Biology subjects

Kato, M.

Publications and source records attributed to Kato, M..

71 records · Page 4Linked to original sources

bioRxiv2020

Dynamic structural order of a low complexity domain facilitates assembly of intermediate filaments

The coiled-coil domains of intermediate filament (IF) proteins are flanked by regions of low sequence complexity. Whereas IF coiled-coil domains assume dimeric and tetrameric conformations on their own, maturation of eight tetramers into cylindrical IFs is dependent upon either "head" or "tail" domains of low sequence complexity. Here we confirm that the tail domain required for assembly of Drosophila Tm1 IFs functions by forming labile cross-{beta} interactions. These interactions are seen in polymers made from the tail domain alone as well as assembled IFs formed by the intact Tm1 protein. The ability to visualize such interactions in situ within the context of a discrete cellular assembly lends support to the concept that equivalent interactions may be used in organizing other dynamic aspects of cell morphology. One Sentence SummaryA new form of protein folding that interconverts between the structured and unstructured states controls assembly of intermediate filaments.

biochemistry↗

bioRxiv2020

Transiently structured head domains control intermediate filament assembly

Low complexity (LC) head domains 92 and 108 residues in length are, respectfully, required for assembly of neurofilament light (NFL) and desmin intermediate filaments (IFs). As studied in isolation, these IF head domains interconvert between states of conformational disorder and labile, {beta}-strand-enriched polymers. Solid state nuclear magnetic resonance (ss-NMR) spectroscopic studies of NFL and desmin head domain polymers reveal spectral patterns consistent with structural order. A combination of intein chemistry and segmental isotope labeling allowed preparation of fully assembled NFL and desmin IFs that could also be studied by ss-NMR. Assembled IFs revealed spectra overlapping with those observed for {beta}-strand-enriched polymers formed from the isolated NFL and desmin head domains. Phosphorylation and disease causing mutations reciprocally alter NFL and desmin head domain self-association, yet commonly impede IF assembly. These observations show how facultative structural assembly of LC domains via labile, {beta}-strand-enriched self-interactions may broadly influence cell morphology.

biochemistry↗

bioRxiv2019

Redox-mediated regulation of a labile, evolutionarily conserved cross-β structure formed by the TDP43 low complexity domain

An evolutionarily conserved low complexity (LC) domain is found within a 152 residue segment localized to the carboxyl-terminal region of the TDP43 RNA-binding protein. This TDP43 LC domain contains ten conserved methionine residues. Self-association of this domain leads to the formation of liquid-like droplets composed of labile, cross-{beta} polymers. Exposure of polymers to low concentrations of H2O2 leads to a phenomenon of droplet melting that can be reversed upon exposure of the oxidized protein to the MsrA and MsrB methionine sulfoxide reductase enzymes, thioredoxin, thioredoxin reductase and NADPH. Morphological features of the cross-{beta} polymers were revealed by a method of H2O2-mediated footprinting. Similar TDP43 LC domain footprints were observed in highly polymerized, hydrogel samples, liquid-like droplet samples, and living cells. The ability of H2O2 to impede cross-{beta} polymerization was abrogated by a prominent ALS-causing mutation that changes methionine residue 337 to valine. These observations offer potentially useful insight into the biological role of TDP43 in facilitating synapse-localized translation, as well as aberrant aggregation of the protein in neurodegenerative disease.

biochemistry↗

bioRxiv2019

Cables2 Is a novel Smad2-regulatory factor essential for early embryonic development in mice

CDK5 and Abl enzyme substrate 2 (Cables2), a member of the Cables family that has a C-terminal cyclin box-like domain, is widely expressed in adult mouse tissues. However, the physiological role of Cables2 in vivo is unknown. We show here that Cables2-deficiency causes post-gastrulation embryonic lethality in mice. The mutant embryos progress to gastrulation, but then arrest, and fail to grow. Analysis of gene expression patterns reveals that formation of the anterior visceral endoderm and the primitive streak is impaired in Cables2-deficient embryos. Tetraploid complementation analyses support the critical requirement of Cables2 in both the epiblast and visceral endoderm for progression of embryogenesis. In addition, we show that Cables2 physically interacts with a key mediator of the canonical Nodal pathway, Smad2, and augments its transcriptional activity. These findings provide novel insights into the essential role of Cables2 in the early embryonic development in mice.

developmental biology↗

bioRxiv2019

Massive parallel variant characterization identifies NUDT15 alleles associated with thiopurine toxicity

