Biology subjects

Tanaka, T.

Publications and source records attributed to Tanaka, T..

At least 91 records · Page 5Linked to original sources

bioRxiv2021

SARS-CoV-2 ORF6 disturbs nucleocytoplasmic trafficking to advance the viral replication

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the virus responsible for the coronavirus disease 2019 pandemic. ORF6 is known to antagonize the interferon signaling by inhibiting the nuclear translocation of STAT1. Here we show that ORF6 acts as a virulence factor through two distinct strategies. First, ORF6 directly interacts with STAT1 in an IFN-independent manner to inhibit its nuclear translocation. Second, ORF6 directly binds to importin 1, which is a nuclear transport factor encoded by KPNA2, leading to a significant suppression of importin 1-mediated nuclear transport. Furthermore, we found that KPNA2 knockout enhances the viral replication, suggesting that importin 1 suppresses the viral propagation. Additionally, the analyses of gene expression data revealed that importin 1 levels decreased significantly in the lungs of older individuals. Taken together, SARS-CoV-2 ORF6 disrupts the nucleocytoplasmic trafficking to accelerate the viral replication, resulting in the disease progression, especially in older individuals.

microbiology↗

bioRxiv2021

Dermal macrophages set pain sensitivity by modulating tissue NGF levels through SNX25-Nrf2 signaling

Crosstalk between peripheral neurons and immune cells plays important roles in pain sensation. We identified sorting nexin 25 (Snx25) as a pain-modulating gene in a transgenic mouse line with reduced pain behavior. Snx25 conditional-KO (cKO) in monocyte/macrophage-lineage cells but not in the peripheral sensory neurons reduced pain responses in both normal and neuropathic conditions. Cross transplantation experiments of bone marrows between cKO and wild type (WT) mice revealed that cKO macrophages caused dull phenotype in WT mice and WT macrophages in turn increased pain behavior in cKO mice. SNX25 in dermal macrophages enhances NGF (one of the key factors in pain sensation) production by inhibiting ubiquitin-mediated degradation of Nrf2, a transcription factor that activates Ngf mRNA synthesis. We conclude that dermal macrophages set pain sensitivity by producing and secreting NGF into the dermis in addition to their host defense functions.

immunology↗

bioRxiv2020

Anti-severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) potency of Mefloquine as an entry inhibitor in vitro

Coronavirus disease 2019 (COVID-19) has caused serious public health, social, and economic damage worldwide and effective drugs that prevent or cure COVID-19 are urgently needed. Approved drugs including Hydroxychloroquine, Remdesivir or Interferon were reported to inhibit the infection or propagation of severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2), however, their clinical efficacies have not yet been well demonstrated. To identify drugs with higher antiviral potency, we screened approved anti-parasitic/anti-protozoal drugs and identified an anti-malarial drug, Mefloquine, which showed the highest anti-SARS-CoV-2 activity among the tested compounds. Mefloquine showed higher anti-SARS-CoV-2 activity than Hydroxychloroquine in VeroE6/TMPRSS2 and Calu-3 cells, with IC50 = 1.28 M, IC90 = 2.31 M, and IC99 = 4.39 M in VeroE6/TMPRSS2 cells. Mefloquine inhibited viral entry after viral attachment to the target cell. Combined treatment with Mefloquine and Nelfinavir, a replication inhibitor, showed synergistic antiviral activity. Our mathematical modeling based on the drug concentration in the lung predicted that Mefloquine administration at a standard treatment dosage could decline viral dynamics in patients, reduce cumulative viral load to 7% and shorten the time until virus elimination by 6.1 days. These data cumulatively underscore Mefloquine as an anti-SARS-CoV-2 entry inhibitor.

microbiology↗

bioRxiv2020

Plasma irradiation efficiently inactivates the coronaviruses mouse hepatitis virus and SARS-CoV-2

Many inactivation methods have been shown to inactivate SARS-CoV-2 for safe and efficient diagnostic methods. COVID-19 is caused by airborne infection of SARS-CoV-2, and therefore, methods of inactivating the virus efficiently and safely are crucial for reducing the risk of airborne infection. In this regard, the effect of plasma discharge on the infectivity of the coronaviruses mouse hepatitis virus (MHV) and SARS-CoV-2 was tested. Plasma discharge efficiently reduced the infectivity of both coronaviruses. Treatment of SARS-CoV-2 in culture medium with a plasma discharge resulted in 95.17% viral inactivation after plasma irradiation after 1 hour (hr), 99.54% inactivation after 2 hrs and 99.93% inactivation after 3 hrs. Similar results were obtained for MHV. The results indicated that plasma discharge effectively and safely inactivated the airborne coronaviruses and may be useful in minimizing the risk of airborne infection of SARS-CoV-2.

