[Show abstract][Hide abstract] ABSTRACT: The cladoceran crustacean Daphnia pulex produces female offspring by parthenogenesis under favorable conditions, but in response to various unfavorable external stimuli, it produces male offspring (environmental sex determination: ESD). We recently established an innovative system for ESD studies using D. pulex WTN6 strain, in which the sex of the offspring can be controlled simply by changes in the photoperiod: the long-day and short-day conditions can induce female and male offspring, respectively. Taking advantage of this system, we demonstrated that de novo methyl farnesoate (MF) synthesis is necessary for male offspring production. These results indicate the key role of innate MF signaling as a conductor between external environmental stimuli and the endogenous male developmental pathway. Despite these findings, the molecular mechanisms underlying up- and downstream signaling of MF have not yet been well elucidated in D. pulex.
To elucidate up- and downstream events of MF signaling during sex determination processes, we compared the transcriptomes of daphnids reared under the long-day (female) condition with short-day (male) and MF-treated (male) conditions. We found that genes involved in ionotropic glutamate receptors, known to mediate the vast majority of excitatory neurotransmitting processes in various organisms, were significantly activated in daphnids by the short-day condition but not by MF treatment. Administration of specific agonists and antagonists, especially for the N-methyl-D-aspartic acid (NMDA) receptor, strongly increased or decreased, respectively, the proportion of male-producing mothers. Moreover, we also identified genes responsible for male production (e.g., protein kinase C pathway-related genes). Such genes were generally shared between the short-day reared and MF-treated daphnids.
We identified several candidate genes regulating ESD which strongly suggests that these genes may be essential factors for male offspring production as an upstream regulator of MF signaling in D. pulex. This study provides new insight into the fundamental mechanisms underlying how living organisms alter their phenotypes in response to various external environments.
[Show abstract][Hide abstract] ABSTRACT: Calcium (Ca) deficiency symptoms in plants often occur in agriculture; however, little is known about the mechanisms for adaptation to low-Ca conditions. To understand the mechanisms, we screened for Arabidopsis thaliana (L.) Heynh mutants sensitive to low Ca. Here, we describe one of the mutants, lcs1-1, isolated from the screen. The relative shoot growth of the mutant was reduced under the low-Ca conditions compared with the wild-type plants. Genetic mapping and genome resequencing revealed that lcs1-1 has one nonsynonymous mutation in the region of the chromosome responsible for the phenotype. The mutation is in Pleiotropic Regulatory Locus 1 (PRL1). An allelism test between lcs1-1 and a T-DNA inserted allele of prl1 demonstrates that the causal gene of lcs1-1 is PRL1. It has been reported that PRL1 is involved in sugar metabolism; however, the involvement of PRL1 in low-Ca tolerance has not been reported. Our results suggest a new insight connecting sugar metabolism with a mechanism for low-Ca tolerance in plants.
[Show abstract][Hide abstract] ABSTRACT: Human papillomavirus type 16 (HPV16) is a major cause of cervical cancer. We previously demonstrated that C-to-T and G-to-A hypermutations accumulated in the HPV16 genome by APOBEC3 expression in vitro. To investigate in vivo characteristics of hypermutation, differential DNA denaturation-PCR (3D-PCR) was performed using three clinical specimens obtained from HPV16-positive cervical dysplasia, and detected hypermutation from two out of three specimens. One sample accumulating hypermutations in both E2 and the long control region (LCR) was further subjected to Next-Generation Sequencing, revealing that hypermutations spread across the LCR and all early genes. Notably, hypermutation was more frequently observed in the LCR, which contains a viral replication origin and the early promoter. APOBEC3 expressed abundantly in an HPV16-positive cervix, suggesting that single-stranded DNA exposed during viral replication and transcription may be efficient targets for deamination. The results further strengthen a role of APOBEC3 in introducing HPV16 hypermutation in vivo.
[Show abstract][Hide abstract] ABSTRACT: Chemical communication is essential for the coordination of complex organisation in ant societies. Recent comparative genomic approaches have revealed that chemosensory genes are diversified in ant lineages, and suggest that this diversification is crucial for social organisation. However, how such diversified genes shape the peripheral chemosensory systems remains unknown. In this study, we annotated and analysed the gene expression profiles of chemosensory proteins (CSPs), which transport lipophilic compounds toward chemosensory receptors in the carpenter ant, Camponotus japonicus. Transcriptome analysis revealed 12 CSP genes and phylogenetic analysis showed that 3 of these are lineage-specifically expanded in the clade of ants. RNA sequencing and real-time quantitative polymerase chain reaction revealed that, among the ant specific CSP genes, two of them (CjapCSP12 and CjapCSP13) were specifically expressed in the chemosensory organs and differentially expressed amongst ant castes. Furthermore, CjapCSP12 and CjapCSP13 had a ratio of divergence at non-synonymous and synonymous sites (dN/dS) greater than 1, and they were co-expressed with CjapCSP1, which is known to bind cuticular hydrocarbons. Our results suggested that CjapCSP12 and CjapCSP13 were functionally differentiated for ant-specific chemosensory events, and that CjapCSP1, CjapCSP12, and CjapCSP13 work cooperatively in the antennal chemosensilla of worker ants.
