Biology subjects

Yamaguchi, K.

Publications and source records attributed to Yamaguchi, K..

69 records · Page 4Linked to original sources

bioRxiv2021

Conservation and diversity in transcriptional responses among host plants forming distinct arbuscular mycorrhizal morphotypes

O_LIThe morphotype of arbuscular mycorrhizal (AM) roots is distinct mostly depending on AM host species: Arum, Paris, and Intermediate types. We previously reported that gibberellin (GA) promotes the establishment of Paris-type AM symbiosis in Eustoma grandiflorum despite its negative effects on Arum-type AM symbiosis in model plants. However, the molecular mechanisms underlying the differential effects of GA on different morphotypes, including Intermediate-type AM symbiosis, remain elusive. C_LIO_LIComparative transcriptomics revealed that several symbiosis-related genes were transcriptionally promoted upon AM fungal colonization in Lotus japonicus (Arum-type), Daucus carota (Intermediate-type), and E. grandiflorum (Paris-type). Interestingly, upon GA treatment, the fungal colonization levels and expression of symbiosis-related genes were suppressed in L. japonicus and D. carota but were promoted in E. grandiflorum. C_LIO_LIExogenous GA transcriptionally inhibited the biosynthetic process of a host-derived signal molecule involved in AM symbiosis, strigolactone, in L. japonicus and E. grandiflorum. Additionally, disaccharides mainly metabolized in AM roots would be different between L. japonicus and D. carota/E. grandiflorum. C_LIO_LIThis study uncovered the conserved transcriptional responses during mycorrhization and diverse responses to GA in AM roots with distinct morphotypes among phylogenetically distant host plants. C_LI

plant biology↗

bioRxiv2021

A genome-wide knock-out screen for actors of epigenetic silencing reveals new regulators of germline genes and 2-cell like cell state

Epigenetic mechanisms are essential to establish and safeguard cellular identities in mammals. They dynamically regulate the expression of genes, transposable elements, and higher-order chromatin structures. Expectedly, these chromatin marks are indispensable for mammalian development and alterations often lead to diseases such as cancer. Molecularly, epigenetic mechanisms rely on factors to establish patterns, interpret them into a transcriptional output, and maintain them across cell divisions. A global picture of these phenomena has started to emerge over the years, yet many of the molecular actors remain to be discovered. In this context, we have developed a reporter system sensitive to epigenetic perturbations to report on repressive pathways based on Dazl, which is normally repressed in mouse ES cells. We used this system for a genome-wide CRISPR knock-out screen, which yielded expected hits (DNMT1, UHRF1, MGA), as well as novel candidates. We prioritized the candidates by secondary screens, and led further experiments on 6 of them: ZBTB14, KDM5C, SPOP, MCM3AP, BEND3, and KMT2D. Our results show that all 6 candidates regulate the expression of germline genes. In addition, we find that removal of ZBTB14, KDM5C, SPOP and MCM3AP led to similar transcriptional responses, including a reactivation of the 2-cell like cell (2CLC) signature. Therefore, our genetic screen has identified new regulators of key cellular states.

molecular biology↗

bioRxiv2021

Transcriptomic analysis of resistant and susceptible responses in a new model root-knot nematode infection system using Solanum torvum and Meloidogyne arenaria

