担当区分：筆頭著者,　責任著者   記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Oxford University Press (OUP)  

Abstract

In vascular plants, roots anchor themselves into the soil and take up water and nutrients to provide them to the shoots. Therefore, continuous growth and development of the roots are important for plant life. To achieve this, photosynthesizing leaves must be able to supply sufficient photoassimilates to the roots. However, the mechanisms by which plants maintain carbon levels in roots remain elusive. Here, we focused on the Arabidopsis (Arabidopsis thaliana) CLAVATA3/ESR-related 2 (CLE2) peptide, which was detected in Arabidopsis xylem exudate, and its homologs. CLE2 and CLE3 genes responded to carbon-deficient conditions. Loss- and gain-of-function mutant analyses showed that CLE genes positively affected root sucrose level. Mutations in the CLE genes resulted in a high shoot/root ratio under sucrose-free conditions. Grafting experiments demonstrated the systemic effect of CLE peptide genes. These findings provide insights into the molecular basis for the relationship between roots and leaves in maintenance of the root sucrose levels and growth.

DOI： 10.1093/plphys/kiac227

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担当区分：筆頭著者,　責任著者   記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：MDPI AG  

The xylem is the main pathway for the transport of water and molecules from roots to shoots. To date, it has been reported that secreted oligopeptides mediate root-to-shoot signaling, and some long-distance mobile oligopeptides have been detected in xylem exudates. However, the conservation of a number of oligopeptides and the overall features of peptide fragments contained in xylem exudates are poorly understood. Here, we conducted a comprehensive analysis of small proteins and peptides in tomato (Solanum lycopersicum) xylem exudates and characterized the identified peptide fragments. We found that putative secreted proteins were enriched in xylem exudates compared with all proteins in the tomato protein database. We identified seven oligopeptides that showed common features of bioactive oligopeptides, including homologs of CLV3/ESR-related (CLE), C-TERMINALLY ENCODED PEPTIDE (CEP), and CASPARIAN STRIP INTEGRITY FACTOR (CIF) peptides. Furthermore, five of the identified oligopeptides were homologs of the soybean xylem exudate-associated oligopeptides that we previously reported. Our results suggest that oligopeptides in xylem exudates are conserved across plant species and provide insights into not only root-to-shoot signaling but also the maintenance of the xylem conduit.

DOI： 10.3390/life12040592

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Oxford University Press (OUP)  

Abstract

Oryza longistaminata, a wild rice, vegetatively reproduces and forms a networked clonal colony consisting of ramets connected by rhizomes. Although water, nutrients, and other molecules can be transferred between ramets via the rhizomes, inter-ramet communication in response to spatially heterogeneous nitrogen availability is not well understood. We studied the response of ramet pairs to heterogeneous nitrogen availability using a split hydroponic system that allowed each ramet root to be exposed to different conditions. Ammonium uptake was compensatively enhanced in the sufficient-side root when roots of the ramet pairs were exposed to ammonium-sufficient and ammonium-deficient conditions. Comparative transcriptome analysis revealed that a gene regulatory network for effective ammonium assimilation and amino acid biosynthesis was activated in the sufficient-side roots. Allocation of absorbed nitrogen from the nitrogen-sufficient to the nitrogen-deficient ramets was rather limited. Nitrogen was preferentially used for newly growing axillary buds on the sufficient-side ramets. Biosynthesis of trans-zeatin (tZ), a cytokinin, was upregulated in response to the nitrogen supply, but tZ appeared not to target the compensatory regulation. Our results also implied that the O. longistaminata putative ortholog of rice (Oryza sativa) C-terminally encoded peptide1 plays a role as a nitrogen-deficient signal in inter-ramet communication, providing compensatory upregulation of nitrogen assimilatory genes. These results provide insights into the molecular basis for efficient growth strategies of asexually proliferating plants growing in areas where the distribution of ammonium ions is spatially heterogeneous.

