Biology subjects

Tanaka, K.

Publications and source records attributed to Tanaka, K..

At least 91 records · Page 5Linked to original sources

bioRxiv2020

Phospholipid flippases and Sfk1 are essential for the retention of ergosterol in the plasma membrane

Sterols are important lipid components of the plasma membrane (PM) in eukaryotic cells, but it is unknown how the PM retains sterols at a high concentration. Phospholipids are asymmetrically distributed in the PM, and phospholipid flippases play an important role in generating this phospholipid asymmetry. Here, we provide evidence that phospholipid flippases are essential for retaining ergosterol in the PM of yeast. A mutant in three flippases, Dnf1-Lem3, Dnf2-Lem3, and Dnf3-Crf1, and a membrane protein, Sfk1, showed a severe growth defect. We recently identified Sfk1 as a PM protein involved in phospholipid asymmetry. The PM of this mutant showed high permeability and low density, and many nutrient transporters failed to localize to the PM. Staining with the sterol probe filipin and the expression of a sterol biosensor revealed that ergosterol was not retained in the PM. Instead, ergosterol accumulated in an esterified form in lipid droplets. We propose that ergosterol is retained in the PM by the asymmetrical distribution of phospholipids and the action of Sfk1. Once phospholipid asymmetry is severely disrupted, sterols might be exposed on the cytoplasmic leaflet of the PM and actively transported to the endoplasmic reticulum by sterol transfer proteins.

cell biology↗

bioRxiv2020

Innate behavior sequence progression by peptide-mediated interorgan crosstalk

Innate behaviors consist of a succession of genetically-hardwired motor and physiological subprograms that can be coupled to drastic morphogenetic changes. How these integrative responses are orchestrated is not completely understood. Here, we provide insight into these mechanisms by studying pupariation, a multi-step innate behavior of fly larvae that is critical for survival during metamorphosis. We find that the steroid-hormone ecdysone triggers parallel pupariation neuromotor and morphogenetic subprograms, which include the induction of the relaxin-peptide hormone, Dilp8, in the epidermis. Dilp8 acts on six Lgr3-positive thoracic interneurons to couple both subprograms in time and to instruct neuromotor subprogram switching during behavior. Our work reveals that interorgan feedback gates progression between subunits of an innate behavior and points to an ancestral neuromodulatory function of relaxin signaling.

animal behavior and cognition↗

bioRxiv2020

Ribosomal protein S7 ubiquitination during ER stress in yeast is associated with selective mRNA translation and stress outcome

eIF2 phosphorylation-mediated translational regulation is crucial for global translation repression by various stresses, including the unfolded protein response (UPR). However, translational control during UPR has not been demonstrated in yeast. This study investigated ribosome ubiquitination-mediated translational controls during UPR. Tunicamycin-induced ER stress enhanced the levels of ubiquitination of the ribosomal proteins uS10, uS3 and eS7. Not4-mediated monoubiquitination of eS7A was required for resistance to tunicamycin, whereas E3 ligase Hel2-mediated ubiquitination of uS10 was not. Ribosome profiling showed that the monoubiquitination of eS7A was crucial for translational regulation, including the upregulation of the spliced form of HAC1 (HAC1i) mRNA and the downregulation of Histidine triad NucleoTide-binding 1 (HNT1) mRNA. Downregulation of the deubiquitinating enzyme complex Upb3-Bre5 increased the levels of ubiquitinated eS7A during UPR in an Ire1-independent manner. These findings suggest that the monoubiquitination of ribosomal protein eS7A plays a crucial role in translational controls during the ER stress response in yeast.

molecular biology↗

bioRxiv2020

Phosphatidylserine prevents the generation of a protein-free giant plasma membrane domain in yeast