Root-knot nematodes (RKNs) are among the most devastating pests in agriculture. Solanum torvum Sw. (turkey berry) has been used as a rootstock for eggplant (aubergine) cultivation because of its resistance to RKNs, including Meloidogyne incognita and M. arenaria. We previously found that a pathotype of M. arenaria, A2-J, is able to infect and propagate in S. torvum. In vitro infection assays showed that S. torvum induces the accumulation of brown pigments during avirulent pathotype A2-O infection, but not during virulent A2-J infection. This experimental system is advantageous because resistant and susceptible responses can be distinguished within a few days, and because a single plant genome can yield information about both resistant and susceptible responses. Comparative RNA-sequencing analysis of S. torvum inoculated with A2-J and A2-O at early stages of infection was used to parse the specific resistance and susceptible responses. Infection with A2-J did not induce statistically significant changes in gene expression within one day post-inoculation (DPI), but afterward, A2-J specifically induced the expression of chalcone synthase, spermidine synthase, and genes related to cell wall modification and transmembrane transport. Infection with A2-O rapidly induced the expression of genes encoding class III peroxidases, sesquiterpene synthases, and fatty acid desaturases at 1 DPI, followed by genes involved in defense, hormone signaling, and the biosynthesis of lignin at 3 DPI. Both isolates induced the expression of suberin biosynthetic genes, which may be triggered by wounding during nematode infection. Histochemical analysis revealed that A2-O, but not A2-J, induced lignin accumulation at the root tip, suggesting that physical reinforcement of cell walls with lignin is an important defense response against nematodes. The S. torvum-RKN system can provide a molecular basis for understanding plant-nematode interactions.

plant biology↗

bioRxiv2021

On the application of BERT models for nanopore methylation detection

MotivationDNA methylation is a common epigenetic modification, which is widely associated with various biological processes, such as gene expression, aging, and disease. Nanopore sequencing provides a promising methylation detection approach through monitoring abnormal signal shifts for detecting modified bases in target motif regions. Recently, model-based approaches, especially those with deep learning models, have achieved significant performance improvements on nanopore methylation detection. In this work, we explore using bidirectional encoder representations from transformers (BERT) for doing the task, which can provide non-recurrent neural structures for fast parallel computation. ResultsWe find original BERT architecture does not work as well as the bidirectional recurrent neural network (biRNN) on the nanopore methylation prediction task. Through further analysis, we observe recurrent patterns of positional-signal-shift in the context window surrounding target 5-methylcytosine (5mC) and N6-methyladenine (6mA) motifs. We propose a refined BERT with relative position representation and center hidden units concatenation, which takes account of task-specific characters into modeling. We perform systematic evaluations in-sample and cross-sample. The experiment results show that the refined BERT model can achieve competitive or even better results than the state-of-the-art biRNN model, while the model inference speed is about 6x faster. Besides, on the cross-sample evaluation of datasets from the different research groups, BERT models demonstrate a good generalization performance. AvailabilityThe source code and data are available at https://github.com/yaozhong/methBERT Contactyaozhong@ims.u-tokyo.ac.jp

bioinformatics↗

bioRxiv2020

Inference of a genome-wide protein-coding gene set of the inshore hagfish Eptatretus burgeri

The group of hagfishes (Myxiniformes) arose from agnathan (jawless vertebrate) lineages and is one of the only two extant cyclostome taxa, together with lampreys (Petromyzontiformes). Even though whole genome sequencing has been achieved for diverse vertebrate taxa, genome-wide sequence information has been highly limited for cyclostomes. Here we sequenced the genome of the inshore hagfish Eptatretus burgeri using DNA extracted from the testis, with a short-read sequencing platform, aiming at reconstructing a high-coverage coding gene catalogue. The obtained genome assembly, scaffolded with mate-pair reads and paired RNA-seq reads, exhibited an N50 scaffold length of 293 Kbp, which allowed the genome-wide prediction of coding genes. This computation resulted in the gene models whose completeness was estimated at the complete coverage of more than 83 % and the partial coverage of more than 93 % by referring to evolutionarily conserved single-copy orthologs. The high contiguity of the assembly and completeness of resulting gene models promises a high utility in various comparative analyses including phylogenomics and phylome exploration.

genomics↗

bioRxiv2020

Development of diagnostic PCR and LAMP markers for MALE STERILITY 1 (MS1) in Cryptomeria japonica D. Don