DOI： 10.1093/plphys/kiac025

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その他リンク： https://academic.oup.com/plphys/article-pdf/188/4/2364/43109834/kiac025.pdf

記述言語：英語   掲載種別：研究論文（学術雑誌）  

Pollen-free varieties are advantageous in promoting cut-flower production. In this study, we identified a candidate mutation which is responsible for pollen sterility in a strain of Lilium × formolongi, which was originally identified as a naturally occurred male-sterile plant in a seedling population. The pollen sterility occurred due to the degradation of pollen mother cells (PMCs) before meiotic cell division. Genetic analysis suggested that the male-sterile phenotype is attributed to one recessive locus. Transcriptome comparison between anthers of sterile and fertile plants in a segregated population identified a transcript that was expressed only in pollen-fertile plants, which is homologous to TDF1 (DEFECTIVE in TAPETAL DEVELOPMENT and FUNCTION1) in Arabidopsis, a gene encoding a transcription factor AtMYB35 that is known as a key regulator of pollen development. Since tdf1 mutant shows male sterility, we assumed that the absence transcript of the TDF1-like gene, named as LflTDF1, is the reason for pollen sterility observed in the mutant. A 30 kbp-long nanopore sequence read containing LflTDF1 was obtained from a pollen-fertile accession. PCR analyses using primers designed from the sequence suggested that at least a 30kbp-long region containing LflTDF1 was deleted or replaced by unknown sequence in the pollen-sterile mutant. Since the cross between L. × formolongi and Easter lily (L. longiflorum) is compatible, we successfully introgressed the male-sterile allele, designated as lfltdf1, to Easter lily. To our knowledge, this is the first report of molecular identification of a pollen-sterile candidate gene in lily. The identification and marker development of LflTDF1 gene will assist pollen-free lily breeding of Easter lilies and other lilies.

DOI： 10.3389/fpls.2022.914671

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Springer Science and Business Media {LLC}  

<jats:title>Abstract</jats:title>
<jats:p>Legumes utilize a shoot-mediated signaling system to maintain a mutualistic relationship with nitrogen-fixing bacteria in root nodules. In <jats:italic>Lotus japonicus</jats:italic>, shoot-to-root transfer of microRNA miR2111 that targets <jats:italic>TOO MUCH LOVE</jats:italic>, a nodulation suppressor in roots, has been proposed to explain the mechanism underlying nodulation control from shoots. However, the role of shoot-accumulating miR2111s for the systemic regulation of nodulation was not clearly shown. Here, we find <jats:italic>L. japonicus</jats:italic> has seven miR2111 loci, including those mapped through RNA-seq. <jats:italic>MIR2111-5</jats:italic> expression in leaves is the highest among miR2111 loci and repressed after rhizobial infection depending on a shoot-acting HYPERNODULATION ABERRANT ROOT FORMATION1 (HAR1) receptor. <jats:italic>MIR2111-5</jats:italic> knockout mutants show significantly decreased nodule numbers and miR2111 levels. Furthermore, grafting experiments using transformants demonstrate scions with altered miR2111 levels influence nodule numbers in rootstocks in a dose-dependent manner. Therefore, miR2111 accumulation in leaves through <jats:italic>MIR2111-5</jats:italic> expression is required for HAR1-dependent systemic optimization of nodule number.</jats:p>

