Biology subjects

Usui, K.

Publications and source records attributed to Usui, K..

6 recordsLinked to original sources

bioRxiv2026

Antiglycation effects of imidazole dipeptides and 2-oxo-imidazole dipeptides on glyceraldehyde-induced intracellular protein glycation and neuronal cell death

Glyceraldehyde (GA) contributes to the development of various diseases, such as diabetes and Alzheimers disease via protein glycation and the formation of advanced glycation end products (AGEs); however, effective strategies for neutralizing GA are limited. Carnosine (Car), an imidazole dipeptide (IDP) that is abundant in meat, suppresses protein glycation by scavenging reactive aldehydes. There are only a few reports on the antiglycation activity of Car against GA. For other IDPs, such as anserine, balenine (Bal), and homocarnosine, there are almost no reports on their antiglycation activity. In this study, we demonstrated the antiglycation activity of four types of IDPs and 2-oxocarnosine (2-oxo-Car), an oxidized form of Car, against GA-induced intracellular protein glycation and neuronal cytotoxicity. Car and Bal exhibited significantly higher reactivity with GA compared with other IDPs and 2-oxo-Car. An in silico analysis suggested that the difference in reactivity is dependent upon intramolecular hydrogen bond formation and the conformation of each IDP. Although there were differences in reactivity with GA, LC-MS analysis revealed that all of the IDPs and 2-oxo-Car reacted with two molecules of GA to form adducts containing pyridinium rings. Car and Bal exhibited high reactivity with GA and markedly suppressed GA-induced cytotoxicity in SH-SY5Y cells. Western blot and qPCR analyses revealed that IDPs suppressed GA-induced protein glycation and the upregulation of endoplasmic reticulum and oxidative stress response genes. Our results indicate that IDPs represent a novel preventive approach to AGE-related diseases and provide a foundation for the development of strategies to treat GA-related neurotoxicity. Graphical abstractCarnosine and balenine, which are imidazole dipeptides (IDPs), scavenged two molecules of glyceraldehyde to form adducts containing pyridinium rings and suppressed intracellular protein glycation and neuronal cell death. The formation of intramolecular hydrogen bond of IDPs played a crucial role in the strength of antiglycation activity. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=126 SRC="FIGDIR/small/734660v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@11d82dorg.highwire.dtl.DTLVardef@187aff0org.highwire.dtl.DTLVardef@12fe482org.highwire.dtl.DTLVardef@14327ee_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2026

Epigenomes in thermophilic microbial communities and their impact on the interaction across prokaryotes and mobilomes

DNA chemical modifications, including methylation, are widespread in prokaryotes and mobilomes, including viruses, plasmids, and other extrachromosomal DNAs, and play important roles in their ecology and interactions. However, current knowledge of these modification systems and their association with interactions between hosts and mobilomes across communities, including those in extreme environments, is severely limited. Here, using single-molecule real-time sequencing and single-cell genome sequencing technologies, we conducted a culture-independent metaepigenomic analysis of microbial communities in hot spring biofilms. A total of 248, 332, and 465 genomes were constructed from diverse prokaryotes, viruses, and extrachromosomal circular DNAs, respectively, from 10 biofilm samples collected from 3 hot spring sites. In total, 1106 candidate methylated motifs and 3280 genes associated with the restriction-modification (RM) system, including DNA methyltransferases (MTases), were identified. In contrast to the varied methylated motifs, the nucleotide-level modification ratios were consistent with those of a common Escherichia coli genome, and an environment-dependent epigenomic preference attributed to the lack of C5-methylcytosine was observed, as supported by direct measurements of modified bases by liquid chromatography-tandem mass spectrometry. A systematic survey revealed various defense systems in the genome, and almost half of the MTase genes were estimated to be genetically involved in defense mechanisms against extracellular DNA, such as RM systems. The mobilomes and their predicted hosts shared epigenomic patterns within each interactive subnetwork, suggesting that mobilome DNA was modified by host MTase during the current infection or transfection, rather than serving as historical records. Our findings highlight that DNA modification shapes multiple ecological and evolutionary strategies in interactions between prokaryotes and mobilomes, and that epigenomes serve as a potential signature for accurate prediction of current host-phage interactions.

microbiology↗

bioRxiv2026

Experimental SARS-CoV-2 infection using horseshoe bats

Horseshoe bats are known as the natural reservoir of sarbecoviruses. To understand how horseshoe bats coexist with sarbecoviruses in nature, experimental infection can provide direct evidence. However, in vivo infection studies using horseshoe bats have been lacking because of the difficulty of maintaining insectivorous bats in a laboratory setting. Here, we established a stable husbandry system for greater horseshoe bats (Rhinolophus ferrumequinum) and performed experimental infection with SARS-CoV-2. In contrast to Syrian hamsters which showed substantial viral replication, infected horseshoe bats exhibited low-level but persistent viral replication in the lung without overt disease. Histological analyses revealed that inflammatory lesions in the bat lungs were spatially restricted and temporally delayed compared with those in hamsters. Transcriptomic analyses further showed preferential activation of tissue repair pathways but limited inflammatory responses following infection. Notably, bats expressed several interferon-stimulated genes prior to infection. Our results suggest that a host strategy combining constitutive antiviral state, limited inflammation and enhanced tissue repair may result in controlled viral replication without overt disease, likely enabling horseshoe bats to coexist with sarbecoviruses.