As a prototype of genomics-guided precision medicine, individualized thiopurine dosing based on pharmacogenetics is a highly effective way to mitigate hematopoietic toxicity of this class of drugs. Recently, NUDT15 deficiency was identified as a novel genetic cause of thiopurine toxicity, and NUDT15-informed preemptive dose reduction is quickly adopted in clinical settings. To exhaustively identify pharmacogenetic variants in this gene, we developed massively parallel NUDT15 function assays to determine variants effect on protein abundance and thiopurine cytotoxicity. Of the 3,097 possible missense variants, we characterized the abundance of 2,922 variants and found 54 hotspot residues at which variants resulted in complete loss of protein stability. Analyzing 2,935 variants in the thiopurine cytotoxicity-based assay, we identified 17 additional residues where variants altered NUDT15 activity without affecting protein stability. We identified structural elements key to NUDT15 stability and/or catalytical activity with single amino-acid resolution. Functional effects for NUDT15 variants accurately predicted toxicity risk alleles in 2,398 patients treated with thiopurines, with 100% sensitivity and specificity, in contrast with poor performance of bioinformatic prediction algorithms. In conclusion, our massively parallel variant function assays identified 1,103 deleterious NUDT15 variants, providing a comprehensive reference of variant function and vastly improving the ability to implement pharmacogenetics-guided thiopurine treatment individualization.

genomics↗

bioRxiv2019

SMAD3 Determines Conventional versus Plasmacytoid Dendritic Cell Fates

Transforming growth factor (TGF)-{beta} plays crucial roles in differentiation of dendritic cells (DC). However, molecular mechanisms how TGF-{beta} regulates DC differentiation remain largely unknown. Here, we show that selective repression of one of the TGF-{beta} receptor-regulated SMADs (R-SMADs), SMAD3 directs conventional DC (cDC) differentiation, whereas maintenance of SMAD3 is indispensable for plasmacytoid DC (pDC) differentiation. Expression of SMAD3 was specifically downregulated in CD115+ common DC progenitor (CDP), pre-cDCs and cDCs. SMAD3 deficient mice showed a significant reduction in pre-pDCs and pDCs with increased CDP, pre-cDCs and cDCs. SMAD3 upregulated the pDC-related genes: SPI-B, E2-2 and IKAROS, while it repressed FLT3 and the cDC-related genes: IRF4 and ID2. STAT3 and a SMAD transcriptional co-repressor, c-SKI repressed SMAD3 for cDC differentiation, whereas canonical SMAD-mediated TGF-{beta} signalling maintained SMAD3 for pDC differentiation. Thus, SMAD3 is the pivotal determinant to bifurcate cDC and pDC differentiation in the steady-state condition.

immunology↗

bioRxiv2019

Single-cell DNA and RNA sequencing reveals the dynamics of intra-tumor heterogeneity in a colorectal cancer model

Intra-tumor heterogeneity (ITH) encompasses cellular differences in tumors and is related to clinical outcomes, such as drug resistance. However, little is known about the dynamics of ITH, owing to the lack of time-series analysis at the single-cell level. We performed single-cell exome and transcriptome sequencing of 200 cells and investigated how ITH is generated from one single cell in a mouse colorectal cancer model. The ITH of the transcriptome increased after transplantation from cultured organoids, while that of the exome decreased. Mutations generated in the culture did not greatly change at the transplantation at the bulk-cell level. The RNA ITH increase was due to the emergence of new transcriptional subpopulations. In contrast to the initial cells expressing mesenchymal-marker genes, new subpopulations repressed these genes at transplantation. Analyses of colorectal cancer data from The Cancer Genome Atlas revealed a high proportion of metastatic cases in human subjects with expression patterns similar to the new cell subpopulations in mouse. These results suggest that the birth of transcriptional subpopulations may be a key for adaptation to drastic micro-environmental changes when cancer cells have sufficient genetic alterations at later tumor stages. This study revealed an evolutionary dynamics of single-cell RNA and DNA changes in tumor progression, giving insights into the mesenchymal-epithelial transformation of tumor cells at metastasis in colorectal cancer.

