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Kato, K.

Publications and source records attributed to Kato, K..

At least 55 records · Page 3Linked to original sources

Effective mechanical potential of cell-cell interaction in tissues harboring cavity and in cell sheet toward morphogenesis

Measuring mechanical forces of cell-cell interactions is important for studying morphogenesis in multicellular organisms. We previously reported an image-based statistical method for inferring effective mechanical potentials of pairwise cell-cell interactions by fitting cell tracking data with a theoretical model. However, whether this method is applicable to tissues with non-cellular components such as cavities remains elusive. Here we evaluated the applicability of the method to cavity-harboring tissues. Using synthetic data generated by simulations, we found that the effect of expanding cavities was added to the pregiven potentials used in the simulations, resulting in the inferred effective potentials having an additional repulsive component derived from the expanding cavities. Interestingly, simulations by using the effective potentials reproduced the cavity-harboring structures. Then, we applied our method to the mouse blastocysts, and found that the inferred effective potentials can reproduce the cavity-harboring structures. Pairwise potentials with additional repulsive components were also detected in two-dimensional cell sheets, by which curved sheets including tubes and cups were simulated. We conclude that our inference method is applicable to tissues harboring cavities and cell sheets, and the resultant effective potentials are useful to simulate the morphologies.

biophysics↗

Structural basis of EHEP-mediated offense against phlorotannin-induced defense from brown algae to protect akuBGL activity

The defensive-offensive associations between algae and herbivores determine marine ecology. Brown algae utilize phlorotannin as their chemical defense against the predator Aplysia kurodai, which uses {beta}-glucosidase (akuBGL) to digest the laminarin in algae to glucose. Moreover, A. kurodai employs Eisenia hydrolysis-enhancing protein (EHEP) as an offense to protect akuBGL activity from phlorotannin inhibition by precipitating phlorotannin. To underpin the molecular mechanism of this digestive-defensive-offensive system, we determined the structures of apo and tannic-acid (TNA, a phlorotannin-analog) bound form of EHEP, as well as apo akuBGL. EHEP consisted of three peritrophin-A domains formed in a triangle and bound TNA in the center without significant conformational changes. Structural comparison between EHEP and EHEP-TNA led us to find that EHEP can be resolubilized from phlorotannin-precipitation at an alkaline pH, which reflects a requirement in the digestive tract. akuBGL contained two GH1 domains, only one of which conserved the active site. Combining docking analysis, we propose the mechanisms by which phlorotannin inhibits akuBGL by occupying the substrate-binding pocket, and EHEP protects akuBGL against the inhibition by binding with phlorotannin to free the akuBGL pocket.

biochemistry↗

ATF4 Expression in Thermogenic Adipocytes is Required for Cold-Induced Thermogenesis in Mice via FGF21-Independent Mechanisms

In brown adipose tissue (BAT), short-term cold exposure induces the activating transcription factor 4 (ATF4), and its downstream target fibroblast growth factor 21 (FGF21). Induction of ATF4 in BAT in response to mitochondrial stress is required for thermoregulation, partially via upregulation of FGF21. In the present study, we tested the hypothesis that Atf4 and Fgf21 induction in BAT are both required for BAT thermogenesis by generating mice selectively lacking either Atf4 (ATF4 BKO) or Fgf21 (FGF21 BKO) in UCP1-expressing adipocytes. After 3 days of cold exposure, core body temperature was significantly reduced in ad-libitum-fed ATF4 BKO mice, which correlated with Fgf21 downregulation in brown and beige adipocytes, and impaired browning of white adipose tissue (WAT). Conversely, despite having reduced browning, FGF21 BKO mice had preserved core body temperature after cold exposure. Mechanistically, ATF4, but not FGF21, regulates amino acid import and metabolism in response to cold, likely contributing to BAT thermogenic capacity under ad libitum-fed conditions. Importantly, under fasting conditions, both ATF4 and FGF21 were required for thermogenesis in cold-exposed mice. Thus, ATF4 regulates BAT thermogenesis by activating amino acid metabolism in BAT in a FGF21-independent manner. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=152 SRC="FIGDIR/small/531964v2_ufig1.gif" ALT="Figure 1"> View larger version (45K): org.highwire.dtl.DTLVardef@1d2c3fdorg.highwire.dtl.DTLVardef@414f5forg.highwire.dtl.DTLVardef@1c1fa9eorg.highwire.dtl.DTLVardef@2cc815_HPS_FORMAT_FIGEXP M_FIG C_FIG

physiology↗

Structure of the Commander endosomal trafficking complex linked to X-linked intellectual disability/Ritscher-Schinzel syndrome

