Yoshiharu Yamamoto

Widespread translational inhibition by plant miRNAs and siRNAs

High complementarity between plant microRNAs (miRNAs) and their messenger RNA targets is thought to cause silencing, prevalently by endonucleolytic cleavage. We have isolated Arabidopsis mutants defective in miRNA action. Their analysis provides evidence that plant miRNA–guided silencing has a widespread translational inhibitory component that is genetically separable from endonucleolytic cleavage. We further show that the same is true of silencing mediated by small interfering RNA (siRNA) populations. Translational repression is effected in part by the ARGONAUTE proteins AGO1 and AGO10. It also requires the activity of the microtubule-severing enzyme katanin, implicating cytoskeleton dynamics in miRNA action, as recently suggested from animal studies. Also as in animals, the decapping component VARICOSE (VCS)/Ge-1 is required for translational repression by miRNAs, which suggests that the underlying mechanisms in the two kingdoms are related.

Identification of Arabidopsis Genes Regulated by High Light–Stress Using cDNA Microarray¶

Abstract In plants, excess light has the potential to damage the photosynthetic apparatus. The damage is caused in part by reactive oxygen species (ROS) generated by electrons leaking from the photosynthetic electron transport system. To investigate the mechanisms equipped in higher plants to reduce high light (HL) stress, we surveyed the response of 7000 Arabidopsis genes to HL, taking advantage of the recently developed microarray technology. Our analysis revealed that 110 genes had a positive response to a 3 h treatment at a light intensity of 150 W m−2. In addition to the scavenging enzymes of ROS, the genes involved in biosynthesis of lignins and flavonoids are activated by HL and actually resulted in increased accumulation of lignins and anthocyanins. Comparing the HL-responsive genes with drought-inducible genes identified with the same microarray system revealed a dense overlap between HL- and drought-inducible genes. In addition, we have identified 10 genes that showed upregulation by HL, drought, cold and also salt stress. These genes include RD29A, ERD7, ERD10, KIN1, LEA14 and COR15a, most of which are thought to be involved in the protection of cellular components.

Advances in heart rate variability signal analysis: Joint position statement by the e-Cardiology ESC Working Group and the European Heart Rhythm Association co-endorsed by the Asia Pacific Heart Rhythm Society

Following the publication of the Task Force document on heart rate variability (HRV) in 1996, a number of articles have been published to describe new HRV methodologies and their application in different physiological and clinical studies. This document presents a critical review of the new methods. A particular attention has been paid to methodologies that have not been reported in the 1996 standardization document but have been more recently tested in sufficiently sized populations. The following methods were considered: Long-range correlation and fractal analysis; Short-term complexity; Entropy and regularity; and Nonlinear dynamical systems and chaotic behaviour. For each of these methods, technical aspects, clinical achievements, and suggestions for clinical application were reviewed. While the novel approaches have contributed in the technical understanding of the signal character of HRV, their success in developing new clinical tools, such as those for the identification of high-risk patients, has been rather limited. Available results obtained in selected populations of patients by specialized laboratories are nevertheless of interest but new prospective studies are needed. The investigation of new parameters, descriptive of the complex regulation mechanisms of heart rate, has to be encouraged because not all information in the HRV signal is captured by traditional methods. The new technologies thus could provide after proper validation, additional physiological, and clinical meaning. Multidisciplinary dialogue and specialized courses in the combination of clinical cardiology and complex signal processing methods seem warranted for further advances in studies of cardiac oscillations and in the understanding normal and abnormal cardiac control processes.

Identification of cis-acting promoter elements in cold- and dehydration-induced transcriptional pathways in Arabidopsis, rice, and soybean.