Membrane phase separation accompanied with micron-scale domains of lipids and proteins occurs in artificial membranes; however, a similar large phase separation has not been reported in the plasma membrane of the living cells. We demonstrate here that a stable micron-scale protein-free region is generated in the plasma membrane of the yeast mutants lacking phosphatidylserine. We named this region the "void zone". Transmembrane proteins, peripheral membrane proteins, and certain phospholipids are excluded from the void zone. The void zone is rich in ergosterol and requires ergosterol and sphingolipids for its formation. These characteristics of the void zone are similar to the properties of the cholesterol-enriched domain in phase-separated artificial membranes. We propose that phosphatidylserine prevents the formation of the void zone by preferentially interacting with ergosterol. We also found that void zones were frequently in contact with vacuoles, in which a membrane domain was also formed at the contact site. Summary statementYeast cells lacking phosphatidylserine generate protein-free plasma membrane domains, and vacuoles contact with this domain. This is the first report of micron-scale plasma membrane domains in living cells.

cell biology↗

bioRxiv2020

TGN/EE SNARE protein SYP61 is ubiquitinated and required for carbon/nitrogen-nutrient responses in Arabidopsis

Ubiquitination is a post-translational modification with reversible attachment of the small protein ubiquitin, which is involved in numerous cellular processes including membrane trafficking. For example, ubiquitination of cargo proteins is known to regulate their subcellular dynamics, and plays important roles in plant growth and stress adaptation. However, the regulatory mechanism of the trafficking machinery components remains elusive. Here, we report Arabidopsis trans-Golgi network/early endosome (TGN/EE) localized soluble N-ethylmaleimide sensitive factor attachment protein receptor (SNARE) protein SYP61 as a novel ubiquitination target of a membrane localized ubiquitin ligase ATL31. SYP61 is a key component of membrane trafficking in Arabidopsis. SYP61 was ubiquitinated with K63-linked chain by ATL31 in vitro and in plants. The knockdown mutants of SYP61 were hypersensitive to the disrupted carbon (C)/nitrogen (N)-nutrient stress, suggesting its critical role in plant homeostasis in response to nutrients. We also found the ubiquitination status of SYP61 is affected by C/N-nutrient availability. These results provided possibility that ubiquitination of SNARE protein has important role in plant physiology.

plant biology↗

bioRxiv2020

Pattern recognition receptors confer plant salt tolerance via WRKY18/WRKY40 transcription factors

Pattern recognition receptors (PRRs) bind microbe- and damage-associated molecular patterns (MAMPs/DAMPs, respectively) to enhance host immunity in animals and plants. Here, we report that PRRs also confer salt tolerance in the model plant Arabidopsis thaliana following recognition of cognate ligands, such as bacterial flagellin and EF-Tu and the endogenous Pep peptides. Pattern-triggered salt tolerance (PTST) requires the PRR-associated kinases BAK1 and BIK1, and the NADPH oxidase RBOHD. Transcriptome profiling reveals an inventory of PTST target genes, which increase or acquire salt responsiveness following an exposure to immunogenic patterns. In their regulatory DNA sequences, specific binding sites for a subset of WRKY transcription factors are over-represented. Accordingly, PTST requires WRKY40 and WRKY18, which activate salt tolerance-related genes but attenuate pathogen defense-related genes, including the EDS1 immunity activator. PRR signaling leads to sustained WRKY40/WRKY18 accumulation under salt stress and utilizes both WRKYs for salt tolerance. The PRR-WRKY40/WRKY18 module also confers salt tolerance after challenge with non-pathogenic bacteria. Our findings give molecular insight into signaling plasticity underlying biotic-abiotic stress cross-tolerance in plants conferred by PRRs.

plant biology↗

bioRxiv2020

Arabidopsis SMN2/HEN2, encoding DEAD-box RNA helicase, governs proper expression of the resistance gene SMN1/RPS6 and is involved in dwarf, autoimmune phenotypes of mekk1 and mpk4