ObjectiveDue to the allergic nature of the pollen of Cryptomeria japonica, the most important Japanese forestry conifer, a pollen-free cultivar is preferred. Mutant trees detected in nature have been used for the production of a pollen-free cultivar. In order to reduce the time and cost needed for the production and breeding, we aimed to develop simple diagnostic molecular markers for mutant alleles of the causative gene MALE STERILITY 1 (MS1) in C. japonica. The expected function of this gene, its two dysfunctional mutations, and genetic diversity were described recently in a related study. ResultsWe have developed PCR and LAMP markers to detect mutant alleles and to present experimental options depending on available laboratory equipment. At field stations, where PCR machines are not available, LAMP markers were developed. LAMP only needs heat-blocks or a water bath to perform the isothermal amplification and assay results can be easily seen by eye. Because the causative mutations were deletions, two kinds of PCR markers, amplified length polymorphism (ALP) and allele specific PCR (ASP) markers, were developed. These assays can be carried out by capillary or agarose gel electrophoresis.

genetics↗

bioRxiv2020

Conservation and host-specific expression of non-tandemly repeated heterogenous ribosome RNA gene in arbuscular mycorrhizal fungi

The ribosomal RNA-encoding gene (rDNA) has a characteristic genomic nature: tens to thousands of copies in a genome, tandemly repeated structure, and intragenomic sequence homogeneity. These features contribute to ribosome productivity via physiological and evolutionary processes. We reported previously the exceptional absence of these features in the model arbuscular mycorrhizal (AM) fungus Rhizophagus irregularis. Here we examine the phylogenetic distribution of the exceptional rDNA features in the genus Rhizophagus via improving the genome sequence of R. clarus. Cross-species comparison indicated similarity of their rDNAs not only in the genomic features but also in the distribution of intragenomic polymorphic sites on the paralogs. Ribosomal RNA comprises multiple domains with different functions. The two Rhizophagus species commonly exhibited a variation enrichment site, ES27L, which is related to translational fidelity and antibiotic sensitivity. Variation enrichment on ES27L has not been observed in other organisms lacking the three rDNA features such as malaria parasites and Cyanidioschyzon merolae. Expression profiling of rDNAs in R. irregularis revealed that rDNA paralogs are expressed differently in association with host plant species. Our results suggest a broad distribution of the disarranged rDNA across AM fungi and its involvement in the successful association with the broad range of host species.

genomics↗

bioRxiv2020

Identification and genetic diversity analysis of a male-sterile gene (MS1) in Japanese cedar (Cryptomeria japonica D. Don)

Identifying causative genes for a target trait in conifer reproduction is challenging for species lacking whole-genome sequences. In this study, we searched for the male-sterility gene (MS1) in Cryptomeria japonica, aiming to promote marker-assisted selection (MAS) of male-sterile C. japonica to reduce the pollinosis caused by pollen dispersal from artificial C. japonica forests in Japan. We searched for mRNA sequences expressed in male strobili and found the gene CJt020762, coding for a lipid transfer protein containing a 4-bp deletion specific to male-sterile individuals. We also found a 30-bp deletion by sequencing the entire gene of another individual with the ms1. All nine breeding materials with the allele ms1 had either a 4-bp or 30-bp deletion in gene CJt020762, both of which are expected to result in faulty gene transcription and function. Furthermore, the 30-bp deletion was detected from three of five individuals in the Ishinomaki natural forest. From our findings, CJt020762 was considered to be the causative gene of MS1. Thus, by performing MAS using two deletion mutations as a DNA marker, it will be possible to find novel breeding materials of C. japonica with the allele ms1 adapted to the unique environment of each region of the Japanese archipelago.

genetics↗

bioRxiv2020

The anti-tubercular activity of simvastatin is mediated by cholesterol-dependent regulation of autophagy via the AMPK-mTORC1-TFEB axis