The Commander complex is required for endosomal recycling of diverse transmembrane cargos and is mutated in Ritscher-Schinzel syndrome. It comprises two subassemblies; Retriever composed of VPS35L, VPS26C and VPS29, and the CCC complex which contains ten subunits COMMD1-COMMD10 and two coiled-coil domain-containing (CCDC) proteins CCDC22 and CCDC93. Combining X-ray crystallography, electron cryomicroscopy and in silico predictions we have assembled a complete structural model of Commander. Retriever is distantly related to the endosomal Retromer complex but has unique features preventing the shared VPS29 subunit from interacting with Retromer-associated factors. The COMMD proteins form a distinctive hetero-decameric ring stabilised by extensive interactions with CCDC22 and CCDC93. These adopt a coiled-coil structure that connects the CCC and Retriever assemblies and recruits a sixteenth subunit, DENND10, to form the complete Commander complex. The structure allows mapping of disease-causing mutations and reveals the molecular features required for the function of this evolutionarily conserved trafficking machinery.

biochemistry↗

GDF15 Is Required for Cold-Induced Thermogenesis and Contributes to Improve Systemic Metabolic Health Following Loss of OPA1 in Brown Adipocytes

Previously we showed that mice lacking the protein optic atrophy 1 (OPA1 BKO) in brown adipose tissue (BAT) have induction of the activating transcription factor 4 (ATF4), which promotes fibroblast growth factor 21 (FGF21) secretion as a batokine. FGF21 increases metabolic rates at baseline conditions but is dispensable for the resistance to diet-induced obesity (DIO) reported in OPA1 BKO mice [1]. To determine alternative mediators of this phenotype, we performed transcriptome analysis, which revealed increased levels of growth differentiation factor 15 (GDF15) in BAT. To determine if ATF4 induction was mediated by the protein kinase R (PKR)-like endoplasmic reticulum kinase (PERK), and to evaluate the contribution of GDF15 to the resistance to DIO, we selectively deleted PERK or GDF15 in OPA1 BKO mice. Mice lacking both OPA1 and PERK in BAT had preserved induction of ATF4. Importantly, simultaneous deletion of OPA1 and GDF15 partially reversed the resistance to diet-induced obesity and abrogated the improvements in glucose tolerance. Furthermore, GDF15 mediated cold-induced thermogenesis, likely via sympathetic activation of iWAT. Taken together, our data indicate that PERK is dispensable for ATF4 induction, but GDF15 contributes to the resistance to DIO, and is required for glucose homeostasis and thermoregulation in OPA1 BKO mice.

physiology↗

Molecular engineering of emicizumab to improve hemophilia A treatment

Emicizumab, a factor (F)VIIIa-function mimetic bispecific antibody (BsAb) to FIXa and FX, has become an indispensable treatment for people with hemophilia A (PwHA). Although emicizumab is very potent, long-term outcomes from the clinical studies suggest that a small proportion of PwHA still experiences bleeds. Additionally, non-clinical studies indicate that the maximum cofactor activity of emicizumab is lower than international standard activity (100 IU/dL of FVIII). An increased cofactor activity BsAb would benefit such patients. Here, we report NXT007, a BsAb binding FIXa and FX developed through further engineering of emicizumab. Emicizumab has a common light chain, but through advances in antibody engineering, we were able to create a more potent BsAb with two new non-common light chains. After extensive optimization of the heavy and light chains, the resulting BsAb, NXT007, exerted in vitro thrombin generation (TG) activity in hemophilia A plasma equivalent to 100 IU/dL of FVIII when triggered by tissue factor. NXT007 demonstrated potent hemostatic activity in an acquired hemophilia A model in non-human primates at a much lower dosage than emicizumab, consistent with an around 30-fold dose shift in the in vitro TG activity between NXT007 and emicizumab. Moreover, together with Fc engineering that enhanced FcRn binding and reduced in vivo clearance, we demonstrate that NXT007 could be effective at a much lower dosage with a longer dosing interval compared to emicizumab. These non-clinical results suggest that NXT007 could maintain a non-hemophilic range of coagulation potential in PwHA and provides a rationale for its clinical testing.

bioengineering↗

The arms race in noncoding regions of transposable element: the evolution of anti-silencing and RNAi