DOI： 10.1038/s41467-020-19037-9

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Scientific Societies  

<jats:p> Legumes survive in nitrogen-limited soil by forming a symbiosis with rhizobial bacteria. During root nodule symbiosis, legumes strictly control the development of their symbiotic organs, the nodules, in a process known as autoregulation of nodulation (AON). The study of hypernodulation mutants has elucidated the molecular basis of AON. Some hypernodulation mutants show an increase in rhizobial infection in addition to developmental alteration. However, the relationship between the AON and the regulation of rhizobial infection has not been clarified. We previously isolated daphne, a nodule inception (nin) allelic mutant, in Lotus japonicus. This mutant displayed dramatically increased rhizobial infection, suggesting the existence of NIN-mediated negative regulation of rhizobial infection. Here, we investigated whether the previously isolated components of AON, especially CLAVATA3/ESR (CLE)-RELATED-ROOT SIGNAL1 (CLE-RS1), CLE-RS2, and their putative receptor HYPERNODULATION AND ABERRANT ROOT FORMATION1 (HAR1), were able to suppress increased infection in the daphne mutant. The constitutive expression of LjCLE-RS1/2 strongly reduced the infection in the daphne mutant in a HAR1-dependent manner. Moreover, reciprocal grafting analysis showed that strong reduction of infection in daphne rootstock constitutively expressing LjCLE-RS1 was canceled by a scion of the har1 or klavier mutant, the genes responsible for encoding putative LjCLE-RS1 receptors. These data indicate that rhizobial infection is also systemically regulated by CLE-HAR1 signaling, a component of AON. In addition, the constitutive expression of NIN in daphne har1 double-mutant roots only partially reduced the rhizobial infection. Our findings indicate that the previously identified NIN-mediated negative regulation of infection involves unknown local signaling, as well as CLE-HAR1 long-distance signaling. </jats:p>

DOI： 10.1094/mpmi-08-19-0223-r

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掲載種別：研究論文（学術雑誌）   出版者・発行元：Oxford University Press (OUP)  

DOI： 10.1093/jxb/ery364

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：CURRENT BIOLOGY LTD  

Nitrogen is an indispensable inorganic nutrient that is required by plants throughout their life. Root nodule symbiosis (RNS) is an important strategy mainly adopted by legumes to enhance nitrogen acquisition, where several key processes required for the establishment of the symbiosis, are pleiotropically controlled by nitrate availability in soil. Although the autoregulation of nodulation (AON), a systemic long-range signaling, has been suggested to be implicated in nitrate-induced control of RNS, AON alone is insufficient to fully explain the pleiotropic regulation that is induced by nitrate. A recent elucidation of the function of a NIN-LIKE PROTEIN transcription factor has provided greater insights into the genetic mechanisms underlying nitrate-induced control of RNS in varying nitrate environments.

DOI： 10.1016/j.pbi.2018.04.006

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担当区分：筆頭著者   記述言語：英語   出版者・発行元：CURRENT BIOLOGY LTD  

Organ-to-organ communication is indispensable for higher organisms to maintain homeostasis over their entire life. Recent findings have uncovered that plants, like animals, mediate organ-to-organ communication by long-distance signaling through the vascular system. In particular, xylem-mobile secreted peptides have attracted much attention as root-to shoot long-distance signaling molecules in response to fluctuating environmental nutrient status. Several leguminous CLE peptides induced by rhizobial inoculation act as 'satiety' signals in long-distance negative feedback of nodule formation. By contrast, Arabidopsis CEP family peptides induced by local nitrogen (N)-starvation behave as systemic 'hunger' signals to promote compensatory N acquisition in other parts of the roots. Xylem sap peptidomics also implies the presence of still uncharacterized long-distance signaling peptides. This review highlights the current understanding of and new insights into the mechanisms and functions of root-to-shoot long-distance peptide signaling during environmental responses.

DOI： 10.1016/j.pbi.2016.07.009

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担当区分：筆頭著者,　責任著者   記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：WILEY  

There is a growing awareness that secreted peptides mediate organ-to-organ communication in higher plants. Xylem sap peptidomics is an effective but challenging approach for identifying long-distance mobile peptides. In this study we developed a simple, gel-free purification system that combines o-chlorophenol extraction with HPLC separation. Using this system, we successfully identified seven oligopeptides from soybean xylem sap exudate that had one or more post-transcriptional modifications: glycosylation, sulfation and/or hydroxylation. RNA sequencing and quantitative PCR analyses showed that the peptide-encoding genes are expressed in multiple tissues. We further analyzed the long-distance translocation of four of the seven peptides using gene-encoding peptides with single amino acid substitutions, and identified these four peptides as potential root-to-shoot mobile oligopeptides. Promoter-GUS analysis showed that all four peptide-encoding genes were expressed in the inner tissues of the root endodermis. Moreover, we found that some of these peptide-encoding genes responded to biotic and/or abiotic factors. These results indicate that our purification system provides a comprehensive approach for effectively identifying endogenous small peptides and reinforce the concept that higher plants employ various peptides in root-to-shoot signaling.