genomics↗

bioRxiv2019

Genome-Wide Fetalization of Enhancer Architecture in Heart Disease

Heart disease is associated with re-expression of key transcription factors normally active only during prenatal development of the heart. However, the impact of this reactivation on the genome-wide regulatory landscape in heart disease has remained obscure. Here we show that pervasive epigenomic changes occur in heart disease, with thousands of regulatory sequences reacquiring fetal-like chromatin signatures. We used RNA-seq and ChIP-seq targeting a histone modification associated with active transcriptional enhancers to generate genome-wide enhancer maps from left ventricle tissue from 18 healthy controls and 18 individuals with idiopathic dilated cardiomyopathy (DCM). Healthy individuals had a highly reproducible epigenomic landscape, consisting of more than 31,000 predicted heart enhancers. In contrast, we observed reproducible disease-associated gains or losses of activity at more than 7,500 predicted heart enhancers. Next, we profiled human fetal heart tissue by ChIP-seq and RNA-seq. Comparison with adult tissues revealed that the heart disease epigenome and transcrip-tome both shift toward a fetal-like state, with 3,400 individual enhancers sharing fetal regulatory properties. Our results demonstrate widespread epigenomic changes in DCM, and we provide a comprehensive data resource (http://heart.lbl.gov) for the mechanistic exploration of heart disease etiology.

genomics↗

bioRxiv2019

Management of family relationship informationfor a three-generation cohort study

A system for inputting and storing family information, named \"BirThree Enrollment,\" was developed to promote a birth and three-generation cohort study (BirThree Cohort Study), and this system was operated successfully. In the study, it was necessary to satisfy many operational demands. Input information is overwritten and changed continuously. Complex kinship information must be quickly and accurately input and corrected, and information on those families not yet recruited must be retrieved. For these purposes, many devices are needed, from an input interface to the internal data structure. In the field of genetic statistics, a simple standard expressive form is used for describing family structure. This form has sufficient information for genetics; however, we developed this form further for our purposes in conducting the BirThree Cohort Study. To provide information about family roles as required in the BirThree Cohort Study, we expanded the data structure, and constructed the system that is able to be used for the daily operation. In our system, family pedigree information is stored along with initial clinical information, and enabled the input of all self-reported information to the data base. Operators are able to input this family information before the day is out. As a result, when recruitment is completed, family information will be completed concurrently. Therefore, it is possible to immediately know a certain persons family structure. By using our system, data correction was improved dramatically, and the system was operated successfully. This study is the first report of the method for storing three generations of family data.

epidemiology↗

bioRxiv2018

Life in coastal pebble sediment: Unique interstitial organism community and selective feeding on meiobenthos by interstitial fishes (Luciogobius: Gobiidae)

Interstitial areas of coastal pebble sediment in the Japanese Archipelago are inhabited by extremely elongated gobies of the genus Luciogobius, which are characterized by an increased number of vertebrae and reduction of scales, eyes, and fins. To explore the little-known interstitial life of Luciogobius gobies, we investigated the diets of two interstitial Luciogobius species, L. elongatus and L. grandis, and the interstitial organism communities of the gobies microhabitats in an exposed gravelly coast in Shirahama, southern Japan. The interstitial organism community in pebbly sediment was dominated by minute arthropods such as harpacticoids, isopods, and ostracods, presenting a marked contrast to the communities in sandy sediments, which are dominated by nematodes and turbellarians. The gut contents of the two goby species were composed exclusively of interstitial organisms, especially harpacticoides and isopods. Although each prey assemblage was roughly similar to the interstitial organism community in the corresponding microhabitat, marked preferences for harpacticoids and flabelliferan isopods were detected in L. elongatus and L. grandis, respectively. Irrespective of their intense feeding of harpacticoids, rare catches of large isopods were suggested to be nutritionally important for the gobies. These results suggest that the Luciogobius gobies are the first known fishes that depend exclusively on interstitial organisms, and that selective feeding upon meiobenthos may facilitate the coexistence of several interstitial goby species in pebbly sediment.

ecology↗

bioRxiv2018

PHASE TRANSITIONED NUCLEAR OSKAR PROMOTES CELL DIVISION OF DROSOPHILA PRIMORDIAL GERM CELLS

Germ granules are non-membranous ribonucleoprotein granules deemed the hubs for post-transcriptional gene regulation and functionally linked to germ cell fate across species. Little is known about the physical properties of germ granules and how these relate to germ cell function. Here we study two types of germ granules in the Drosophila embryo: cytoplasmic germ granules that instruct primordial germ cells (PGCs) formation and nuclear germ granules within early PGCs with unknown function. We show that cytoplasmic and nuclear germ granules are phase transitioned condensates nucleated by Oskar protein that display liquid as well as hydrogel-like properties. Focusing on nuclear granules, we find that Oskar drives their formation in heterologous cell systems. Multiple, independent Oskar protein domains synergize to promote granule phase separation. Deletion of Oskars nuclear localization sequence specifically ablates nuclear granules in cell systems. In the embryo, nuclear germ granules promote germ cell divisions thereby increasing PGC number for the next generation.

