Biology subjects

Tanigawa, T.

Publications and source records attributed to Tanigawa, T..

4 recordsLinked to original sources

bioRxiv2024

High-Recovery Desalting Tip Columns for a Wide Variety of Peptides in Mass Spectrometry-based Proteomics

In mass spectrometry-based proteomics, loss-minimized peptide purification techniques play a key role in improving sensitivity and coverage. We have developed a desalting tip column packed with thermoplastic polymer-coated chromatographic particles, named ChocoTip, to achieve high recoveries in peptide purification by pipette-tip-based LC with centrifugation (tipLC). ChocoTip identified more than twice as many peptides from 20 ng of tryptic peptides from Hela cell lysate compared to a typical StageTip packed with chromatographic particles entangled in a Teflon mesh in tipLC. The high recovery of ChocoTip in tipLC was maintained for peptides with a wide variety of physical properties over the entire retention time range of the LC/MS/MS analysis, and was especially noteworthy for peptides with long retention times. These excellent properties are attributable to the unique morphology of ChocoTip, in which the thermoplastic polymer covers the pores, thereby inhibiting irreversible adsorption of peptides into mesopores of the chromatographic particles. ChocoTip is expected to find applications, especially in clinical proteomics and single-cell proteomics, where sample amounts are limited.

biochemistry↗

bioRxiv2022

Wsc1 acts as a piezosensor in Saccharomyces cerevisiae, enhancing glycerol efflux via aquaglyceroporin Fps1 in response to high hydrostatic pressure

The fungal cell wall is the initial barrier for the fungi against diverse external stresses, such as osmolarity changes, harmful drugs, and mechanical injuries. This study explores the roles of osmoregulation and the cell wall integrity (CWI) pathway in response to high hydrostatic pressure in the yeast Saccharomyces cerevisiae. We demonstrate the roles of the transmembrane mechanosensor Wsc1 and aquaglyceroporin Fps1 in a general mechanism to maintain cell growth under high-pressure regimes. The promotion of water influx into cells at 25 MPa, as evident by an increase in cell volume and a loss of the plasma membrane eisosome structure, promotes the activation of Wsc1, an activator of the CWI pathway. The downstream mitogen-activated protein kinase Slt2 was hyperphosphorylated at 25 MPa. Glycerol efflux increases via Fps1 phosphorylation, which is initiated by downstream components of the CWI pathway and contributes to the reduction in intracellular osmolarity under high pressure. The elucidation of the mechanisms underlying adaption to high pressure through the well-established CWI pathway could potentially translate to mammalian cells and provide novel insights into cellular mechanosensation.

cell biology↗

bioRxiv2022

Classification of Extracellular Vesicles based on Surface Glycan Structures by Spongy-like Separation Media

Extracellular vesicles (EVs) are lipid bilayer vesicles that enclose various biomolecules. EVs hold promise as sensitive biomarkers to detect and monitor various diseases. However, they have heterogenous molecular compositions. The compositions of EVs from identical donor cells obtained using the same purification methods may differ, which is a significant obstacle for elucidating objective biological functions. Herein the potential of a novel lectin-based affinity chromatography (LAC) method to classify EVs based on their glycan structures is demonstrated. The proposed method utilizes a spongy-like monolithic polymer (spongy monolith, SPM), which consists of poly(ethylene-co-glycidyl methacrylate) with continuous micropores and allows an efficient in-situ protein reaction with epoxy groups. Two distinct lectins with different specificities, Sambucus sieboldiana agglutinin and concanavalin A, are effectively immobilized on SPM without impacting the binding activity. Moreover, high recovery rates of liposomal nanoparticles as a model of EVs are achieved due to the large flow-through pores (>10 m) of SPM. Finally, lectin-immobilized SPMs are employed to classify EVs based on the surface glycan structures and demonstrate different subpopulations by proteome profiling.

biochemistry↗

bioRxiv2018

Changes in clock gene expression in esophagus in rat reflux esophagitis

Background and AimsGastroesophageal reflux disease (GERD) is strongly associated with sleep disturbances. Clock genes harmonize circadian rhythms by their periodic expression and regulate several physiological functions. However, the association between clock genes and GERD is still unknown. We investigated whether reflux esophagitis affects circadian variability of clock genes in the esophagus and other organs using a rat reflux esophagitis model.\n\nMethodsReflux esophagitis was induced in 7-week-old male Wistar rats. Sham-operated rats were used as controls. Rats were sacrificed at 09:00 (light period) and 21:00 (dark period) 3 days (acute phase) and 21 days (chronic phase) after induction of esophagitis. The expression levels of clock gene mRNAs such as Per1, Per2, Per3, Cryl, Cry2, Arntl, and Clock in the esophagus were investigated by qPCR. Arntl expression was examined in stomach, small intestine, colon, and liver tissues. Serum melatonin and IL-6 levels were measured by ELISA.\n\nResultsHistological examination of reflux esophagitis mainly revealed epithelial defects with marked inflammatory cell infiltration in the acute phase, and mucosal thickening with basal cell hyperplasia in the chronic phase. Circadian variability of clock genes, except Cry1, was present in the normal esophagus, and was completely disrupted in reflux esophagitis during the acute phase. The circadian variability of Per2, Per3, and Arntl returned to normal, but disruption of Per1, Cry2, and Clock was present in the chronic phase. Disruption of circadian variability of Arntl was observed in the esophagus, as well as in the stomach, small intestine, and liver tissues in reflux esophagitis during the acute phase. There were no significant differences in serum melatonin and IL-6 levels between control and reflux esophagitis animals in both acute and chronic phases.\n\nConclusionDisruption to circadian variability of clock genes may play a role in the pathogenesis of GERD.

pathology↗