWおよびBDのカシミヤ毛生産量は,それぞれ333.11±8.32g, 268.07±12.30gおよび222.86±5.32gであった。カシミヤ毛の直径は16-19,15-16.5および12-15μmであることから,BD集団のカシミヤ毛が最も細く,毛色形質を考慮すると優良な品種と判断された。冬季間の体重損耗率が最も少ないのは,細い毛による体温保温効果と関係しているものと推察された。3. BD集団におけるカシミヤ毛の長さ(5.68±0.39cm),直径(16.53±0.10μm)および毛生産量(222.86±5.32g)のそれぞれの間における相関では,長さと生産量の間に強い相関(0.747)が認められた。To establish excellent goat breeds in Mongolia, we have performed the cloning of genes related to cashmere production. As a genetic pool for this purpose, we have maintained Bayandelger (BD), Gobi Gurvan Saikhan (GGS), and Zalaajinst White (ZW) in Mongolia. In this study, the breed traits of the 3 goat populations were evaluated based on age-related and seasonal changes in body weight and cashmere production, and the following results were obtained. 1. Weight loss during winter was compared among individuals in the GGS, ZW, and BD populations. No significant difference was noted between GGS and ZW, but a significant difference was observed between the two populations and BD (p<0.05-0.01) in spring at the age of 4 years. Weight loss was the lowest in BD (23.70%), followed by GGS (24.90%). The heaviest ZW showed the greatest weight loss during winter (36.06%) among the 3 populations. 2. The cashmere yields in GGS, ZW and BD were 333.11±8.32, 268.07±12.30, and 222.86±5.32g, respectively, and the cashmere diameters in these populations were 16-19, 15-16.5 and 12-15μm, respectively. Therefore, considering the hair color trait, BD, with the thinnest cashmere, may be an excellent breed. The lowest weight loss during winter seen in this population may be associated with the heat-retaining effects of the thin cashmere. 3. In BD, the correlation coefficients between cashmere length (5.68±0.39cm) and production (222.86±5.32g), and between cashmere diameter (16.53±0.10μm) and production were 0.747 and 0.137, respectively, showing a marked correlation between length and production

LDL-Induced Impairment of Human Vascular Smooth Muscle Cells Repair Function Is Reversed by HMG-CoA Reductase Inhibition

Growing human atherosclerotic plaques show a progressive loss of vascular smooth muscle cells (VSMC) becoming soft and vulnerable. Lipid loaded-VSMC show impaired vascular repair function and motility due to changes in cytoskeleton proteins involved in cell-migration. Clinical benefits of statins reducing coronary events have been related to repopulation of vulnerable plaques with VSMC. Here, we investigated whether HMG-CoA reductase inhibition with rosuvastatin can reverse the effects induced by atherogenic concentrations of LDL either in the native (nLDL) form or modified by aggregation (agLDL) on human VSMC motility. Using a model of wound repair, we showed that treatment of human coronary VSMC with rosuvastatin significantly prevented (and reversed) the inhibitory effect of nLDL and agLDL in the repair of the cell depleted areas. In addition, rosuvastatin significantly abolished the agLDL-induced dephosphorylation of myosin regulatory light chain as demonstrated by 2DE-electrophoresis and mass spectrometry. Besides, confocal microscopy showed that rosuvastatin enhances actin-cytoskeleton reorganization during lipid-loaded-VSMC attachment and spreading. The effects of rosuvastatin on actin-cytoskeleton dynamics and cell migration were dependent on ROCK-signalling. Furthermore, rosuvastatin caused a significant increase in RhoA-GTP in the cytosol of VSMC. Taken together, our study demonstrated that inhibition of HMG-CoA reductase restores the migratory capacity and repair function of VSMC that is impaired by native and aggregated LDL. This mechanism may contribute to the stabilization of lipid-rich atherosclerotic plaques afforded by statins

The ventral anterior temporal lobe has a necessary role in exception word reading

An influential account of reading holds that words with exceptional spelling-to-sound correspondences (e.g., PINT) are read via activation of their lexical-semantic representations, supported by the anterior temporal lobe (ATL). This account has been inconclusive because it is based on neuropsychological evidence, in which lesion-deficit relationships are difficult to localize precisely, and functional neuroimaging data, which is spatially precise but cannot demonstrate whether the ATL activity is necessary for exception word reading. To address these issues, we used a technique with good spatial specificity - repetitive transcranial magnetic stimulation (rTMS) - to demonstrate a necessary role of ATL in exception word reading. Following rTMS to left ventral ATL, healthy Japanese adults made more regularization errors in reading Japanese exception words. We successfully simulated these results in a computational model in which exception word reading was underpinned by semantic activations. The ATL is critically and selectively involved in reading exception words

Guidelines for the use and interpretation of assays for monitoring autophagy (3rd edition)

In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. For example, a key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process versus those that measure fl ux through the autophagy pathway (i.e., the complete process including the amount and rate of cargo sequestered and degraded). In particular, a block in macroautophagy that results in autophagosome accumulation must be differentiated from stimuli that increase autophagic activity, defi ned as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (inmost higher eukaryotes and some protists such as Dictyostelium ) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the fi eld understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. It is worth emphasizing here that lysosomal digestion is a stage of autophagy and evaluating its competence is a crucial part of the evaluation of autophagic flux, or complete autophagy. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. Along these lines, because of the potential for pleiotropic effects due to blocking autophagy through genetic manipulation it is imperative to delete or knock down more than one autophagy-related gene. In addition, some individual Atg proteins, or groups of proteins, are involved in other cellular pathways so not all Atg proteins can be used as a specific marker for an autophagic process. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field