The genomes of three plants, Arabidopsis (Arabidopsis thaliana), rice (Oryza sativa), and soybean (Glycine max), have been sequenced, and their many genes and promoters have been predicted. In Arabidopsis, cis-acting promoter elements involved in cold- and dehydration-responsive gene expression have been extensively analysed; however, the characteristics of such cis-acting promoter sequences in cold- and dehydration-inducible genes of rice and soybean remain to be clarified. In this study, we performed microarray analyses using the three species, and compared characteristics of identified cold- and dehydration-inducible genes. Transcription profiles of the cold- and dehydration-responsive genes were similar among these three species, showing representative upregulated (dehydrin/LEA) and downregulated (photosynthesis-related) genes. All (46 = 4096) hexamer sequences in the promoters of the three species were investigated, revealing the frequency of conserved sequences in cold- and dehydration-inducible promoters. A core sequence of the abscisic acid-responsive element (ABRE) was the most conserved in dehydration-inducible promoters of all three species, suggesting that transcriptional regulation for dehydration-inducible genes is similar among these three species, with the ABRE-dependent transcriptional pathway. In contrast, for cold-inducible promoters, the conserved hexamer sequences were diversified among these three species, suggesting the existence of diverse transcriptional regulatory pathways for cold-inducible genes among the species.

Identification of plant promoter constituents by analysis of local distribution of short sequences

BackgroundPlant promoter architecture is important for understanding regulation and evolution of the promoters, but our current knowledge about plant promoter structure, especially with respect to the core promoter, is insufficient. Several promoter elements including TATA box, and several types of transcriptional regulatory elements have been found to show local distribution within promoters, and this feature has been successfully utilized for extraction of promoter constituents from human genome.ResultsLDSS (Local Distribution of Short Sequences) profiles of short sequences along the plant promoter have been analyzed in silico, and hundreds of hexamer and octamer sequences have been identified as having localized distributions within promoters of Arabidopsis thaliana and rice. Based on their localization patterns, the identified sequences could be classified into three groups, pyrimidine patch (Y Patch), TATA box, and REG (Regulatory Element Group). Sequences of the TATA box group are consistent with the ones reported in previous studies. The REG group includes more than 200 sequences, and half of them correspond to known cis-elements. The other REG subgroups, together with about a hundred uncategorized sequences, are suggested to be novel cis-regulatory elements. Comparison of LDSS-positive sequences between Arabidopsis and rice has revealed moderate conservation of elements and common promoter architecture. In addition, a dimer motif named the YR Rule (C/T A/G) has been identified at the transcription start site (-1/+1). This rule also fits both Arabidopsis and rice promoters.ConclusionLDSS was successfully applied to plant genomes and hundreds of putative promoter elements have been extracted as LDSS-positive octamers. Identified promoter architecture of monocot and dicot are well conserved, but there are moderate variations in the utilized sequences.

Role of a COP1 interactive protein in mediating light-regulated gene expression in arabidopsis.

Arabidopsis seedlings display distinct patterns of gene expression and morphogenesis according to the ambient light condition. An Arabidopsis nuclear protein, CONSTITUTIVE PHOTOMORPHOGENIC1 (COP1), acts to repress photomorphogenesis in the absence of light. The Arabidopsis CIP7 protein was identified by its capability to interact with COP1. CIP7 is a novel nuclear protein that contains transcriptional activation activity without a recognizable DNA binding motif. CIP7 requires light for its high level of expression, and COP1 seems to play a role in repressing its expression in darkness. Decreasing CIP7 expression by introducing antisense CIP7 RNA resulted in defects in light-dependent anthocyanin and chlorophyll accumulation. Antisense plants also displayed reduced expression of light-inducible genes for anthocyanin biosynthesis and photosynthesis. However, no defect was observed in light-dependent inhibition of hypocotyl elongation. Taken together, our data indicate that CIP7 acts as a positive regulator of light-regulated genes and is a potential direct downstream target of COP1 for mediating light control of gene expression.