Statins, which inhibit both cholesterol biosynthesis and protein prenylation branches of the mevalonate pathway, increase anti-tubercular antibiotic efficacy in animal models. We investigated the mechanism of anti-tubercular action of simvastatin in Mycobacterium tuberculosis-infected human monocytic cells. We found that the anti-tubercular activity of statins was phenocopied by cholesterol-branch but not prenylation-branch inhibitors. Moreover, statin treatment blocked activation of mechanistic target of rapamycin complex 1 (mTORC1), activated AMP-activated protein kinase (AMPK) through increased intracellular AMP:ATP ratios, and favored nuclear translocation of transcription factor EB (TFEB). These mechanisms all induce autophagy, which is anti-mycobacterial. The biological effects of simvastatin on the AMPK-mTORC1-TFEB-autophagy axis were reversed by adding exogenous cholesterol to the cells. Overall, our data demonstrate that the anti-tubercular activity of simvastatin requires inhibiting cholesterol biosynthesis, reveal novel links between cholesterol homeostasis, AMPK-mTORC1-TFEB axis, and intracellular infection control, and uncover new anti-tubercular therapy targets.

immunology↗

bioRxiv2020

Switching secretory pathway direction for organelle acquisition in plants

Eukaryotic cells acquired novel organelles during evolution through mechanisms that remain largely obscure. The existence of the unique oil body compartment is a synapomorphy of liverworts that represents lineage-specific acquisition of this organelle during evolution, although its origin, biogenesis, and physiological function are yet unknown. We found that two Syntaxin 1 paralogs in the liverwort, Marchantia polymorpha, are distinctly targeted to forming cell plates and the oil body, suggesting these structures share some developmental similarity. Oil body formation is under the regulation of an ERF/AP2-type transcription factor and loss of the oil body increased M. polymorpha herbivory. These findings highlight a common strategy for the acquisition of organelles with distinct functions in plants, via periodical switching in secretion direction depending on cellular phase transition.

plant biology↗

bioRxiv2019

The minimal behavioral time window for reward conditioning in the nucleus accumbens of mice

The temporal precision of reward-reinforcement learning is determined by the minimal time window of the reward action--theoretically known as the eligibility trace. In animal studies, however, such a minimal time window and its origin have not been well understood. Here, we used head-restrained mice to accurately control the timing of sucrose water as an unconditioned stimulus (US); we found that the reinforcement effect of the US occurred only within 1 s after a short tone of a conditioned stimulus (CS). The conditioning required the dopamine D1 receptor and CaMKII signaling in the nucleus accumbens (NAc). The time window was not reduced by replacing CS with optogenetic stimulation of the synaptic inputs to the NAc, which is in agreement with previous reports on the effective dopamine timing of NAc synapses. Thus, our data suggest that the minimal reward time window is 1 s, and is formed in the NAc.

neuroscience↗

bioRxiv2018

A comprehensive reference transcriptome resource for the Iberian ribbed newt Pleurodeles waltl, an emerging model for developmental and regeneration biology

Urodele amphibian newts have unique biological properties, notably including prominent regeneration ability. Iberian ribbed newt, Pleurodeles waltl, is a promising model newt along with the successful development of the easy breeding system and efficient transgenic and genome editing methods. However, genetic information of P. waltl was limited. In the present study, we conducted an intensive transcriptome analysis of P. waltl using RNA-sequencing to build gene models and annotate them. We generated 1.2 billion Illumina reads from a wide variety of samples across 11 different tissues and 9 time points during embryogenesis. They were assembled into 202,788 non-redundant contigs that appear to cover nearly complete (~98%) P. waltl protein-coding genes. Using the gene set as a reference, our gene network analysis identified regeneration-, developmental-stage-, and tissue-specific co-expressed gene modules. Ortholog analyses with other vertebrates revealed the gene repertoire evolution of amphibians which includes urodele-specific loss of bmp4 and duplications of wnt11b. Our transcriptome resource will enhance future research employing this emerging model animal for regeneration research as well as other areas such as developmental biology, stem cell biology, cancer research, ethology and toxico-genomics. These data are available via our portal website, iNewt (http://www.nibb.ac.jp/imori/main/).