Transposable elements (TEs) are among the most dynamic parts of genomes. Since TEs are potentially deleterious, eukaryotes silence them through epigenetic mechanisms such as DNA methylation and RNAi. We have previously reported that Arabidopsis TEs, called VANDALs, counteract epigenetic silencing through a group of sequence-specific anti-silencing proteins, VANCs. VANC proteins bind to noncoding regions of specific VANDAL copies and induce a loss of silent chromatin marks. Sequence-specific anti-silencing allows these TEs to proliferate with minimum host damage. Here, we show that RNAi efficiently targets noncoding regions of VANDAL TEs to silence them de novo. Target motifs of VANC, in turn, evolved to escape RNAi. Escaping RNAi could be the primary event leading to the differentiation of sequence-specific anti-silencing systems. We propose that this selfish behaviour of TEs paradoxically could make them less harmful to the host.

genetics↗

PNPO-PLP Axis Senses Prolonged Hypoxia by Regulating Lysosomal Activity

Oxygen is critical for all metazoan organisms on the earth and impacts various biological processes in physiological and pathological conditions. While oxygen sensing systems inducing acute hypoxic response, including HIF pathway, have been identified, those operating in prolonged hypoxia remain to be elucidated. Here, we show that pyridoxine 5-phosphate oxidase (PNPO) that catalyzes bioactivation of vitamin B6 serves as an oxygen sensor and regulates lysosomal activity in macrophages. Decline of PNPO activity under prolonged hypoxia reduced an active form of vitamin B6, pyridoxal 5-phosphate (PLP), and inhibited lysosomal activity, leading to the augmentation of inflammatory response of macrophages. The PNPO-PLP axis creates a distinct layer of oxygen sensing, which gradually turns down and up the PLP-dependent metabolism according to prolonged changes in oxygen availability.

biochemistry↗

Novel monoclonal antibodies showing broad neutralizing activity for SARS-CoV-2 variants including Omicrons BA.5 and BA.2.75

We identified novel neutralizing monoclonal antibodies against SARS-CoV-2 variants (including Omicron) from individuals received two doses of mRNA vaccination after they had been infected with wildtype. We named them MO1, MO2 and MO3. MO1 shows high neutralizing activity against authentic variants: D614G, Delta, BA.1, BA.1.1, BA.2, and BA.2.75 and BA.5. Our findings confirm that the wildtype-derived vaccination can induce neutralizing antibodies that recognize the epitopes conserved among the SARS-CoV-2 variants (including BA.5 and BA.2.75). The monoclonal antibodies obtained herein could serve as novel prophylaxis and therapeutics against not only current SARS-CoV-2 viruses but also future variants that may arise.

microbiology↗

Orchestration of Proteins in cyanobacterial Circadian Clock System 1

Circadian rhythm by Cyanobacteria is one of the simplest biological clocks: the clock consists of only three proteins, KaiA, KaiB and KaiC. Their oligomers, KaiA dimer (A2), KaiB tetramer (B4) and KaiC hexamer (C6) oscillate an association- disassociation cycle with 24hr period. In a widely accepted model, the oscillation process is as follows. From the viewpoint of a base unit (C6), C6 homo-oligomer [->] A2C6 complex [->] B6C6 complex [->] AnB6C6 complex (n[≤]12) [->]C6 homo-oligomer. In this study, Small-Angle X-ray Scattering, Contrast Matching-Small-Angle Neutron Scattering, Analytical Ultracentrifuge and phosphorylation-analysis PAGE measurements were performed to reveal the kinetics not only of KaiC hexamer but also of all components in a working Kai clock. The complementary analysis disclosed that the oscillation is not the single process as the widely accepted model but composed with synchronized multiple association-dissociation reactions between components. Namely, there are various reactions between components, which proceed simultaneously, in a working Kai-clock.

biophysics↗

RNA-triggered protein cleavage and cell death by the RNA-guided type III-E CRISPR-Cas nuclease-protease complex

The type III-E Cas7-11 effector nuclease forms a complex with a CRISPR RNA (crRNA) and the putative caspase-like protease Csx29, catalyzes crRNA-guided target RNA cleavage, and has been used for RNA targeting in eukaryotic cells. Here, we report cryo-electron microscopy structures of the Cas7-11-crRNA-Csx29 complex with and without target RNA, and demonstrate that target RNA binding induces a conformational change in Csx29 and results in the protease activation. Biochemical analysis confirmed that Cas7-11-bound Csx29 cleaves Csx30 in a target RNA-dependent manner. Reconstitution of the system in bacteria uncovered Csx30-dependent cellular toxicity regulated by Csx31, and that Csx29-mediated cleavage produces toxic Csx30 fragments, promoting cell death. We find that Csx30 can bind both Csx31 and the associated sigma factor RpoE, suggesting Csx30 can inhibit RpoE and modulate cellular stress response towards infection. Overall, the RNA-guided nuclease-protease activities of the Cas7-11-Csx29 effector complex facilitate protease-based programmed cell death.