microbiology↗

bioRxiv2020

Leukemia stemness and co-occurring mutations drive resistance to IDH inhibitors in acute myeloid leukemia

Allosteric inhibitors of mutant IDH1 or IDH2 induce terminal differentiation of the mutant leukemic blasts and provide durable clinical responses in approximately 40% of acute myeloid leukemia (AML) patients with the mutations. However, primary resistance and acquired resistance to the drugs are major clinical issues. To understand the molecular underpinnings of clinical resistance to IDH inhibitors (IDHi), we performed multipronged genomic analyses (DNA sequencing, RNA sequencing and cytosine methylation profiling) in longitudinally collected specimens from 68 IDH1- or IDH2-mutant AML patients treated with the inhibitors. The analysis revealed that leukemia stemness is a major driver of primary resistance to IDHi, whereas selection of mutations in RUNX1/CEBPA or RAS-RTK pathway genes was the main driver of acquired resistance to IDHi, along with BCOR, homologous IDH gene, and TET2. These data suggest that targeting stemness and certain high-risk co-occurring mutations may overcome resistance to IDHi in AML.

cancer biology↗

bioRxiv2020

Epileptic Spike Detection by Using a Linear-Phase Convolutional Neural Network

To cope with the lack of highly skilled professionals, machine learning with proper signal processing is key for establishing automated diagnostic-aid technologies with which to conduct epileptic electroencephalogram (EEG) testing. In particular, frequency filtering with the appropriate passbands is essential for enhancing the biomarkers--such as epileptic spike waves--that are noted in the EEG. This paper introduces a novel class of neural networks (NNs) that have a bank of linear-phase finite impulse response filters at the first layer as a preprocessor that can behave as bandpass filters that extract biomarkers without destroying waveforms because of a linear-phase condition. Besides, the parameters of the filters are also data-driven. The proposed NNs were trained with a large amount of clinical EEG data, including 15,833 epileptic spike waveforms recorded from 50 patients, and their labels were annotated by specialists. In the experiments, we compared three scenarios for the first layer: no preprocessing, discrete wavelet transform, and the proposed data-driven filters. The experimental results show that the trained data-driven filter bank with supervised learning behaves like multiple bandpass filters. In particular, the trained filter passed a frequency band of approximately 10-30 Hz. Moreover, the proposed method detected epileptic spikes, with the area under the receiver operating characteristic curve of 0.967 in the mean of 50 intersubject validations.

neuroscience↗

bioRxiv2020

High-resolution Crystal Structures of Transient Intermediates in the Phytochrome Photocycle

Phytochromes are red/far-red light photoreceptors in bacteria to plants, which elicit a variety of important physiological responses. They display a reversible photocycle between the resting (dark) Pr state and the light activated Pfr state, in which light signals are received and transduced as structural change through the entire protein to modulate the activity of the protein. It is unknown how the Pr-to-Pfr interconversion occurs as the structure of intermediates remain notoriously elusive. Here, we present short-lived crystal structures of the classical phytochrome from myxobacterium Stigmatella aurantiaca captured by an X-ray Free Electron Laser 5 ns and 33ms after light illumination of the Pr state. We observe large structural displacements of the covalently bound bilin chromophore, which trigger a bifurcated signaling pathway. The snapshots show with atomic precision how the signal progresses from the chromophore towards the output domains, explaining how plants, bacteria and fungi sense red light.

biophysics↗

bioRxiv2020

Structural basis for no spectral shift of heliorhodopsin by counterion mutation

Microbial rhodopsins comprise an opsin protein with seven transmembrane helices and a retinal as the chromophore. An all-trans retinal is covalently-bonded to a lysine residue through the retinal Schiff base (RSB) and stabilized by a negatively-charged counterion. The distance between the RSB and counterion is closely related to the light energy absorption. However, in heliorhodopsin-48C12 (HeR-48C12), while Glu107 acts as the counterion, E107D mutation exhibits an identical absorption spectrum to the wild-type, suggesting that the distance does not affect its absorption spectra. Here we present the 2.6 [A] resolution crystal structure of the Thermoplasmatales archaeon HeR E108D mutant, which also has an identical absorption spectrum to the wild-type. The structure revealed that D108 does not form a hydrogen bond with the RSB, and its counterion interaction becomes weaker. Alternatively, serine cluster, S78, S112, and S238 form a distinct interaction network around the RSB. The absorption spectra of the E to D and S to A double mutants suggested that S112 influences the spectral shift by compensating for the weaker counterion interaction. Our structural and spectral studies have revealed the unique spectral shift mechanism of HeR and clarified the physicochemical properties of HeRs.