In Arabidopsis thaliana, a mitogen-activated protein kinase pathway, MEKK1-MKK1/MKK2-MPK4, is important for basal resistance, and disruption of this pathway results in dwarf, autoimmune phenotypes. To elucidate the complex mechanisms activated by the disruption of this pathway, we have previously developed a mutant screening system based on a dwarf autoimmune line that overexpressed the N-terminal regulatory domain of MEKK1. Here, we report that the second group of mutants, smn2, had defects in the SMN2 gene, encoding a DEAD-box RNA helicase. SMN2 is identical to HEN2, whose function is vital for the nuclear RNA exosome because it provides non-ribosomal RNA specificity for RNA turnover, RNA quality control, and RNA processing. Aberrant SMN1/RPS6 transcripts were detected in smn2 and hen2 mutants. Disease resistance against Pseudomonas syringae pv. tomato DC3000 (hopA1), which is conferred by SMN1/RPS6, was decreased in smn2 mutants, suggesting a functional connection between SMN1/RPS6 and SMN2/HEN2. We produced double mutants mekk1smn2 and mpk4smn2 to determine whether the smn2 mutations suppress the dwarf, autoimmune phenotypes of the mekk1 and mpk4 mutants, as the smn1 mutations do. As expected, the mekk1 and mpk4 phenotypes were suppressed by the smn2 mutations. These results suggested that SMN2 is involved in proper function of SMN1/RPS6. The GO enrichment analysis using RNA-seq data showed that defense genes were downregulated in smn2, suggesting positive contribution of SMN2 to genome-wide expression of defense-related genes. In conclusion, this study provides novel insight into plant immunity via SMN2/HEN2, an essential component of the nuclear RNA exosome.

plant biology↗

bioRxiv2020

Impaired expression of chloroplast HSP90C chaperone activates plant defense responses leading to a disease symptom-like phenotype

RNA-seq analysis of a transgenic tobacco plant, i-hpHSP90C, in which chloroplast HSP90C genes can be silenced in an artificially inducible manner resulting in the development of chlorosis, revealed the up- and down-regulation of 2746 and 3490 genes, respectively. Gene Ontology analysis of these differentially expressed genes indicated the upregulation of ROS-responsive genes, the activation of the innate immunity and cell death pathways, and the downregulation of genes involved in photosynthesis, plastid organization, and cell cycle. Cell death was confirmed by trypan blue staining and electrolyte leakage assay and the H2O2 production by diaminobenzidine staining. The upregulation of ER stress-responsive genes suggested the interplay between ER protein quality control and chloroplast or immune response. The results collectively suggest that the reduced levels of HSP90C chaperone leads the plant to develop chlorosis primarily through the global downregulation of chloroplast and photosynthesis-related genes and additionally through the light-dependent production of ROS, followed by the activation of immune responses including the cell death. HighlightInduced silencing of HSP90C gene caused the upregulation of stress-responsive genes and the activation of innate immune response, which resulted in the chlorosis development accompanying cell death.

plant biology↗

bioRxiv2020

High resolution spatial transcriptome analysis by photo-isolation chemistry

In multicellular organisms, individual cells are characterized by their gene expression profiles and the spatial interactions among cells enable the elaboration of complex functions. Expression profiling in spatially defined regions is crucial to elucidate cell interactions and functions. Here, we established a transcriptome profiling method coupled with photo-isolation chemistry (PIC) that allows the determination of expression profiles specifically from photo-irradiated regions of whole tissues. PIC uses photo-caged oligodeoxynucleotides for in situ reverse transcription. After photo-irradiation of limited areas, gene expression was detected from at least 10 cells in the tissue sections. PIC transcriptome analysis detected genes specifically expressed in small distinct areas of the mouse embryo. Thus, PIC enables transcriptome profiles to be determined from limited regions at a spatial resolution up to the diffraction limit.

genomics↗

bioRxiv2020

Mediator-Microorganism Interaction in Microbial Solar Cell: a Fluo-Electrochemical Insight