molecular biology↗

Structure of a monomeric photosystem I core associated with iron-stress-induced-A proteins from Anabaena sp. PCC 7120

Iron-stress-induced-A proteins (IsiAs) are expressed in cyanobacteria under iron-deficient conditions. The cyanobacterium Anabaena sp. PCC 7120 has four isiA genes; however, their binding property and functional roles in PSI are still missing. We analyzed a cryo-electron microscopy structure of a PSI-IsiA supercomplex isolated from Anabaena grown under an iron-deficient condition. The PSI-IsiA structure contains six IsiA subunits associated with the PsaA side of a PSI core monomer. Three of the six IsiA subunits are identified as IsiA1 and IsiA2. The PSI-IsiA structure lacks a PsaL subunit; instead, a C-terminal domain of IsiA2 is inserted at the position of PsaL, which inhibits the oligomerization of PSI, leading to the formation of a monomer. Furthermore, excitation-energy transfer from IsiAs to PSI appeared with a time constant of 55 ps. These findings provide novel insights into both the molecular assembly of the Anabaena IsiA family and the functional roles of IsiAs.

plant biology↗

The double-layered structure of amyloid-β assemblage on GM1-containing membranes catalytically promotes fibrillization

Alzheimers disease (AD) is associated with progressive accumulation of amyloid-{beta} (A{beta}) cross-{beta} fibrils in the brain. A{beta} species tightly associated with GM1 ganglioside, a glycosphingolipid abundant in neuronal membranes, promote amyloid fibril formation; therefore, they could be attractive clinical targets. However, the active conformational state of A{beta} in GM1-containing lipid membranes is still unknown. The present solid-state nuclear magnetic resonance study revealed a nonfibrillar A{beta} assemblage characterized by a double-layered antiparallel {beta}-structure specifically formed on GM1 ganglioside clusters. Our data show that this unique assemblage was not transformed into fibrils on GM1-containing membranes, but could promote conversion of monomeric A{beta} into fibrils, suggesting that a solvent-exposed hydrophobic layer provides a catalytic surface evoking A{beta} fibril formation. Our findings will offer structural clues for designing drugs targeting catalytically active A{beta} conformational species for the development of anti-AD therapeutics.

biophysics↗

Structural insights into an evolutionary turning-point of photosystem I from prokaryotes to eukaryotes

Photosystem I (PSI) contributes to light-conversion reactions; however, its oligomerization state is variable among photosynthetic organisms. Herein we present a 3.8-[A] resolution cryo-electron microscopic structure of tetrameric PSI isolated from a glaucophyte alga Cyanophora paradoxa. The PSI tetramer is organized in a dimer of dimers form with a C2 symmetry. Different from cyanobacterial PSI tetramer, two of the four monomers are rotated around 90{degrees}, resulting in a totally different pattern of monomer-monomer interactions. Excitation-energy transfer among chlorophylls differs significantly between Cyanophora and cyanobacterial PSI tetramers. These structural and spectroscopic features reveal characteristic interactions and energy transfer in the Cyanophora PSI tetramer, thus offering an attractive idea for the changes of PSI from prokaryotes to eukaryotes.

plant biology↗

Appropriate tension sensitivity of α-catenin ensures rounding morphogenesis of epithelial spheroids

The adherens junction (AJ) is an actin filament-anchoring junction. It plays a central role in epithelial morphogenesis through cadherin-based recognition and adhesion among cells. The stability and plasticity of AJs are required for the morphogenesis. An actin-binding -catenin is an essential component of the cadherin-catenin complex and functions as a tension transducer that changes its conformation and induces AJ development in response to tension. Despite much progress in understanding molecular mechanisms of tension sensitivity of -catenin, its significance on epithelial morphogenesis is still unknown. Here we show that the tension sensitivity of -catenin is essential for epithelial cells to form round spheroids through proper multicellular rearrangement. Using a novel in vitro suspension culture model, we found that epithelial cells form round spheroids even from rectangular-shaped cell masses with high aspect ratios without using high tension and that hypersensitive mutants affected this morphogenesis. Analyses of AJ formation and cellular tracking during rounding morphogenesis showed cellular rearrangement, probably through AJ remodeling. The rearrangement occurs at the cell mass level, but not single-cell level. Hypersensitive -catenin mutant-expressing cells did not show cellular rearrangement at the cell mass level, suggesting that proper AJ plasticity requires appropriate tension sensitivity of -catenin.

