Biology subjects

Morioka, Y.

Publications and source records attributed to Morioka, Y..

6 recordsLinked to original sources

bioRxiv2023

Akaluc bioluminescence offers superior sensitivity to track in vivo dynamics of SARS-CoV-2 infection

Monitoring in vivo viral dynamics can improve our understanding of pathogenicity and tissue tropism. For positive-sense, single-stranded RNA viruses, several studies have attempted to monitor viral kinetics in vivo using reporter genomes. The application of such recombinant viruses can be limited by challenges in accommodating bioluminescent reporter genes in the viral genome. Conventional luminescence also exhibits relatively low tissue permeability and thus less sensitivity for visualization in vivo. Here we show that unlike NanoLuc bioluminescence, the improved method, termed AkaBLI, allows visualization of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in Syrian hamsters. By successfully incorporating a codon-optimized Akaluc luciferase gene into the SARS-CoV-2 genome, we visualized in vivo infection, including the tissue-specific differences associated with particular variants. Additionally, we could evaluate the efficacy of neutralizing antibodies and mRNA vaccination by monitoring changes in Akaluc signals. Overall, AkaBLI is an effective technology for monitoring viral dynamics in live animals.

microbiology↗

bioRxiv2022

Virological characteristics of SARS-CoV-2 BA.2 variant

Soon after the emergence and global spread of a new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron lineage, BA.1 (ref1, 2), another Omicron lineage, BA.2, has initiated outcompeting BA.1. Statistical analysis shows that the effective reproduction number of BA.2 is 1.4-fold higher than that of BA.1. Neutralisation experiments show that the vaccine-induced humoral immunity fails to function against BA.2 like BA.1, and notably, the antigenicity of BA.2 is different from BA.1. Cell culture experiments show that BA.2 is more replicative in human nasal epithelial cells and more fusogenic than BA.1. Furthermore, infection experiments using hamsters show that BA.2 is more pathogenic than BA.1. Our multiscale investigations suggest that the risk of BA.2 for global health is potentially higher than that of BA.1.

microbiology↗

bioRxiv2020

Establishment of a reverse genetics system for SARS-CoV-2 using circular polymerase extension reaction

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been identified as the causative agent of coronavirus disease 2019 (COVID-19). While the development of specific treatments and a vaccine is urgently needed, functional analyses of SARS-CoV-2 have been limited by the lack of convenient mutagenesis methods. In this study, we established a PCR-based, bacterium-free method to generate SARS-CoV-2 infectious clones. Recombinant SARS-CoV-2 could be rescued at high titer with high accuracy after assembling 10 SARS-CoV-2 cDNA fragments by circular polymerase extension reaction (CPER) and transfection of the resulting circular genome into susceptible cells. Notably, the construction of infectious clones for reporter viruses and mutant viruses could be completed in two simple steps: introduction of reporter genes or mutations into the desirable DNA fragments (~5,000 base pairs) by PCR and assembly of the DNA fragments by CPER. We hope that our reverse genetics system will contribute to the further understanding of SARS-CoV-2.

microbiology↗

bioRxiv2020

A synthetic synaptic organizer protein restores glutamatergic neuronal circuits

Neuronal synapses undergo structural and functional changes throughout life, essential for nervous system physiology. However, these changes may also perturb the excitatory/inhibitory neurotransmission balance and trigger neuropsychiatric and neurological disorders. Molecular tools to restore this balance are highly desirable. Here, we report the design and characterization of CPTX, a synthetic synaptic organizer combining structural elements from cerebellin-1 and neuronal pentraxin-1 to interact with presynaptic neurexins and postsynaptic AMPA-type ionotropic glutamate receptors. CPTX induced the formation of excitatory synapses in vitro and in vivo and restored synaptic functions, motor coordination, spatial and contextual memories, and locomotion in mouse models for cerebellar ataxia, Alzheimers disease and spinal cord injury, respectively. Thus, CPTX represents a prototype for novel structure-guided biologics that can efficiently repair or remodel neuronal circuits. One Sentence SummaryStructural biology information was used to design CPTX, a synthetic protein that induces functional excitatory synapses and restores normal behaviors in mouse models of neurological diseases.