biochemistry↗

bioRxiv2020

Induction of Alzheimer's disease pathology by early-life stress

BackgroundAlzheimers disease (AD), a progressive neurodegenerative disorder, is a serious social problem. Recently, several early-life factors have been associated with an increased risk of a clinical diagnosis of AD. MethodsWe investigated the involvement of early-life stress in AD pathogenesis using heterozygous the amyloid precursor protein (APP) mutant mice (AppNL-G-F/wt) and wild-type (Appwt/wt) mice. Maternal separation was used as an animal paradigm for early-life stress. Object location and fear conditioning tests were performed to measure cognitive functions, in addition to biochemical tests. Immunohistochemical analyses were performed after the behavioral tests. ResultsWe found that maternal-separated Appwt/wt mice showed narrowing of vessels and decreased pericyte coverage of capillaries in prefrontal cortex, while maternal-separated AppNL-G-F/wt mice additionally showed impairment of cognitive function, and earlier formation of A{beta} plaques and disruption of the blood-brain barrier. Severe activation of microglia was detected in the maternal-separated AppNL-G-F/wt mice and maternal-separated Appwt/wt mice. At the early stage, morphological changes and inflammatory responses were observed in the microglia of the maternal-separated AppNL-G-F/wt mice and maternal-separated Appwt/wt mice, as well as morphological changes in the microglia of the non-maternal-separated AppNL-G-F/wt mice. ConclusionsMicroglia activation induced by maternal separation in combination with the APP mutation may impairs the vascular system, leading to AD progression. These findings therefore suggest that maternal separation causes early induction of AD pathology via angiopathy.

neuroscience↗

bioRxiv2020

Multidrug treatment with nelfinavir and cepharanthine against COVID-19

Antiviral treatments targeting the emerging coronavirus disease 2019 (COVID-19) are urgently required. We screened a panel of already-approved drugs in a cell culture model of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and identified two new antiviral agents: the HIV protease inhibitor Nelfinavir and the anti-inflammatory drug Cepharanthine. In silico modeling shows Nelfinavir binds the SARS-CoV-2 main protease consistent with its inhibition of viral replication, whilst Cepharanthine inhibits viral attachment and entry into cells. Consistent with their different modes of action, in vitro assays highlight a synergistic effect of this combined treatment to limit SARS-CoV-2 proliferation. Mathematical modeling in vitro antiviral activity coupled with the known pharmacokinetics for these drugs predicts that Nelfinavir will facilitate viral clearance. Combining Nelfinavir/Cepharanthine enhanced their predicted efficacy to control viral proliferation, to ameliorate both the progression of disease and risk of transmission. In summary, this study identifies a new multidrug combination treatment for COVID-19.

microbiology↗

bioRxiv2020

Multiband entropy-based feature-extraction method for automatic identification of epileptic focus based on high- frequency components in interictal iEEG

Presurgical investigations for categorizing focal patterns are crucial, leading to localization and surgical removal of the epileptic focus. This paper presents a machine learning approach using information theoretic features extracted from high-frequency subbands to detect the epileptic focus from interictal intracranial electroencephalogram (iEEG). It is known that high-frequency subbands (>80 Hz) include important biomarkers such as high-frequency oscillations (HFOs) for identifying epileptic focus commonly referred to as the seizure on-set zone (SOZ). In this analysis, the multi-channel interictal iEEG signals were splitted into segments and each segment was decomposed into multiple high-frequency subbands. The different types of entropy were calculated for each of the subbands and the sparse linear discriminant analysis (sLDA) was applied to select the prominent entropy features. Due to the imbalance of SOZ and non-SOZ channels in iEEG data, the use of machine learning techniques is always tricky. To deal with the imbalanced learning problem, an adaptive synthetic oversampling approach (ADASYN) with radial basis function kernel-based SVM was used to detect the focal segments. Finally, the epileptic focus was identified based on detection of focal segments on SOZ and non-SOZ channels. Eight patients were examined to observe the efficiency of the automatic detector. The experimental results and statistical tests indicate that the proposed automatic detector can identify the epileptic focus accurately and efficiently.