Microbial solar cells that mainly rely on the use of photosynthesic organisms are a promising alternative to photovoltaics for solar electricity production. In that way, we propose a new approach involving electrochemistry and fluorescence techniques. The coupled set-up Electro-Pulse-Amplitude-Modulation ("e-PAM") enables the simultaneous recording of the produced photocurrent and fluorescence signals from the photosynthetic chain. This methodology was validated with a suspension of green alga Chlamydomonas reinhardtii in interaction with an exogenous redox mediatior (2,6-dichlorobenzoquinone; DCBQ). The balance between photosynthetic chain events (PSII photochemical yield, quenching) and the extracted electricity can be monitored overtime. More particularly, the non photochemical quenching induced by DCBQ mirrors the photocurrent. This set-up thus helps to distinguish the electron harvesting from some side effects due to quinones in real time. It therefore paves the way for future analyses devoted to the choice of the experimental conditions (redox mediator, photosynthetic organisms...) to find the best electron extraction.

biophysics↗

bioRxiv2020

Latrophilins are essential for endothelial junctional fluid shear stress mechanotransduction

Endothelial cell (EC) responses to fluid shear stress (FSS) are crucial for vascular development, adult physiology and disease. PECAM1 is an important transducer but earlier events remain poorly understood. We therefore investigated heterotrimeric G proteins in FSS sensing. Knockdown (KD) in ECs of single G proteins had little effect but combined depletion of Gi and Gq/11 blocked all known PECAM1-dependent responses. Re-expression of Gi2 and Gq but not Gi1 and Gi3 rescued these effects. Sequence alignment and mutational studies identified that K307 in Gi2 and Gq/11 (Q306 in Gi1/3), determines participation in flow signaling. We developed pull-down assays for measuring G activation and found that this residue, localized to the GPCR interface, determines activation by FSS. We developed a protocol for affinity purification of GPCRs on activated Gs, which identified latrophilins (ADGRLs) as specific upstream interactors for Gi2 and Gq/11. Depletion of latrophilin-2 blocked EC activation of Gi2 and Gq, downstream events in vitro, and flow-dependent vascular morphogenesis in zebrafish embryos. Surprisingly, latrophilin-2 depletion also blocked flow activation of two additional pathways activated at cell-cell junctions, Smad1/5 and Notch1, independently of G proteins. Latrophilins are thus central mediators of junctional shear stress mechanotransduction via G protein-dependent and -independent mechanisms.

molecular biology↗

bioRxiv2020

Inactive variants of death receptor p75NTR reduce Alzheimer's neuropathology by interfering with APP internalization

A prevalent model of Alzheimers disease (AD) pathogenesis postulates the generation of neurotoxic fragments derived from the amyloid precursor protein (APP) after its internalization to endocytic compartments. However, the molecular pathways that regulate APP internalization and intracellular trafficking in neurons are unknown. Here we report that 5xFAD mice, an animal model of AD, expressing signaling-deficient variants of the p75 neurotrophin receptor (p75NTR) show greater neuroprotection from AD neuropathology than animals lacking this receptor. p75NTR knock-in mice lacking the death domain or transmembrane Cys259 showed lower levels of A{beta} species, amyloid plaque burden, gliosis, mitochondrial stress and neurite dystrophy than global knock-outs. Strikingly, long-term synaptic plasticity and memory, which are completely disrupted in 5xFAD mice, were fully recovered in the knock-in mice. Mechanistically, we found that p75NTR interacts with APP and regulates its internalization in hippocampal neurons. Inactive p75NTR variants internalized much slower and to lower levels than wild type p75NTR, favoring non-amyloidogenic APP cleavage by reducing APP internalization and colocalization with BACE1, the critical protease for generation of neurotoxic APP fragments. These results reveal a novel pathway that directly and specifically regulates APP internalization, amyloidogenic processing and disease progression, and suggest that inhibitors targeting the p75NTR transmembrane domain may be an effective therapeutic strategy in AD.

neuroscience↗

bioRxiv2019

Comparative population genetic structure of two Ixodidae ticks (Ixodes ovatus and Haemaphysalis flava) in Niigata Prefecture, Japan