cell biology↗

bioRxiv2018

A cross-organism framework for supervised enhancer prediction with epigenetic pattern recognition and targeted validation

Enhancers are important noncoding elements, but they have been traditionally hard to characterize experimentally. Only a few mammalian enhancers have been validated, making it difficult to train statistical models for their identification properly. Instead, postulated patterns of genomic features have been used heuristically for identification. The development of massively parallel assays allows for the characterization of large numbers of enhancers for the first time. Here, we developed a framework that uses Drosophila STARR-seq data to create shape-matching filters based on enhancer-associated meta-profiles of epigenetic features. We combined these features with supervised machine learning algorithms (e.g., support vector machines) to predict enhancers. We demonstrated that our model could be applied to predict enhancers in mammalian species (i.e., mouse and human). We comprehensively validated the predictions using a combination of in vivo and in vitro approaches, involving transgenic assays in mouse and transduction-based reporter assays in human cell lines. Overall, the validations involved 153 enhancers in 6 mouse tissues and 4 human cell lines. The results confirmed that our model can accurately predict enhancers in different species without re-parameterization. Finally, we examined the transcription-factor binding patterns at predicted enhancers and promoters in human cell lines. We demonstrated that these patterns enable the construction of a secondary model effectively discriminating between enhancers and promoters.

bioinformatics↗

bioRxiv2018

Evolution of shell flattening and the loss of coiling in top shells (Gastropoda: Trochidae: Fossarininae) on wave-swept rock reefs

Flattening of coiled shells has occurred in numerous gastropod lineages, probably as an adaptation to life in narrow protected spaces, such as crevices or the undersides of rocks. While several genera in the top snail family (Trochidae) have flattened shells, two Fossarininae genera, Broderipia and Roya, are unique in having shells that are limpet-like and zygomorphic, lacking any trace of coiling. The sister genera of these two genera are Fossarina and Synaptocochlea, both of which have coiled shells and live in rock crevices or the vacant shells of sessile organisms. Although Broderipia has recently been identified as living symbiotically in the pits of sea urchins, the habitat and biology of Roya are poorly known. After an extensive search for rare Roya snails on rocky shores of the Japanese Archipelago, we found live Roya eximia snails on intertidal/subtidal rock surfaces exposed to strong waves. The Roya snails crawled swiftly over wave-swept rock surfaces at low tide, while they retreated into the vacant shells of barnacles at high tide, where they adhered firmly to the inner wall. A survey of the macrobenthic communities around the snail habitat showed that Roya snails inhabited only wave-swept rocks of exposed reefs, where the substrata was covered by encrusting red algae and barnacles. Despite the abnormal shell morphology, the radula was similar to other species in the subfamily, and the diet of Roya snails was mainly pennate diatoms. The limpet-like shell of Roya caused loss of coiling and contraction of the soft body, acquisition of a zygomorphic flat body, expansion of the foot sole and loss of the operculum. All of these changes improved tolerance of strong waves and the ability to cling to rock surfaces, and thus enabled a lifestyle split between wave-swept rock surfaces and refugia of vacant barnacle shells.

evolutionary biology↗

bioRxiv2018

Immuno-genomic PanCancer Landscape Reveals Diverse Immune Escape Mechanisms and Immuno-Editing Histories

Immune reactions in the tumor micro-environment are one of the cancer hallmarks and emerging immune therapies have been proven effective in many types of cancer. To investigate cancer genome-immune interactions and the role of immuno-editing or immune escape mechanisms in cancer development, we analyzed 2,834 whole genomes and RNA-seq datasets across 31 distinct tumor types from the PanCancer Analysis of Whole Genomes (PCAWG) project with respect to key immuno-genomic aspects. We show that selective copy number changes in immune-related genes could contribute to immune escape. Furthermore, we developed an index of the immuno-editing history of each tumor sample based on the information of mutations in exonic regions and pseudogenes. Our immuno-genomic analyses of pan-cancer analyses have the potential to identify a subset of tumors with immunogenicity and diverse background or intrinsic pathways associated with their immune status and immuno-editing history.