[Show abstract][Hide abstract] ABSTRACT: Plant developmental processes are co-ordinated with the status of cell metabolism, not only in mitochondria but also in plastids.
In Arabidopsis thaliana, succinic semialdehyde (SSA), a GABA shunt metabolite, links the specific mitochondrial metabolic pathway to shoot development.
To understand the mechanism of SSA-mediated development, we isolated a succinic semialdehyde dehydrogenase (ssadh) suppressor mutant, affected in its ability to catalyze SSA to succinic acid. We found that pleiotropic developmental phenotypes
of ssadh are suppressed by a mutation in GLUTAMATE-1-SEMIALDEHYDE 2, 1-AMINOMUTASE 2 (GSA2), which encodes a plastidial enzyme converting glutatamate-1-semialdehyde to 5-aminolevulinic acid (5-ALA). In addition,
a mutation in either HEMA1 or GSA1, two other enzymes for 5-ALA synthesis, also suppressed ssadh fully and partially, respectively. Furthermore, exogenous application of 5-ALA and SSA disturbed leaf development. These
results suggest that metabolism in both mitochondria and plastids affect shoot development.
[Show abstract][Hide abstract] ABSTRACT: The low frequency of meiotic recombination in chromosomal regions other than hotspots is a general obstacle to efficient breeding. A number of active genes are present in recombination-repressed centromeric regions in higher eukaryotes, suggesting that suppression of meiotic recombination prevents shuffling of genes within a centromeric region. In this study, by using an inter-subspecific cross of Oryza sativa L., we show that modification of inactive chromatin states by either genetic or chemical inhibition of chromatin modifying proteins induced changes in both the position of meiotic recombination and, unexpectedly, the pattern of segregation distortion of parental alleles. Antisense knockdown of rice homologues of DECREASE IN DNA METHYLATION1, which is required for the maintenance of heterochromatin in Arabidopsis thaliana, induced a recombination hotspot in a centromeric region accompanied by a steep increase in the proportion of heterozygotes. Our results describe a previously undocumented phenomenon in which artificial chromatin modification could be used to change the pattern of segregation distortion in rice and open up novel possibilities for efficient crop breeding.
[Show abstract][Hide abstract] ABSTRACT: Maternal investment for offspring’s growth and survival is widespread among diverse organisms [1, 2 and 3]. Vertical symbiont transmission via maternal passage is also pivotal for offspring’s growth and survival in many organisms [4, 5 and 6]. Hence, it is expected that vertical symbiont transmission may coevolve with various organismal traits concerning maternal investment in offspring. Here we report a novel phenotypic syndrome entailing morphological, histological, behavioral, and ecological specializations for maternal investment and vertical symbiont transmission in stinkbugs of the family Urostylididae [7, 8 and 9]. Adult females develop huge ovaries exaggerated for polysaccharide excretion, possess novel ovipositor-associated organs for vertical transmission of a bacterial symbiont (“Candidatus Tachikawaea gelatinosa”), and lay eggs covered with voluminous symbiont-supplemented jelly. Newborns hatch in midwinter, feed solely on the jelly, acquire the symbiont, and grow during winter. In spring, the insects start feeding on plant sap, wherein the symbiont localizes to a specialized midgut region and supplies essential amino acids deficient in the host’s diet. The reduced symbiont genome and host-symbiont cospeciation indicate their obligate association over evolutionary time. Experimental deprivation of the jelly results in nymphal mortality, whereas restoration of the jelly leads to recovered nymphal growth, confirming that the jelly supports nymphal growth in winter. Chemical analyses demonstrate that the galactan-based jelly contains a sufficient quantity of amino acids to sustain nymphal growth to the third instar. The versatile biological roles of the symbiont-containing egg-covering jelly highlight intricate evolutionary interactions between maternal resource investment and vertical symbiont transmission, which are commonly important for offspring’s growth, survival, and ecological adaptation.
Current Biology 09/2014; 24(20). DOI:10.1016/j.cub.2014.08.065 · 9.57 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Cell-to-cell communication is a fundamental mechanism for coordinating developmental and physiological events in multicellular organisms. Heterotrimeric G proteins are key molecules that transmit extracellular signals; similarly, CLAVATA signaling is a crucial regulator in plant development. Here, we show that Arabidopsis thaliana Gβ mutants exhibit an enlarged stem cell region, which is similar to that of clavata mutants. Our genetic and cell biological analyses suggest that the G protein beta-subunit1 AGB1 and RPK2, one of the major CLV3 peptide hormone receptors, work synergistically in stem cell homeostasis through their physical interactions. We propose that AGB1 and RPK2 compose a signaling module to facilitate meristem development.
