Biology subjects

Hatano, Y.

Publications and source records attributed to Hatano, Y..

6 recordsLinked to original sources

bioRxiv2025

Androgen receptors expressed in the primary sensory neurons regulate mechanical pain sensitivity

The expression of hormonal receptors in pain-processing regions complicates understanding the hormonal effects on pain mechanisms. This study investigates androgen receptor (AR) involvement in pain sensitivity and sex differences in pain perception. Mechanical pain thresholds were higher in normal male mice compared to gonadectomized (GDX) male and normal female mice, correlating with serum testosterone levels. In the dorsal root ganglia (DRG), AR was expressed in normal males but undetectable in GDX males and normal females. In male sensory neuron-selective AR conditional knockout (AR-cKO) mice, mechanical pain thresholds were significantly lower than in wild-type males. In female mice, administration of testosterone propionate or dihydrotestosterone significantly raised mechanical pain thresholds, accompanied by increased AR expression in the DRG. This effect was abolished in AR-cKO females, consistent with male findings. These results indicate that primary sensory neurons are critical targets of androgen signaling in regulating mechanical pain sensitivity.

neuroscience↗

bioRxiv2023

Sexually dimorphic effects of pexidartinib on nerve injury-induced neuropathic pain in mice

BackgroundIt is well-established that spinal microglia and peripheral macrophages play critical roles in the etiology of neuropathic pain; however, growing evidence suggests sex differences in pain hypersensitivity owing to microglia and macrophages. Therefore, it is crucial to understand sex- and androgen-dependent characteristics of pain-related myeloid cells in mice with nerve injury-induced neuropathic pain. MethodsThe current study was performed using normal male and female mice, as well as gonadectomized (GDX) male mice. To deplete microglia and macrophages, pexidartinib (PLX3397), an inhibitor of the colony-stimulating factor 1 receptor, was orally administered, and mice were subjected to partial sciatic nerve ligation (PSL). Immunohistochemistry was performed to visualize microglia and macrophages, and PSL-induced mechanical allodynia was evaluated using the von Frey test. ResultsFollowing PSL induction, healthy male and female mice and male GDX mice exhibited similar levels of spinal microglial activation, peripheral macrophage accumulation, and mechanical allodynia. Treatment with PLX3397 significantly suppressed mechanical allodynia in normal males; this was not observed in female and GDX male mice. Sex- and androgen-dependent differences in the PLX3397-mediated preventive effects were observed on spinal microglia and dorsal root ganglia (DRG) macrophages, as well as in expression patterns of pain-related inflammatory mediators in these cells. Conversely, no sex- or androgen-dependent differences were detected in sciatic nerve macrophages, and inhibition of peripheral CC-chemokine receptor 5 prevented neuropathic pain in both sexes. ConclusionCollectively, these findings demonstrate the presence of considerable sex- and androgen-dependent differences in the etiology of neuropathic pain in spinal microglia and DRG macrophages but not in sciatic nerve macrophages. Given that the mechanisms of neuropathic pain may differ among experimental models and clinical conditions, accumulating several lines of evidence is crucial to comprehensively clarifying the sex-dependent regulatory mechanisms of pain.

pharmacology and toxicology↗

bioRxiv2022

Accuracy of a machine learning method based on structural and locational information from AlphaFold2 for predicting the pathogenicity of TARDBP and FUS gene variants in ALS

BackgroundIn the sporadic form of amyotrophic lateral sclerosis (ALS), the pathogenicity of rare variants in the causative genes characterizing the familial form remains largely unknown. To predict the pathogenicity of such variants, in silico analysis is commonly used. In some cases of ALS, the gene mutations are concentrated in specific regions, and the resulting alterations in protein structure are thought to significantly affect pathogenicity. However, existing methods have not taken this issue into account. To address this, we have developed a technique termed MOVA (method for evaluating the pathogenicity of missense variants using AlphaFold2), which applies positional information for structural variants predicted by AlphaFold2. Here we examined the utility of MOVA for analysis of several causative genes of ALS. MethodsWe analyzed variants of six ALS-related genes (TARDBP, FUS, SETX, TBK1, OPTN, and SOD1) and classified them as pathogenic or neutral. For each gene, the features of the variants, including their positions in the 3D structure predicted by AlphaFold2, were entered into a random forest algorithm and evaluated by leave-one-out cross-validation. We compared how accurately MOVA was able to classify the pathogenic and neutral mutation variants. ResultsMOVA yielded useful results (AUC [≥]0.70 for 3 (TARDBP 0.755, FUS 0.844, and SOD1 0.787) of the 6 genes) and was particularly useful for genes where pathogenic mutations were concentrated at specific sites (TARDBP, FUS). ConclusionsMOVA is useful for predicting the virulence of rare variants of ALS-causing genes in which mutations are concentrated at specific structural sites.