bioengineering↗

bioRxiv2020

Selective and controlled myelin formation by individual interfascicular oligodendrocytes in the mouse corpus callosum

Single oligodendrocytes produce myelin sheath around multiple axons in the central nervous system. Interfascicular oligodendrocytes (IOs) in the white matter are aligned in rows and facilitate nerve conduction, but their detailed morphologies remain largely unknown. In the present study, we three-dimensionally reconstructed seven IOs in a row in the murine corpus callosum using serial block face-scanning electron microscopy (SBF-SEM). These morphologically polarized IOs extended a thick process with numerous branches from the cytoplasm-rich part of the cell and formed myelin sheaths preferentially around distant axons. The multiple branched processes of each IO myelinated multiple axons having similar diameters with restricted myelin thicknesses, indicating that individual IOs have their own myelination profiles even on distinct target neurons. Consistent with the finding, the IOs transduced and visualized with the rabies viral vector expressing GFP showed statistically significant variations in the myelination patterns. We further reconstructed the sheath immediately adjacent to those derived from the seven IOs; the thicknesses of both sheaths were significantly correlated despite emanating from different IOs. These results proposed a rule that myelination by individual IOs, regulated by interaction with ensheathed axons, is selective in specific axons and at the same time orchestrated at the whole cell level. Main pointsCallosal oligodendrocytes form myelin sheaths preferentially around distant axons. Each oligodendrocyte myelinate multiple axons having similar diameters with restricted myelin thicknesses. The thicknesses of adjacent myelin sheaths are similar.

neuroscience↗

bioRxiv2020

Angiopathy with impaired glucose logistics observed in schizophrenia-like model mice

Metabolic dysfunction is thought to contribute to the severity of psychiatric disorders; however, it has been unclear whether current high-simple-sugar diets contribute to pathogenesis of these diseases. Here we demonstrate that a high-sucrose diet during adolescence induces psychosis-related behavioral endophenotypes, including hyperactivity, poor working memory, impaired sensory gating, and disrupted interneuron function in mice deficient for glyoxalase-1 (Glo1), an enzyme involved in detoxification of sucrose metabolites. Further, the high-sucrose diet induced microcapillary impairments and reduced brain glucose uptake in brains of Glo1 deficient mice. Aspirin protected against this angiopathy, enhancing brain glucose uptake, and preventing abnormal behavioral phenotypes. Similar vascular damage to our model mice was found in the brains of randomly collected schizophrenia and bipolar disorder patients, suggesting that psychiatric disorders are associated with angiopathy in the brain caused by various environmental stresses, including metabolic stress.

neuroscience↗

bioRxiv2020

Therapeutic Efficacy and Safety of Lenvatinib for Unresectable Hepatocellular Carcinoma Beyond Progression with Sorafenib

Background & AimsThe efficacy and safety of lenvatinib (LEN) as a second/third-line treatment for unresectable hepatocellular carcinoma (HCC) after sorafenib (SOR) therapy remains unknown. We evaluated the outcomes of second/third-line treatment of LEN, investigated the sensitivity of SOR-resistant HCC cell line (PLC/PRF5-R2) to LEN, and their signal transduction pathway by protein array analysis. MethodsWe retrospectively enrolled 57 unresectable HCC patients. Radiologic responses in 53 patients were evaluated by modified Response Evaluation Criteria in Solid Tumors. Active signal transduction pathways in cells were identified by protein array analysis, including 1205 proteins. ResultsPatients comprised 34 tyrosine kinase inhibitor (TKI)-naive (first-line), nine SOR-intolerant (second-line), and ten resistant to regorafenib (third-line). Objective response rates (ORRs) were 61.8% (21/34) in TKI-naive, 33.3% (3/9) in second-line, and 20.0% (2/10) in third-line groups. The overall survival (OS) and the progression free survival (PFS) in the first-line was significantly longer than those in third-line group (p<0.05). Patients with better liver functional reserve (Child score, ALBI grade) exhibited higher ORR and longer OS. LEN was well-tolerated as second/third-line treatment. The IC50 value of LEN against PLC/PRF5-R2 cells (30 M) was significantly higher than that against PLC/PRF5 cells (6.4 M). LEN inhibited significantly more signal transduction pathways related to FRS2, a crucial FGFR downstream molecule, in PLC/PRF5 than PLC/PRF5-R2 cells. ConclusionsLEN was active and safe as a second/third-line treatment for unresectable HCC. LEN seems to be more effective for HCC patients with better hepatic reserve function or before TKI-resistance is acquired because of partial cross-resistance to SOR.