cell biology↗

Cancer malignancy is correlated with up-regulation of PCYT2-mediated glycerol phosphate modification of α-dystroglycan

The dystrophin-glycoprotein complex connects the cytoskeleton with base membrane components such as laminin through unique O-glycans displayed on -dystroglycan (-DG). Genetic impairment of elongation of these glycans causes congenital muscular dystrophies. We previously identified that glycerol phosphate (GroP) can cap the core part of the -DG O-glycans and terminate their further elongation. This study examined the possible roles of the GroP modification in cancer malignancy, focusing on colorectal cancer. We found that the GroP modification critically depends on PCYT2, which serves as CDP-Gro synthase. Furthermore, we identified a significant positive correlation between cancer progression and GroP modification, which also correlated positively with PCYT2 expression. Moreover, we demonstrate that GroP modification promotes the migration of cancer cells. Based on these findings, we propose that the GroP modification by PCYT2 disrupts the glycan-mediated cell adhesion to the extracellular matrix and thereby enhances cancer metastasis. Thus, the present study suggests the possibility of novel approaches for cancer treatment by targeting the PCYT2-mediated GroP modification.

biochemistry↗

Structural basis for the absence of low-energy chlorophylls responsible for photoprotection from a primitive cyanobacterial PSI

Photosystem I (PSI) of photosynthetic organisms is a multi-subunit pigment-protein complex and functions in light harvesting and photochemical charge-separation reactions, followed by reduction of NADP to NADPH required for CO2 fixation. PSI from different photosynthetic organisms has a variety of chlorophylls (Chls), some of which are at lower-energy levels than its reaction center P700, a special pair of Chls, and are called low-energy Chls. However, the site of low-energy Chls is still under debate. Here, we solved a 2.04-[A] resolution structure of a PSI trimer by cryo-electron microscopy from a primitive cyanobacterium Gloeobacter violaceus PCC 7421, which has no low-energy Chls. The structure showed absence of some subunits commonly found in other cyanobacteria, confirming the primitive nature of this cyanobacterium. Comparison with the known structures of PSI from other cyanobacteria and eukaryotic organisms reveals that one dimeric and one trimeric Chls are lacking in the Gloeobacter PSI. The dimeric and trimeric Chls are named Low1 and Low2, respectively. Low2 does not exist in some cyanobacterial and eukaryotic PSIs, whereas Low1 is absent only in Gloeobacter. Since Gloeobacter is susceptible to light, this indicates that Low1 serves as a main photoprotection site in most oxyphototrophs, whereas Low2 is involved in either energy transfer or energy quenching in some of the oxyphototrophs. Thus, these findings provide insights into not only the functional significance of low-energy Chls in PSI, but also the evolutionary changes of low-energy Chls responsible for the photoprotection machinery from photosynthetic prokaryotes to eukaryotes.

plant biology↗

Aqueous extract of Phyllanthus niruri protects against severe malaria by blocking erythrocyte invasion and modulating the host immune response

Plasmodium falciparum parasites are the major cause of malaria across Africa. Due to the appearance of multi-drug resistant parasites, new antimalarial drugs are needed. The medicinal plant Phyllanthus niruri is being used to treat fever and other symptoms of malaria in Nigeria; however, little is known about its antimalarial mechanisms. Here, we show that aqueous extract of P. niruri (PE) has multiple antimalarial effects, including anti-parasitic and host immunomodulatory activity. We found that co-culture of P. falciparum with PE drastically reduced parasite number, but PE did not inhibit parasite development or rupture; rather, it blocked erythrocytes invasion. In addition, we identified Astragalin as one of the antimalarial compounds which are contained in PE. Moreover, we found that PE suppresses the inflammatory activity and apoptosis of immune cells (T cells) and astrocytes and neurons in the central nervous system (CNS). Furthermore, we confirmed that oral administration of PE to mice suppressed parasite growth, excessive inflammation, CNS dysfunction, and the development of experimental cerebral malaria in an in vivo murine malaria model. Our findings demonstrate that PE has multiple effects on malaria progression, targeting both parasite and host cells.

pathology↗