DOI： 10.1111/tpj.13015

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担当区分：筆頭著者,　責任著者   記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：TAYLOR & FRANCIS INC  

Nitrate is a major environmental factor in the inhibition of nodulation. In a model legume Lotus japonicus, a CLV1-like receptor kinase, HAR1, mediates nitrate inhibition and autoregulation of nodulation. Autoregulation of nodulation involves root-to-shoot-to-root long-distance communication, and HAR1 functions in shoots. However, it remains elusive where HAR1 functions in the nitrate inhibition of nodulation. We performed grafting experiments with the har1 mutant under various nitrate conditions, and found that shoot HAR1 is critical for the inhibition of nodulation at 10mM nitrate. Combined with our recent finding that the nitrate-induced CLE-RS2 glycopeptide binds directly to the HAR1 receptor, this result suggests that CLE-RS2/HAR1 long-distance signaling plays an important role in the both nitrate inhibition and the autoregulation of nodulation.

DOI： 10.1080/15592324.2014.1000138

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：FRONTIERS RESEARCH FOUNDATION  

Plant vascular systems are constructed by specific cell wall modifications through which cells are highly specialized to make conduits for water and nutrients. Xylem vessels are formed by thickened cell walls that remain after programmed cell death, and serve as water conduits from the root to the shoot. In contrast, phloem tissues consist of a complex of living cells, including sieve tube elements and their neighboring companion cells, and translocate photosynthetic assimilates from mature leaves to developing young tissues. Intensive studies on the content of vascular flow fluids have unveiled that plant vascular tissues transport various types of gene product, and the transport of some provides the molecular basis for the long-distance communications. Analysis of xylem sap has demonstrated the presence of proteins in the xylem transpiration stream. Recent studies have revealed that CLE and CEP peptides secreted in the roots are transported to above ground via the xylem in response to plant-microbe interaction and soil nitrogen starvation, respectively. Their leucine-rich repeat transmembrane receptors localized in the shoot phloem are required for relaying the signal from the shoot to the root. These findings well fit to the current scenario of root-to-shoot-to-root feedback signaling, where peptide transport achieves the root-to-shoot signaling, the first half of the signaling process. Meanwhile, it is now well-evidenced that proteins and a range of RNAs are transported via the phloem translocation system, and some of those can exert their physiological functions at their destinations, including roots. Thus, plant vascular systems may serve not only as conduits for the translocation of essential substances but also as long-distance communication pathways that allow plants to adapt to changes in internal and external environments at the whole plant level.

DOI： 10.3389/fpls.2015.00161

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担当区分：筆頭著者   記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：NATURE PUBLISHING GROUP  

Leguminous plants establish a symbiosis with rhizobia to enable nitrogen fixation in root nodules under the control of the presumed root-to-shoot-to-root negative feedback called autoregulation of nodulation. In Lotus japonicus, autoregulation is mediated by CLE-RS genes that are specifically expressed in the root, and the receptor kinase HAR1 that functions in the shoot. However, the mature functional structures of CLE-RS gene products and the molecular nature of CLE-RS/HAR1 signalling governed by these spatially distant components remain elusive. Here we show that CLE-RS2 is a post-translationally arabinosylated glycopeptide derived from the CLE domain. Chemically synthesized CLE-RS glycopeptides cause significant suppression of nodulation and directly bind to HAR1 in an arabinose-chain and sequence-dependent manner. In addition, CLE-RS2 glycopeptide specifically produced in the root is found in xylem sap collected from the shoot. We propose that CLE-RS glycopeptides are the long sought mobile signals responsible for the initial step of autoregulation of nodulation.