molecular biology↗

bioRxiv2019

Large scale genome-wide association study in a Japanese population identified 45 novel susceptibility loci for 22 diseases

INTRODUCTORY PARAGRAPHThe overwhelming majority of participants in current genetic studies are of European ancestry1-3, limiting our genetic understanding of complex disease in non-European populations. To address this, we aimed to elucidate polygenic disease biology in the East Asian population by conducting a genome-wide association study (GWAS) with 212,453 Japanese individuals across 42 diseases. We detected 383 independent signals in 331 loci for 30 diseases, among which 45 loci were novel (P < 5 x 10-8). Compared with known variants, novel variants have lower frequency in European populations but comparable frequency in East Asian populations, suggesting the advantage of this study in discovering these novel variants. Three novel signals were in linkage disequilibrium (r2 > 0.6) with missense variants which are monomorphic in European populations (1000 Genomes Project) including rs11235604(p.R220W of ATG16L2, a autophagy-related gene) associated with coronary artery disease. We further investigated enrichment of heritability within 2,868 annotations of genome-wide transcription factor occupancy, andidentified 378 significant enrichments across nine diseases (FDR < 0.05) (e.g. NF-{kappa}B for immune-related diseases). This large-scale GWAS in a Japanese population provides insights into the etiology of common complex diseases and highlights the importance of performing GWAS in non-European populations.

genetics↗

bioRxiv2019

The primary structural photoresponse of phytochrome proteins captured by a femtosecond X-ray laser

Phytochrome proteins control the growth, reproduction, and photosynthesis of plants, fungi, and bacteria. Light is detected by a bilin cofactor, but it remains elusive how this leads to activation of the protein through structural changes. We present serial femtosecond X-ray crystallographic data of the chromophore-binding domains of a bacterial phytochrome at delay times of 1 ps and 10 ps after photoexcitation. The structures reveal a twist of the D-ring, which lead to partial detachment of the chromophore from the protein. Unexpectedly, the conserved so-called pyrrole water is photodissociated from the chromophore, concomitant with movement of the A-ring and a key signalling aspartate. The changes are wired together by ultrafast backbone and water movements around the chromophore, channeling them into signal transduction towards the output domains. We suggest that the water dissociation is key to the phytochrome photoresponse, explaining the earliest steps of how plants, fungi and bacteria sense red light.

biophysics↗

bioRxiv2019

Comparing serial X-ray crystallography and microcrystal electron diffraction (MicroED) as methods for routine structure determination from small macromolecular crystals

Innovative new crystallographic methods are facilitating structural studies from ever smaller crystals of biological macromolecules. In particular, serial X-ray crystallography and microcrystal electron diffraction (MicroED) have emerged as useful methods for obtaining structural information from crystals on the nanometer to micron scale. Despite the utility of these methods, their implementation can often be difficult, as they present many challenges not encountered in traditional macromolecular crystallography experiments. Here, we describe XFEL serial crystallography experiments and MicroED experiments using batch-grown microcrystals of the enzyme cyclophilin A (CypA). Our results provide a roadmap for researchers hoping to design macromolecular microcrystallography experiments, and they highlight the strengths and weaknesses of the two methods. Specifically, we focus on how the different physical conditions imposed by the sample preparation and delivery methods required for each type of experiment effect the crystal structure of the enzyme.

biophysics↗

bioRxiv2019

Inverse correlation between heme synthesis and the Warburg effect in cancer cells

AbstractsCancer cells show a bias toward the glycolytic system over the conventional mitochondrial electron transfer system for obtaining energy. This biased metabolic adaptation is called the Warburg effect. Cancer cells also exhibit a characteristic metabolism, a decreased heme synthesizing ability. Here we show that heme synthesis and the Warburg effect are inversely correlated. We used human gastric cancer cell lines to investigate glycolytic metabolism and electron transfer system toward promotion/inhibition of heme synthesis. Under hypoxic conditions, heme synthesis was suppressed and the glycolytic system was enhanced. Addition of a heme precursor for the promotion of heme synthesis led to an enhanced electron transfer system and inhibited the glycolytic system and vice versa. Enhanced heme synthesis leads to suppression of cancer cell proliferation by increasing intracellular reactive oxygen species levels. Collectively, the promotion of heme synthesis in cancer cells eliminated the Warburg effect by shifting energy metabolism from glycolysis to oxidative phosphorylation.

biochemistry↗