[Show abstract][Hide abstract] ABSTRACT: The ciliate Paramecium bursaria harbors several hundred cells of the green-alga Chlorella sp. in their cytoplasm. Irrespective of the mutual relation between P. bursaria and the symbiotic algae, both cells retain the ability to grow without the partner. They can easily reestablish endosymbiosis when put in contact with each other. Consequently, P. bursaria is an excellent model for studying cell-cell interaction and the evolution of eukaryotic cells through secondary endosymbiosis between different protists. Despite the importance of this organism, no genomic resources have been identified for P. bursaria to date. This investigation compared gene expressions through RNA-Seq analysis and de novo transcriptome assembly of symbiont-free and symbiont-bearing host cells.
To expedite the process of gene discovery related to the endosymbiosis, we have undertaken Illumina deep sequencing of mRNAs prepared from symbiont-bearing and symbiont-free P. bursaria cells. We assembled the reads de novo to build the transcriptome. Sequencing using Illumina HiSeq2000 platform yielded 232.3 million paired-end sequence reads. Clean reads filtered from the raw reads were assembled into 68,175 contig sequences. Of these, 10,557 representative sequences were retained after removing Chlorella sequences and lowly expressed sequences. Nearly 90% of these transcript sequences were annotated by similarity search against protein databases. We identified differentially expressed genes in the symbiont-bearing P. bursaria cells relative to the symbiont-free cells, including heat shock 70 kDa protein and glutathione S-transferase.
This is the first reported comprehensive sequence resource of Paramecium - Chlorella endosymbiosis. Results provide some keys for the elucidation of secondary endosymbiosis in P. bursaria. We identified P. bursaria genes that are differentially expressed in symbiont-bearing and symbiont-free conditions.
[Show abstract][Hide abstract] ABSTRACT: Sex chromosomes turn over rapidly in some taxonomic groups, where closely related species have different sex chromosomes. Although there are many examples of sex chromosome turnover, we know little about the functional roles of sex chromosome turnover in phenotypic diversification and genomic evolution. The sympatric pair of Japanese threespine stickleback (Gasterosteus aculeatus) provides an excellent system to address these questions: the Japan Sea species has a neo-sex chromosome system resulting from a fusion between an ancestral Y chromosome and an autosome, while the sympatric Pacific Ocean species has a simple XY sex chromosome system. Furthermore, previous quantitative trait locus (QTL) mapping demonstrated that the Japan Sea neo-X chromosome contributes to phenotypic divergence and reproductive isolation between these sympatric species. To investigate the genomic basis for the accumulation of genes important for speciation on the neo-X chromosome, we conducted whole genome sequencing of males and females of both the Japan Sea and the Pacific Ocean species. No substantial degeneration has yet occurred on the neo-Y chromosome, but the nucleotide sequence of the neo-X and the neo-Y has started to diverge, particularly at regions near the fusion. The neo-sex chromosomes also harbor an excess of genes with sex-biased expression. Furthermore, genes on the neo-X chromosome showed higher non-synonymous substitution rates than autosomal genes in the Japan Sea lineage. Genomic regions of higher sequence divergence between species, genes with divergent expression between species, and QTL for inter-species phenotypic differences were found not only at the regions near the fusion site, but also at other regions along the neo-X chromosome. Neo-sex chromosomes can therefore accumulate substitutions causing species differences even in the absence of substantial neo-Y degeneration.
[Show abstract][Hide abstract] ABSTRACT: Background
Although the infection rate of Helicobacter suis is significantly lower than that of Helicobacter pylori, the H. suis infection is associated with a high rate of gastric mucosa-associated lymphoid tissue (MALT) lymphoma. In addition, in vitro cultivation of H. suis remains difficult, and some H. suis-infected patients show negative results on the urea breath test (UBT).Materials and Methods
Female C57BL/6J mice were orally inoculated with mouse gastric mucosal homogenates containing H. suis strains TKY or SNTW101 isolated from a cynomolgus monkey or a patient suffering from nodular gastritis, respectively. The high-purity chromosomal DNA samples of H. suis strains TKY and SNTW101 were prepared from the infected mouse gastric mucosa. The SOLiD sequencing of two H. suis genomes enabled comparative genomics of 20 Helicobacter and 11 Campylobacter strains for the identification of the H. suis-specific nucleotide sequences.ResultsOral inoculation with mouse gastric mucosal homogenates containing H. suis strains TKY and SNTW101 induced gastric MALT lymphoma and the formation of gastric lymphoid follicles, respectively, in C57BL/6J mice. Two conserved nucleotide sequences among six H. suis strains were identified and were used to design diagnostic PCR primers for the detection of H. suis.Conclusions
There was a strong association between the H. suis infection and gastric diseases in the C57BL/6 mouse model. PCR diagnosis using an H. suis-specific primer pair is a valuable method for detecting H. suis in gastric biopsy specimens.