Ixodid tick species such as Ixodes ovatus and Haemaphysalis flava are essential vectors of tick-borne diseases in Japan. In this study, we investigated the population genetic structures and gene flow of I. ovatus and H. flava as affected by the tick host mobility. We hypothesized that I. ovatus and H. flava may have differences in their genetic structure due to the low mobility of small rodent hosts of I. ovatus at the immature stage in contrast to the mediated dispersal of avian hosts for immature H. flava. We collected 307 adult I. ovatus and 220 adult H. flava from 29 and 17 locations across Niigata Prefecture, Japan. We investigated the genetic structure at two mitochondrial loci (cox1, 16S rRNA gene). For I. ovatus, pairwise FST and analysis of molecular variance (AMOVA) analyses of cox1 sequences indicated significant genetic variation among populations. Both cox1 and 16S rRNA markers showed non-significant genetic variation among locations for H. flava. The Bayesian tree and haplotype network of cox1 marker for I. ovatus samples in Niigata Prefecture found 3 genetic groups wherein most haplotypes in group 2 were distributed in low altitudinal areas. When we added cox1 sequences of I. ovatus from China to the phylogenetic analysis, three genetic groups (China 1, China 2, and Niigata and Hokkaido, Japan) were formed in the tree suggesting the potential for cryptic species in the genetic group in Japan. Our results support our hypothesis and suggest that the host preference of ticks at the immature stage may influence the genetic structure and gene flow of the ticks. This information is vital in understanding the tick-host interactions in the field to better understand the tick-borne disease transmission and in designing an effective tick control program.

genetics↗

bioRxiv2019

Time-of-day dependent responses of cyanobacterial cellular viability against oxidative stress

As an adaptation to periodic fluctuations of environmental light, photosynthetic organisms have evolved a circadian clock. Control by the circadian clock of many cellular physiological functions, including antioxidant enzymes, metabolism and the cell cycle, has attracted attention in the context of oxidative stress tolerance. However, since each physiological function works in an integrated manner to deal with oxidative stress, whether or not cell responses to oxidative stress are under circadian control remains an open question. In fact, circadian rhythms of oxidative stress tolerance have not yet been experimentally demonstrated. In the present work, we applied an assay using methyl viologen (MV), which generates reactive oxygen species (ROS) under light irradiation, and experimentally verified the circadian rhythms of oxidative stress tolerance in photosynthetic cells of the cyanobacterium Synechococcus elongatus PCC7942, a standard model species for investigation of the circadian clock. Here, we report that ROS generated by MV treatment causes damage to stroma components and not to the photosynthetic electron transportation chain, leading to reduced cell viability. The degree of decrease in cell viability was dependent on the subjective time at which oxidative stress was applied. Thus, oxidative stress tolerance was shown to exhibit circadian rhythms. In addition, the rhythmic pattern of oxidative stress tolerance disappeared in mutant cells lacking the essential clock genes. Notably, ROS levels changed periodically, independent of the MV treatment. Thus, we demonstrate for the first time that in cyanobacterial cells, oxidative stress tolerance shows circadian oscillation.

plant biology↗

bioRxiv2019

Population-specific and transethnic genome-wide analyses reveal distinct and shared genetic risks of coronary artery disease

To elucidate the genetics of coronary artery disease (CAD) in the Japanese population, we conducted a large-scale genome-wide association study (GWAS) of 168,228 Japanese (25,892 cases and 142,336 controls) with genotype imputation using a newly developed reference panel of Japanese haplotypes including 1,782 CAD cases and 3,148 controls. We detected 9 novel disease-susceptibility loci and Japanese-specific rare variants contributing to disease severity and increased cardiovascular mortality. We then conducted a transethnic meta-analysis and discovered 37 additional novel loci. Using the result of the meta-analysis, we derived a polygenic risk score (PRS) for CAD, which outperformed those derived from either Japanese or European GWAS. The PRS prioritized risk factors among various clinical parameters and segregated individuals with increased risk of long-term cardiovascular mortality. Our data improves clinical characterization of CAD genetics and suggests the utility of transethnic meta-analysis for PRS derivation in non-European populations.

genetics↗

bioRxiv2019

Convergent evolution of sex-specific leg ornaments in Drosophilidae - from cells to structures