genomics↗

bioRxiv2017

Systematic mapping of chromatin state landscapes during mouse development

Embryogenesis requires epigenetic information that allows each cell to respond appropriately to developmental cues. Histone modifications are core components of a cells epigenome, giving rise to chromatin states that modulate genome function. Here, we systematically profile histone modifications in a diverse panel of mouse tissues at 8 developmental stages from 10.5 days post conception until birth, performing a total of 1,128 ChIP-seq assays across 72 distinct tissue-stages. We combine these histone modification profiles into a unified set of chromatin state annotations, and track their activity across developmental time and space. Through integrative analysis we identify dynamic enhancers, reveal key transcriptional regulators, and characterize the role of chromatin-based repression in developmental gene regulation. We also leverage these data to link enhancers to putative target genes, revealing connections between coding and non-coding sequence variation in disease etiology. Our study provides a compendium of resources for biomedical researchers, and achieves the most comprehensive view of embryonic chromatin states to date.

genomics↗

bioRxiv2017

Structure Of Allium Lachrymatory Factor Synthase Elucidates Catalysis On Sulfenic Acid Substrate

Natural lachrymatory effects are invoked by small volatile S-oxide compounds. They are produced through alkene sulfenic acids by the action of lachrymatory factor synthase (LFS). Here we present the crystal structures of onion LFS (AcLFS) revealed in solute-free and two solute-stabilized forms. Each structure adopts a single seven-stranded helix-grip fold possessing an internal pocket. Mutagenesis analysis localized the active site to a layer near the bottom of the pocket, which is adjacent to the deduced key residues Arg71, Glu88, and Tyr114. Solute molecules visible on the active site have suggested that AcLFS accepts various small alcohol compounds as well as its natural substrate, and they inhibit this substrate according to their chemistry. Structural homologs have been found in the SRPBCC superfamily, and comparison of the active sites has demonstrated that the electrostatic potential unique to AcLFS could work in capturing the substrate in its specific state. Finally, we propose a rational catalytic mechanism based on intramolecular proton shuttling in which the microenvironment of AcLFS can bypass the canonical [1,4]-sigmatropic rearrangement principle established by microwave studies. Beyond revealing how AcLFS generates the lachrymatory compound, this study provides insights into the molecular machinery dealing with highly labile organosulfur species.\n\nSignificance statementCrushing of onion liberates a volatile compound, syn-propanethial S-oxide (PTSO), which causes lachrymatory effect on humans. We present the crystal structures of onion LFS (AcLFS), the enzyme responsible for natural production of PTSO. AcLFS features a barrel-like fold, and mutagenic and inhibitory analyses revealed that the key residues are present in the central pocket, harboring highly concentrated aromatic residues plus a dyad motif. The architecture of AcLFS is widespread among proteins with various biological functions, such as abscisic acid receptors and polyketide cyclases, and comparisons with these homologs indicate that unique steric and electronic properties maintain the pocket as a reaction compartment. We propose the molecular mechanism behind PTSO generation and shed light on biological decomposition of short-lived sulfur species.

biochemistry↗

bioRxiv2016

Empirical Bayes estimation of semi-parametric hierarchical mixture models for unbiased characterization of polygenic disease architectures

Genome-wide association studies (GWAS) suggest that the genetic architecture of complex diseases consists of unexpectedly numerous variants with small effect sizes. However, the polygenic architectures of many diseases have not been well characterized due to lack of simple and fast methods for unbiased estimation of the underlying proportion of disease-associated variants and their effect-size distribution. Applying empirical Bayes estimation of semi-parametric hierarchical mixture models to GWAS summary statistics, we confirmed that schizophrenia was extremely polygenic ([~] 40% risk variants of independent genome-wide SNPs, most within odds ratio (OR)=1.03), whereas rheumatoid arthritis was less polygenic ([~] 4 to 8% risk variants, significant portion reaching OR=1.05 to 1.1). For rheumatoid arthritis, stratified estimations revealed that expression quantitative loci in blood explained large genetic variance, and low- and high-frequency derived alleles were prone to be risk and protective, respectively, suggesting a predominance of deleterious-risk and advantageous-protective mutation. Despite genetic correlation, effect-size distributions for schizophrenia and bipolar disorder differed across allele frequency. These analyses distinguished disease polygenic architectures and provided clues for etiological differences in complex diseases.

genomics↗