[Show abstract][Hide abstract] ABSTRACT: Endomembrane organization is important for various aspects of cell physiology, including membrane protein trafficking. To explore the molecular mechanisms regulating the trafficking of plasma membrane-localized proteins in plants, we screened for Arabidopsis mutants with defective localization of GFP-NIP5;1. Fluorescence imaging-based screening led to the isolation of a mutant, which accumulated abnormal intracellular aggregates labeled by GFP-NIP5;1. The aggregates appeared in epidermal cells in the root elongation zone and included the trans-Golgi network/early endosomes. Rough mapping and whole-genome sequencing identified the mutant as an allele of UDP-glucose 4-epimerase (uge) 4/root hair defective (rhd) 1/root epidermal bulgar (reb) 1, which was originally defined as a cell wall mutant. The responsible gene encodes UDP-glucose 4-epimerase 4 (UGE4), which functions in the biosynthesis of d-galactose, especially for the synthesis of the cell wall polysaccharide xyloglucan and arabinogalactan proteins (AGPs). The endomembrane aggregates in the mutants were absent in the presence of d-galactose, indicative of a requirement for a d-galactose-containing component in endomembrane organization. Genetic and pharmacological analyses suggested that the aggregates were not caused by the disruption of cell wall polysaccharides or the cytoskeleton. Overall, our results suggest that UGE4 activity in d-galactose synthesis is required for the structure of cell wall polysaccharides and endomembrane organization.
[Show abstract][Hide abstract] ABSTRACT: We isolated Burkholderia symbiont strain RPE64 from the bean bug Riptortus pedestris. Analysis of the complete 6.96-Mb genome, which consists of three chromosomes and two plasmids, will facilitate further understanding of insect-microbe symbiosis and the development of pest-control technologies.
[Show abstract][Hide abstract] ABSTRACT: Aims:
Dilated cardiomyopathy (DCM) is characterized by ventricular dilation associated with systolic dysfunction, which could be caused by mutations in lamina/C gene (LMNA). LMNA-linked DCM is severe in males in both human patients and a knock-in mouse model carrying a homozygous p.H222P mutation (LmnaH222P/H222P). The aim of this study was to investigate the molecular mechanisms underlying the gender difference of LMNA-linked DCM.
Methods and results:
A whole-exome analysis of a multiplex family with DCM exhibiting the gender difference revealed a DCM-linked LMNA mutation, p.R225X. Immunohistochemical analyses of neonatal rat cardiomyocytes expressing mutant LMNA constructs and heart samples from the LMNA-linked DCM patients and LmnaH222P/H222P mice demonstrated a nuclear accumulation of androgen receptor (AR) and its co-activators, serum response factor, and four-and-a-half LIM protein-2. Role of sex hormones in the gender difference was investigated in vivo using the LmnaH222P/H222P mice, where male and female mice were castrated and ovariectomized, respectively, or treated with testosterone or an antagonist of AR. Examination of the mice by echocardiography, followed by the analyses of histological changes and gene/protein expression profiles in the hearts, confirmed the involvement of testicular hormone in the disease progression and enhanced cardiac remodelling in the LmnaH222P/H222P mice.
These observations indicated that nuclear accumulation of AR was associated with the gender difference in LMNA-linked DCM.
Cardiovascular Research 04/2013; 99(3). DOI:10.1093/cvr/cvt106 · 5.94 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Next-generation sequencing (NGS) technologies enable the rapid production of an enormous quantity of sequence data. These powerful new technologies allow the identification of mutations by whole-genome sequencing. However, most reported NGS-based mapping methods, which are based on bulked segregant analysis, are costly and laborious. To address these limitations, we designed a versatile NGS-based mapping method that consists of a combination of low- to medium-coverage multiplex SOLiD (Sequencing by Oligonucleotide Ligation and Detection) and classical genetic rough mapping. Using only low to medium coverage reduces the SOLiD sequencing costs and, since just 10 to 20 mutant F 2 plants are required for rough mapping, the operation is simple enough to handle in a laboratory with limited space and funding. As a proof of principle, we successfully applied this method to identify the CTR1, which is involved in boron-mediated root development, from among a population of high boron requiring Arabidopsis thaliana mutants. Our work demonstrates that this NGS-based mapping method is a moderately priced and versatile method that can readily be applied to other model organisms.