DOI： 10.1038/ncomms3191

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担当区分：筆頭著者   記述言語：英語  

DOI： 10.21769/BioProtoc.795

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

The interaction of legumes with N2-fixing bacteria collectively called rhizobia results in root nodule development. The number of nodules formed is tightly restricted through the systemic negative feedback control by the host called autoregulation of nodulation (AON). Here, we report the characterization and gene identification of TOO MUCH LOVE (TML), a root factor that acts during AON in a model legume Lotus japonicus. In our genetic analyses using another root-regulated hypernodulation mutant, plenty, the tml-1 plenty double mutant showed additive effects on the nodule number, whereas the tml-1 har1-7 double mutant did not, suggesting that TML and PLENTY act in different genetic pathways and that TML and HAR1 act in the same genetic pathway. The systemic suppression of nodule formation by CLE-RS1/RS2 overexpression was not observed in the tml mutant background, indicating that TML acts downstream of CLE-RS1/RS2. The tml-1 Snf2 double mutant developed an excessive number of spontaneous nodules, indicating that TML inhibits nodule organogenesis. Together with the determination of the deleted regions in tml-1/-2/-3, the fine mapping of tml-4 and the next-generation sequencing analysis, we identified a nonsense mutation in the Kelch repeat-containing F-box protein. As the gene knockdown of the candidate drastically increased the number of nodules, we concluded that it should be the causative gene. An expression analysis revealed that TML is a root-specific gene. In addition, the activity of ProTML-GUS was constitutively detected in the root tip and in the nodules/nodule primordia upon rhizobial infection. In conclusion, TML is a root factor acting at the final stage of AON. © 2013 The Author 2013. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

DOI： 10.1093/pcp/pct022

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担当区分：筆頭著者,　責任著者   記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：OXFORD UNIV PRESS  

Plant aerial parts are differentiated from stem cells that are located in the shoot apical meristem (SAM). CLAVATA3 (CLV3)-CLV1 is a well-known ligand-receptor pair, which functions in SAM maintenance. In Lotus japonicus, HYPERNODULATION ABERRANT ROOT FORMATION1 (HAR1) shows the highest similarity with CLV1 of all Arabidopsis receptor-like kinases (RLKs). However, HAR1 functions in the systemic regulation of root nodule development, but does not appear to function in SAM maintenance. Therefore, the gene that is responsible for SAM maintenance in L. japonicus is largely unknown. Here, we identified the L. japonicus CLV3-like (LjCLV3) gene as a counterpart of AtCLV3 and performed expression and functional analysis. LjCLV3 transcripts were detected in the central region of the shoot meristems. However, unlike AtCLV3, LjCLV3 expression was not detected in the epidermal layer, but in the inner layers of the shoot meristems. RNA interference (RNAi) of LjCLV3 caused enlargement of not only the SAM, but also the primary and secondary inflorescence meristems (IMs). Furthermore, LjCLV3-silenced plants exhibited fasciated stems and an increased number of flowers per peduncle. These results reveal that LjCLV3 is responsible for the maintenance of the SAM as well as the primary and secondary IMs.