genomics↗

bioRxiv2018

The RopGEF KARAPPO is Essential for the Initiation of Vegetative Reproduction in Marchantia

Many plants can reproduce vegetatively, producing clonal progeny from vegetative cells; however, little is known about the molecular mechanisms underlying this process. Liverwort (Marchantia polymorpha), a basal land plant, propagates asexually via gemmae, which are clonal plantlets formed in gemma cups on the dorsal side of the vegetative thallus [1]. The initial stage of gemma development involves elongation and asymmetric divisions of a specific type of epidermal cell, called a gemma initial, which forms on the floor of the gemma cup [2, 3]. To investigate the regulatory mechanism underlying gemma development, we focused on two allelic mutants in which no gemma initial formed; these mutants were named karappo, meaning \"empty\". We used whole-genome sequencing of both mutants, and molecular genetic analyses to identify the causal gene, KARAPPO (KAR), which encodes a Rop guanine nucleotide exchange factor (RopGEF) carrying a PRONE catalytic domain. In vitro GEF assays showed that the full-length KAR protein and the PRONE domain have significant GEF activity toward MpRop, the only Rop GTPase in M. polymorpha. Moreover, genetic complementation experiments showed a significant role for the N- and C-terminal variable regions in gemma development. Our investigation demonstrated an essential role for KAR/RopGEF in the initiation of plantlet development from a differentiated cell, which may involve cell polarity formation and subsequent asymmetric cell division via activation of Rop signaling, implying a similar developmental mechanism in vegetative reproduction of various land plants.

plant biology↗

bioRxiv2018

Repeated inversions at the pannier intron drive diversification of intraspecific colour patterns of ladybird beetles

How genetic information is modified to generate phenotypic variation within a species is one of the central questions in evolutionary biology. Here we focus on the striking intraspecific diversity of more than 200 aposematic elytral (forewing) colour patterns of the multicoloured Asian ladybird beetle, Harmonia axyridis, which is regulated by a tightly linked genetic locus h. Our loss-of-function analyses, genetic association studies, de novo genome assemblies, and gene expression data reveal that the GATA transcription factor gene pannier is the major regulatory gene located at the h locus, and suggest that repeated inversions and cis-regulatory modifications at pannier led to the expansion of colour pattern variation in H. axyridis. Moreover, we show that the colour patterning function of pannier is conserved in the seven spotted ladybird beetle, Coccinella septempunctata, suggesting that H. axyridis extraordinary intra-specific variation may have arisen from ancient modifications in a conserved elytral colour patterning mechanisms in ladybird beetles.

evolutionary biology↗

bioRxiv2017

Evidence of non-tandemly repeated rDNAs and their intragenomic heterogeneity in Rhizophagus irregularis

Arbuscular mycorrhizal fungus (AMF) species are one of the most widespread symbionts of land plants. Our substantially improved reference genome assembly of a model AMF, Rhizophagus irregularis DAOM-181602 (total contigs = 210), facilitated discovery of repetitive elements with unusual characteristics. R. irregularis has only ten or eleven copies of complete 45S rDNAs, whereas the general eukaryotic genome has tens to thousands of rDNA copies. R. irregularis rDNAs are highly heterogeneous and lack a tandem repeat structure. These findings provide evidence for the hypothesis that rDNA heterogeneity depends on the lack of tandem repeat structures. RNA-Seq analysis confirmed that all rDNA variants are actively transcribed. Observed rDNA/rRNA polymorphisms may modulate translation by using different ribosomes depending on biotic and abiotic interactions. The non-tandem repeat structure and intragenomic heterogeneity of AMF rDNA/rRNA may facilitate adaptation to a various environmental condition including the broad host range.

genomics↗