microbiology↗

bioRxiv2024

Extracellular vesicle-mediated secretion of chlorophyll biosynthetic intermediates in the cyanobacterium Leptolyngbya boryana

Extracellular vesicles (EVs) are derived from the outer membrane (OM) in Gram-negative bacteria and have diverse physiological functions such as toxin transport, extracellular signal transduction, nutrient acquisition, and horizontal gene transfer. EV-mediated secretion of monovinyl protochlorophyllide (MV-Pchlide), the chlorophyll a (Chl) biosynthetic intermediate, was previously reported in a mutant lacking dark-operative Pchlide reductase in the cyanobacterium Leptolyngbya boryana. This study showed a detailed characterization of EVs from the wild-type (WT) of L. boryana grown under photoautotrophic and dark heterotrophic conditions, focusing on the accumulation of Chl intermediates. WT L. boryana cells produce two types of EVs, low-density EVs (L-EVs) and high-density EVs (H-EVs), both under light and dark conditions. L-EVs and H-EVs showed distinct morphological features and protein compositions. L-EVs from cells grown under both light and dark conditions commonly contained carotenoids, myxol glycoside, and zeaxanthin, as major pigments. Based on the protein compositions of EVs and other cellular membrane fractions, L-EVs and H-EVs are probably derived from low-density OM and high-density OM interacting with cell walls, respectively. Fluorescence detection of pigments was applied to EVs, and the three Chl intermediates, protoporphyrin IX, demetallated Mg-protoporphyrin IX monomethyl ester, pheophorbide, and were commonly detected both L-EVs from light- and dark-grown cells, whereas L-EVs from dark-grown cells contained additional MV-Pchlide and MV-protopheophorbide. These Chl intermediates appear to transfer from the thylakoid membrane to L-EVs via an unknown transport system. Cyanobacterial EVs may play a novel function in alleviating the accumulation of Chl intermediates in cells.

microbiology↗

bioRxiv2023

Genetic and functional diversity of beta-N-acetylgalactosamine residue-targeting glycosidases expanded by deep-sea metagenome

{beta}-N-Acetylgalactosamine-containing glycans play essential roles in several biological processes, including cell adhesion, signal transduction, and immune responses. {beta}-N-Acetylgalactosaminidases hydrolyze {beta}-N-acetylgalactosamine linkages of various glycoconjugates. However, their biological significance remains ambiguous, primarily because only one type of enzyme, exo-{beta}-N-acetylgalactosaminidases that specifically act on {beta}-N-acetylgalactosamine residues, has been documented so far. In this study, we identified three novel glycoside hydrolase families distributed among all three domains of life and characterized eight novel {beta}-N-acetylgalactosaminidases and {beta}-N-acetylhexosaminidase through sequence-based screening of deep-sea metagenomes and subsequent searching of public protein databases. Despite low sequence similarity, the crystal structures of these enzymes demonstrate that all enzymes share a prototype structure and diversify their substrate specificities (endo-, dual-endo/exo-, and exo-) through the accumulation of mutations and insertional amino acid sequences. The diverse {beta}-N-acetylgalactosaminidases reported in this study could facilitate the comprehension of their structures and functions and present novel evolutionary pathways for expanding their substrate specificity.

biochemistry↗

bioRxiv2020

Development of a Tet-On inducible expression system for the anhydrobiotic cell line, Pv11

The Pv11 cell line established from an African chironomid, Polypedilum vanderplanki, is the only cell line tolerant to complete desiccation. In Pv11 cells, a constitutive expression system for Pv11 cells was previously exploited and several reporter genes were successfully expressed. Here we report the identification of an effective minimal promoter for Pv11 cells and its application to the Tet-On inducible expression system. First, using a luciferase reporter assay, we showed that a 202 bp deletion fragment derived from the constitutively active 121-promoter functions in Pv11 cells as an appropriate minimal promoter with the Tet-On inducible expression system. The AcGFP1 protein was also successfully expressed in Pv11 cells using the inducible system. In addition to these reporter genes, avian myeloblastosis virus reverse transcriptase subunit (AMV RT), which is one of the most widely commercially available RNA-dependent DNA polymerases, was successfully expressed through the inducible expression system and its catalytic activity was verified. These results demonstrate the establishment of an inducible expression system in cells that can be preserved in the dry state and highlight a possible application to the production of large and complex proteins. HighlightsO_LIA 202 bp-deletion fragment derived from a constitutively active promoter was identified as a minimal promoter in Pv11 cells. C_LIO_LIA Tet-On inducible expression system was developed for Pv11 cells using the minimal promoter. C_LIO_LITypical reporter genes (GFP and luciferase) and an enzyme with complex structure, i.e. a viral reverse transcriptase, were successfully and inducibly expressed in Pv11 cells using the Tet-On system. C_LI

bioengineering↗