DOI： 10.1093/pcp/pcr071

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：WILEY-BLACKWELL  

The number of root nodules developing on legume roots after rhizobial infection is controlled by the plant shoot through autoregulation and mutational inactivation of this mechanism leads to hypernodulation. We have characterised the Pisum sativum (pea) Sym28 locus involved in autoregulation and shown that it encodes a protein similar to the Arabidopsis CLAVATA2 (CLV2) protein. Inactivation of the PsClv2 gene in four independent sym28 mutant alleles, carrying premature stop codons, results in hypernodulation of the root and changes to the shoot architecture. In the reproductive phase sym28 shoots develops additional flowers, the stem fasciates, and the normal phyllotaxis is perturbed. Mutational substitution of an amino acid in one leucine rich repeat of the corresponding Lotus japonicus LjCLV2 protein results in increased nodulation. Similarly, down-regulation of the Lotus Clv2 gene by RNAi mediated reduction of the transcript level also resulted in increased nodulation. Gene expression analysis of LjClv2 and Lotus hypernodulation aberrant root formation Har1 (previously shown to regulate nodule numbers) indicated they have overlapping organ expression patterns. However, we were unable to demonstrate a direct protein-protein interaction between LjCLV2 and LjHAR1 proteins in contrast to the situation between equivalent proteins in Arabidopsis. LjHAR1 was localised to the plasma membrane using a YFP fusion whereas LjCLV2-YFP localised to the endoplasmic reticulum when transiently expressed in Nicotiana benthamiana leaves. This finding is the most likely explanation for the lack of interaction between these two proteins.

DOI： 10.1111/j.1365-313X.2010.04474.x

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担当区分：筆頭著者   記述言語：英語   掲載種別：研究論文（学術雑誌）  

Host legumes control root nodule numbers by sensing external and internal cues. A major external cue is soil nitrate, whereas a feedback regulatory system in which earlier formed nodules suppress further nodulation through shoot-root communication is an important internal cue. The latter is known as autoregulation of nodulation (AUT), and is believed to consist of two long-distance signals: a root-derived signal that is generated in infected roots and transmitted to the shoot; and a shoot-derived signal that systemically inhibits nodulation. In Lotus japonicus, the leucine-rich repeat receptor-like kinase, HYPERNODULATION ABERRANT ROOT FORMATION 1 (HAR1), mediates AUT and nitrate inhibition of nodulation, and is hypothesized to recognize the root-derived signal. Here we identify L. japonicus CLE-Root Signal 1 (LjCLE-RS1) and LjCLE-RS2 as strong candidates for the root-derived signal. A hairy root transformation study shows that overexpressing LjCLE-RS1 and -RS2 inhibits nodulation systemically and, furthermore, that the systemic suppression depends on HAR1. Moreover, LjCLE-RS2 expression is strongly up-regulated in roots by nitrate addition. Based on these findings, we propose a simple model for AUT and nitrate inhibition of nodulation mediated by LjCLE-RS1, -RS2 peptides and the HAR1 receptor-like kinase.

DOI： 10.1093/pcp/pcn194

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：OXFORD UNIV PRESS  

Nitrogen fixation in nodules provides leguminous plants with an ability to grow in nitrogen-starved soil. Infection of the host plants by microsymbionts triggers various physiological and morphological changes during nodule formation. In Lotus japonicus, expression of early nodulin (ENOD) genes is triggered by perception of bacterial signal molecules, nodulation factors (Nod factors). We examined the expression patterns of ENOD40 genes during the nodule formation process. Two ENOD40 genes of L. japonicus were specifically expressed in the nodule formation process, but they showed different expression patterns upon infection. Each ENOD40 gene demonstrates an individual specificity and regulation with regard to rhizobial infection.

DOI： 10.1093/pcp/pci138

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記述言語：英語   掲載種別：研究発表ペーパー・要旨（国際会議）   出版者・発行元：OXFORD UNIV PRESS  

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開催年月日： 2019年3月

記述言語：日本語   会議種別：ポスター発表  

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開催年月日： 2017年3月

記述言語：日本語   会議種別：ポスター発表  

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開催年月日： 2016年11月 - 2016年12月

記述言語：英語   会議種別：ポスター発表  

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開催年月日： 2016年3月

記述言語：日本語   会議種別：ポスター発表  

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開催年月日： 2015年10月

記述言語：日本語   会議種別：ポスター発表  

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開催年月日： 2015年9月

記述言語：英語   会議種別：口頭発表（一般）  

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