bioinformatics↗

bioRxiv2021

A chemogenetic platform for controlling plasma membrane signaling and synthetic signal oscillation

Chemogenetic methods that enable the rapid translocation of specific signaling proteins in living cells using small molecules are powerful tools for manipulating and interrogating intracellular signaling networks. However, existing techniques rely on chemically induced dimerization of two protein components and have certain limitations, such as a lack of reversibility, bioorthogonality, and usability. Here, by expanding our self-localizing ligand-induced protein translocation (SLIPT) approach, we have developed a versatile chemogenetic system for plasma membrane (PM)-targeted protein translocation. In this system, a novel engineered Escherichia coli dihydrofolate reductase in which a hexalysine (K6) sequence is inserted in a loop region (iK6DHFR) is used as a universal protein tag for PM-targeted SLIPT. Proteins of interest that are fused to the iK6DHFR tag can be specifically recruited from the cytoplasm to the PM within minutes by addition of a myristoyl-O_SCPLOWDC_SCPLOW-Cys-tethered trimethoprim ligand (mDcTMP). We demonstrated the broad applicability and robustness of this engineered protein-synthetic ligand pair as a tool for the conditional activation of various types of signaling molecules, including protein and lipid kinases, small GTPases, heterotrimeric G proteins, and second messengers. In combination with a competitor ligand and a culture-medium flow chamber, we further demonstrated the application of the system for chemically manipulating protein localization in a reversible and repeatable manner to generate synthetic signal oscillations in living cells. The present bioorthogonal iK6DHFR/mDcTMP-based SLIPT system affords rapid, reversible, and repeatable control of the PM recruitment of target proteins, offering a versatile and easy-to-use chemogenetic platform for chemical and synthetic biology applications.

synthetic biology↗

bioRxiv2021

Chromosome counting in the mouse and human zygote using low-invasive super-resolution live-cell imaging

In preimplantation embryos, an abnormal chromosome number causes developmental failure and a reduction in the pregnancy rate. Conventional chromosome testing methods requiring biopsy reduce the risk of associated genetic diseases; nevertheless, the reduction in cell number also reduces the pregnancy rate. Therefore, we attempted to count the chromosomes in mouse (Slc:ICR) embryos using super-resolution live-cell imaging as a new method of chromosome counting that does not reduce the cell number or viability. We counted the forty chromosomes at the first mitosis by injecting embryos with histone H2B-mCherry mRNA under conditions by which pups could be obtained; however, the results were often an underestimation of chromosome number and varied by embryo and time point. Therefore, we developed a method to count the chromosomes via CRISPR/dCas-mediated live-cell fluorescence in situ hybridization targeting the sequence of the centromere region, enabling us to count the chromosomes more accurately in mouse embryos. The methodology presented here may provide useful information for assisted reproductive technologies, such as those used in livestock animals/humans, as a technique for assessing the chromosomal integrity of embryos prior to transfer. Impact StatementLow-toxic super-resolution observation enables chromosome counting in preimplantation embryos without cell collection.

developmental biology↗

bioRxiv2020

Improved synthetic lipidation-based protein translocation system for SNAP-tag fusion proteins

The ability to artificially attach lipids to specific intracellular protein targets would be a valuable approach for controlling protein localization and function in cells. We recently devised a chemogenetic method in which a SNAP-tag fusion protein can be translocated from the cytoplasm to the plasma membrane by post-translationally and covalently conjugating a synthetic lipopeptide in cells. However, the first-generation system lacked general applicability. Herein, we present an improved synthetic lipidation system that enables efficient plasma membrane translocation of SNAP-tag fusion proteins in cells. This second-generation system is now applicable to the control of various cell-signaling molecules, offering a new and useful research tool in chemical biology and synthetic biology.

biochemistry↗