Biology subjects

Watabe, M.

Publications and source records attributed to Watabe, M..

7 recordsLinked to original sources

bioRxiv2026

Chemical Boosting of Foldase Condensates Accelerates Oxidative Protein Folding

Dynamic biomolecular condensates play crucial roles in intracellular compartmentalization and physiological functions. While engineering tools for compartmentalization have expanded add-on functionalities, directly amplifying the inherent catalytic machinery within biological phase-separated droplets has remained elusive. Herein, we developed a phase-separated oxidative folding reaction chamber based on protein disulfide isomerase A6 (PDIA6) by chemically targeting its active site CxxC motif to enhance enzymatic activity within PDIA6 droplets. A para-substituted N-methylated pyridinylmethanethiol (pMePySH) enhanced the catalytic oxidative folding of bovine pancreatic trypsin inhibitor, proinsulin, and antibody up to 12-fold within in vitro PDIA6 droplets. Furthermore, pMePySH targeted PDIA6 foci within the endoplasmic reticulum, significantly promoting insulin secretion. These findings offer a powerful platform for the spatiotemporal manipulation of protein folding, with profound implications for the scalable manufacturing of therapeutic antibodies and other complex biopharmaceuticals.

bioengineering↗

bioRxiv2025

Lipids Are Involved in Heterochromatin Condensation: A Quantitative Raman and Brillouin Microscopy Study

Chromatin, a fundamental component of eukaryotic genomes, is categorized into euchromatin and heterochromatin, which play distinct roles in gene regulation. Although these two chromatin states are distinguished by their degree of condensation, quantitatively measuring the degree of chromatin condensation, as well as the physical properties of chromatin in living cells, remains challenging. In this study, label-free in situ quantitative imaging was performed using a Raman-Brillouin microscope to visualize the spatial distribution of molecular concentration and viscoelasticity within the nuclear environment of a living cell. A quantitative concentration distribution image of each intracellular biomolecule was obtained by combining Raman imaging with multivariate curve resolution analysis, using a water Raman band as an internal standard. Simultaneous Raman-Brillouin imaging enables the quantitative visualization of viscoelasticity within a cell. Using this approach, we found that, in addition to DNA, heterochromatin is enriched in lipids and that lipids play a critical role in heterochromatin formation, determining its mechanical properties. These findings provide new insights into the mechanism of heterochromatin formation and its chemical and physical properties, leading to a comprehensive understanding of gene regulation and nuclear organization.

biophysics↗

bioRxiv2024

Ca2+-driven PDIA6 phase separation to ensure proinsulin quality control

The endoplasmic reticulum (ER) plays key roles in protein quality control1,2 and dynamic Ca2+ storage3,4 in eukaryotic cells. However, the protein homeostasis (proteostasis) system that regulates these ER functions is still incompletely characterised. Previous study revealed the importance of oligomerization in the function PDIA1, an ER-resident disulfide isomerase and molecular chaperone, regulates oligomeric states in accordance with client folding5. This result suggests that at least some of the 20 members of other PDI family may undergo regulated self-assembly in order to optimally function. Here, we show that Ca2+ triggers the phase separation of PDIA6 into liquid-like condensates. In contrast to the condensation mechanism observed for proteins containing low-complexity domains, our results indicate that transient but specific electrostatic interactions occur between the first and the third folded thioredoxin-like domains of PDIA6. We further show that the Ca2+-driven condensation of PDIA6 recruits PDIA3 and proinsulin, thus increasing their local concentrations. This process results in the 30-fold enhancement of proinsulin folding and in the inhibition of proinsulin aggregation. Our findings shed light on a condensation-driven Ca2+-mediated proteostasis cascade in the ER by revealing how the efficiency of the protein folding process can be enhanced within quality control granules.

biochemistry↗

bioRxiv2024

Conserved loop of a phase modifier endows protein condensates with fluidity

Dipeptide repeats (DPRs) that are gene products from abnormal hexanucleotide repeat expansion in C9orf72 trigger amyotrophic lateral sclerosis (ALS) through unknown mechanism. This study highlights, importin Karyopherin{beta}2 (Kap{beta}2), which is responsible for nuclear transport and phase modification of RNA-binding proteins (RBPs), as a major DPR target. We demonstrate DPR accumulation in the nucleus via Kap{beta}2-mediated transport, which results in dose-dependent toxicity observed in nematode and yeast models. In vitro interaction studies exploiting chemical probe arrays and biophysical measurements reveal multivalent DPR binding to Kap{beta}2, including at the conserved acidic loop. Refractive index and fluorescence imaging coupled with biochemical assays unveiled that binding of excess DPRs to the acidic loop turns a phase modifier Kap{beta}2 into phase disrupter, resulting more condensed and viscous RBP condensates. Our findings provides molecular insight into C9orf72-ALS related to age and repeat expansion.

biochemistry↗

bioRxiv2023

Zinc finger domains bind low-complexity domain polymers

Self-association of low-complexity protein sequences (LC domains) is important for polymer formation. Several molecular chaperones are involved in the regulation of LC domain polymer formation. However, the mechanisms underlying cell recognition of LC domain polymers remain unclear. Here we show that zinc finger domains (ZnFs) bind LC domains of RNA-binding proteins in a cross-{beta} polymer-dependent manner. ZnFs bound to LC domain hydrogels and suppressed LC domain polymer formation. Moreover, ZnFs preferentially recognize LC domains in the polymeric state. These findings suggest that ZnFs act as physiological regulators of LC domain polymer formation.

biochemistry↗

bioRxiv2023

Leveraging Machine Learning to Uncover the Hidden Links between Trusting Behavior and Biological Markers

Understanding the decision-making mechanisms underlying trust is essential, particularly for patients with mental disorders who experience difficulties in developing trust. We aimed to explore biomarkers associated with trust-based decision-making by quantitative analysis. However, quantification of decision-making properties is difficult because it cannot be directly observed. Here, we developed a machine learning method based on Bayesian hierarchical model to quantitatively decode the decision-making properties from behavioral data of a trust game. By applying the method to data of patients with MDD and healthy controls, we estimated model parameters regulating trusting decision-making. The estimated model was able to predict behaviors of each participant. Although there is no difference of the estimated parameters between MDD and healthy controls, several biomarkers were associated with the decision-making properties in trusting behavior. Our findings provide valuable insights into the trusting decision-making, offering a basis for developing targeted interventions to improve their social functioning and overall well-being.

bioinformatics↗

bioRxiv2020

A unique adhesive motif of protein disulfide isomerase P5 supports its function via dimerization

P5, also known as PDIA6, is a PDI-family member that plays an important role in the ER quality control. Herein, we revealed that P5 dimerizes via a unique adhesive motif contained in the N-terminal thioredoxin-like domain. This motif is apparently similar to, but radically different from conventional leucine-zipper motifs, in that the former includes a periodic repeat of leucine or valine residues at the third or fourth position spanning five helical turns on 15-residue anti-parallel -helices, unlike the latter of which the leucine residues appear every two helical turns on [~]30-residue parallel -helices at dimer interfaces. A monomeric P5 mutant with the impaired adhesive motif showed structural instability and local unfolding, and behaved as an aberrant protein that induces the ER stress response. Disassembly of P5 to monomers compromised its ability to inactivate IRE1 via reduction of intermolecular disulfide bonds and its Ca2+-dependent regulation of chaperone function in vitro. Thus, the leucine-valine adhesive motif supports structure and physiological function of P5.

biochemistry↗