neuroscience↗

bioRxiv2020

Various miRNAs are involved in efficient HCV replication

One of the determinants for tissue tropism of hepatitis C virus (HCV) is miR-122, a liver-specific microRNA. Recently, it has been reported that interaction of miR-122 to HCV RNA induces a conformational change of the 5UTR internal ribosome entry site (IRES) structure to form stem-loop II structure (SLII) and hijack of translating 80S ribosome through the binding of SLIII to 40S subunit, which leads to efficient translation. On the other hand, low levels of HCV-RNA replication have also been detected in some non-hepatic cells; however, the details of extrahepatic replication remain unknown. These observations suggest the possibility that miRNAs other than miR-122 can support efficient replication of HCV-RNA in non-hepatic cells. Here, we identified a number of such miRNAs and show that they could be divided into two groups: those that bind HCV-RNA at two locations (miR-122 binding sites I and II), in a manner similar to miR-122 (miR-122-like), and those that target a single site that bridges sites I and II and masking both G28 and C29 in the 5UTR (non-miR-122-like). Although the enhancing activity of these non-hepatic miRNAs were lower than those of miR-122, substantial expression was detected in various normal tissues. Furthermore, structural modeling indicated that both miR-122-like and non-miR-122-like miRNAs not only can facilitate the formation of an HCV IRES SLII but also can stabilize IRES 3D structure in order to facilitate binding of SLIII to the ribosome. Together, these results suggest that HCV facilitates miR-122-independent replication in non-hepatic cells through recruitment of miRNAs other than miR-122. And our findings can provide a more detailed mechanism of miR-122-dependent enhancement of HCV-RNA translation by focusing on IRES tertiary structure. Author summaryOne of the determinants for tissue tropism of hepatitis C virus (HCV) is miR-122, a liver-specific microRNA, which is required for efficient propagation. Recently, it has been reported that interaction of miR-122 with the 5UTR of HCV contributes to the folding of a functional IRES structure that is required for efficient translation of viral RNA. In this study, we examined the minimum motifs in the seed region of miRNAs required for the enhancement of HCV replication. As a result, we found two groups of non-hepatic miRNAs: "miR-122-like miRNAs" that can bind HCV-RNA at two locations in a manner similar to miR-122, and "non-miR-122-like miRNAs" that target a single site that masking both G28 and C29 in the 5UTR. The interaction of these non-hepatic miRNAs with the 5UTR can facilitate not only the folding of active HCV IRES but also the stabilization of IRES 3D structure in order to facilitate binding to the ribosome. These results suggest the possibility of replication of HCV in non-hepatic cells through interaction with miRNAs other than miR-122 and provide insight into the establishment of persistent infection of HCV in non-hepatic tissues that lead to the development of extrahepatic manifestations.

microbiology↗

bioRxiv2017

Anterior cingulate cortex connectivity is associated with suppression of behavior in a rat model of chronic pain

A cardinal feature of persistent pain that follows injury is a general suppression of behavior, in which motivation is inhibited in a way that promotes energy conservation and recuperation. Across species, the anterior cingulate cortex (ACC) is associated with the motivational aspects of phasic pain, but whether it mediates motivational functions in persistent pain is less clear. Using burrowing behavior as an marker of non-specific motivated behavior in rodents, we studied the suppression of burrowing following painful CFA or control injection into the right knee-joint of 37 rats (18 with pain), and examined associated neural connectivity with ultra-high-field resting state functional MRI. We found that connectivity between ACC and the subcortex correlated with the reduction in burrowing behavior observed following the pain manipulation. In a full replication study we confirmed these findings in a group of 44 rats (23 with pain). Across both datasets, reduced burrowing was assosciated with increased connectivity between ACC and subcortical structures including hypothalamic/preoptic nuclei and the bed nucleus of the stria terminalis. Together, the findings implicate ACC connectivity as a robust correlate of the motivational aspect of persistent pain in rodents.

neuroscience↗