Synchronization of spontaneous eyeblinks while viewing video stories

Blinks are generally suppressed during a task that requires visual attention and tend to occur immediately before or after the task when the timing of its onset and offset are explicitly given. During the viewing of video stories, blinks are expected to occur at explicit breaks such as scene changes. However, given that the scene length is unpredictable, there should also be appropriate timing for blinking within a scene to prevent temporal loss of critical visual information. Here, we show that spontaneous blinks were highly synchronized between and within subjects when they viewed the same short video stories, but were not explicitly tied to the scene breaks. Synchronized blinks occurred during scenes that required less attention such as at the conclusion of an action, during the absence of the main character, during a long shot and during repeated presentations of a similar scene. In contrast, blink synchronization was not observed when subjects viewed a background video or when they listened to a story read aloud. The results suggest that humans share a mechanism for controlling the timing of blinks that searches for an implicit timing that is appropriate to minimize the chance of losing critical information while viewing a stream of visual events.

Heterogeneity of Arabidopsis core promoters revealed by high-density TSS analysis

Our limited understanding of plant promoters does not allow us to recognize any core promoter elements for the majority of plant promoters. To understand the promoter architecture of Arabidopsis, we used the combined approach of in silico detection of novel core promoter elements and large-scale determination of transcription start sites (TSSs). To this end, we developed a novel methodology for TSS identification, using a combination of the cap-trapper and massively parallel signature sequencing methods. This technique, CT-MPSS, allowed us to identify 158 237 Arabidopsis TSS tags corresponding to 38 311 TSS loci, which provides an opportunity for quantitative analysis of plant promoters. The expression characteristics of these promoters were analyzed with respect to core promoter elements detected by our in silico analyses, revealing that Arabidopsis promoters contain two main types of elements with exclusive characteristics, the TATA type and the GA type. The TATA-type promoters tend to be associated with the Y Patch and the Inr motif, and cause high expression with sharp-peak TSS clusters. By contrast, the GA type produces broad-type TSS clusters. Unlike mammalian promoters, plant promoters are not associated with CpG islands. However, plant-specific GA-type promoters share some characteristics with mammalian CpG-type promoters.

ppdb: a plant promoter database

ppdb (http://www.ppdb.gene.nagoya-u.ac.jp) is a plant promoter database that provides promoter annotation of Arabidopsis and rice. The database contains information on promoter structures, transcription start sites (TSSs) that have been identified from full-length cDNA clones and also a vast amount of TSS tag data. In ppdb, the promoter structures are determined by sets of promoter elements identified by a position-sensitive extraction method called local distribution of short sequences (LDSS). By using this database, the core promoter structure, the presence of regulatory elements and the distribution of TSS clusters can be identified. Although no differentiation of promoter architecture among plant species has been reported, there is some divergence of utilized sequences for promoter elements. Therefore, ppdb is based on species-specific sets of promoter elements, rather than on general motifs for multiple species. Each regulatory sequence is hyperlinked to literary information, a PLACE entry served by a plant cis-element database, and a list of promoters containing the regulatory sequence.

INCREASED LEVEL OF POLYPLOIDY1, a Conserved Repressor of CYCLINA2 Transcription, Controls Endoreduplication in Arabidopsis

Endoreduplication is a type of cell cycle in which DNA replication continues without cell division. We have isolated several dominant mutants from Arabidopsis thaliana activation tagging lines by flow cytometry. One of the mutants, increased level of polyploidy1-1D (ilp1-1D), showed increased polyploidy in both light- and dark-grown hypocotyls. The corresponding gene of ilp1-1D encodes a protein homologous to the C-terminal region of mammalian GC binding factor. We demonstrate that this protein functions as a transcriptional repressor in vivo. The expression of all members of the CYCLINA2 (CYCA2) family was reduced in an ILP1 overexpressing line, and the mouse (Mus musculus) homolog of ILP1 repressed cyclin A2 expression in mouse NIH3T3 cells. T-DNA insertion mutants of ILP1 showed reduced polyploidy and upregulated all CYCA2 expression. Furthermore, loss of CYCA2;1 expression induces an increase in polyploidy in Arabidopsis. We demonstrate that this protein regulates endoreduplication through control of CYCA2 expression in Arabidopsis.