The origin, diversification, and secondary loss of sexually dimorphic characters are common in animal evolution. In some cases, structurally and functionally similar traits have evolved independently in multiple lineages. Prominent examples of such traits include the male-specific grasping structures that develop on the front legs of many dipteran insects. In this report, we describe the evolution and development of one of these structures, the male-specific "sex brush". The sex brush is composed of densely packed, irregularly arranged modified bristles and is found in several distantly related lineages in the family Drosophilidae. Phylogenetic analysis using 250 genes from over 200 species provides modest support for a single origin of the sex brush followed by many secondary losses; however, independent origins of the sex brush cannot be ruled out completely. We show that sex brushes develop in very similar ways in all brush-bearing lineages. The dense packing of brush hairs is explained by the specification of bristle precursor cells at a near-maximum density permitted by the lateral inhibition mechanism, as well as by the reduced size of the surrounding epithelial cells. In contrast to the female and the ancestral male condition, where bristles are arranged in stereotypical, precisely spaced rows, cell migration does not contribute appreciably to the formation of the sex brush. The complex phylogenetic history of the sex brush can make it a valuable model for investigating coevolution of sex-specific morphology and mating behavior.

evolutionary biology↗

bioRxiv2019

Transethnic meta-analysis of genome-wide association studies identifies three new loci and characterizes population-specific differences for coronary artery disease

BackgroundGenome-wide association studies (GWAS) provided many biological insights into coronary artery disease (CAD), but these studies were mainly performed in Europeans. GWAS in diverse populations have the potential to advance our understanding of CAD.\n\nMethods and ResultsWe conducted two GWAS for CAD in the Japanese population, which included 12,494 cases and 28,879 controls, and 2,808 cases and 7,261 controls, respectively. Then, we performed transethnic meta-analysis using the results of the CARDIoGRAMplusC4D 1000 Genomes meta-analysis with UK Biobank. We identified 3 new loci on chromosome 1q21 (CTSS), 10q26 (WDR11-FGFR2), and 11q22 (RDX-FDX1). Quantitative trait locus analyses suggested the association of CTSS and RDX-FDX1 with atherosclerotic immune cells. Tissue/cell type enrichment analysis showed the involvement of arteries, adrenal glands and fat tissues in the development of CAD. Finally, we performed tissue/cell type enrichment analysis using East Asian-frequent and European-frequent variants according to the risk allele frequencies, and identified significant enrichment of adrenal glands in the East Asian-frequent group while the enrichment of arteries and fat tissues was found in the European-frequent group. These findings indicate biological differences in CAD susceptibility between Japanese and Europeans.\n\nConclusionsWe identified 3 new loci for CAD and highlighted the genetic differences between the Japanese and European populations. Moreover, our transethnic analyses showed both shared and unique genetic architectures between the Japanese and Europeans. While most of the underlying genetic bases for CAD are shared, further analyses in diverse populations will be needed to elucidate variations fully.

genetics↗

bioRxiv2019

A Pro-Diabetogenic mtDNA Polymorphism in the Mitochondrial-Derived Peptide, MOTS-c

Type 2 Diabetes (T2D) is an emerging public health problem in Asia. An Asian mitochondrial DNA variation m.1382A>C (rs111033358) leads to a K14Q amino acid replacement in MOTS-c, an insulin sensitizing mitochondrial-derived peptide. Meta-analysis of three cohorts (n=27,527, J-MICC, MEC, and TMM) showed that males but not females with the C-allele exhibit a higher prevalence of T2D. Furthermore, in J-MICC, only males with the C-allele in the lowest tertile of physical activity increased their prevalence of T2D, demonstrating a kinesio-genomic interaction. High-fat fed, male mice injected with MOTS-c showed reduced weight and improved glucose tolerance, but not K14Q-MOTS-c treated mice. Like the human data, female mice were unaffected. Mechanistically, K14Q-MOTS-c leads to diminished insulin-sensitization in vitro. Thus, the m.1382A>C polymorphism is associated with susceptibility to T2D in men, possibly interacting with exercise, and contributing to the risk of T2D in sedentary males by reducing the activity of MOTS-c.

genetics↗