Biology subjects

Takeuchi, A.

Publications and source records attributed to Takeuchi, A..

8 recordsLinked to original sources

bioRxiv2025

Significant improvement of environmental DNA assay by targeting retrotransposon sequences characteristic to Anguilla eels

Environmental DNA (eDNA) analysis faces challenges regarding sensitivity and quantification accuracy, particularly in areas with low target species densities or during seasons when eDNA release decreases. Multi-copy markers such as mitochondrial genomic DNA and ribosomal DNA (rDNA) have been widely used in eDNA analysis to address this issue. However, the copy number of the DNA markers per cell remains a potential bottleneck for eDNA sensitivity. In this study, we aimed to increase the sensitivity of eDNA assays by using retrotransposons, which are abundant in the genome, as novel target markers. We developed an assay targeting UnaSINE1, a short interspersed nucleotide element (SINE) characteristic of Anguilla eels, and compared its sensitivity and accuracy with that of an established mitochondrial 16S rRNA marker. Our results demonstrated that UnaSINE1 was detected at over 100 times the copy number of the mitochondrial marker in both genomic and eDNA samples. In the river surveys, the 16S marker was positive in 32 of the 81 samples, whereas the UnaSINE1 marker was positive in 62 samples, indicating that the use of SINE marker remarkably reduced false negatives. Furthermore, both biological and technical replicates exhibited improved positive consistency and reduced variability in quantification, leading to more robust presence/absence determination and quantitative results. Utilizing retrotransposon sequences as markers requires additional effort for sequence acquisition and organization and may limit taxonomic resolution to the genus level. However, this approach significantly improves sensitivity without increasing the labor or cost of sampling and PCR analysis, making it highly practical for eDNA studies.

ecology↗

bioRxiv2024

Viral vectors with cluster of differentiation gene promoters to target specific cell types in the brain

Understanding brain function and developing targeted therapies for neurological disorders require precise access to specific cell types, but current methods are limited. Here, we report the development of cell-type specific adeno-associated virus (AAV) vectors utilizing promoters of cluster of differentiation genes (CD promoters) for targeted gene delivery in the brain. We newly identified CD promoters that induce gene expression selectively in specific cell types in the mouse brain. These AAVs enable in vivo calcium imaging and chemogenetic applications in specific cerebellar cells, revealing distinct and shared roles of two types of cerebellar interneurons. Notably, chemogenetic modulation of cerebellar molecular layer interneurons using a CD promoter rescued social behavior and motor deficits in a mouse model of autism spectrum disorder. Our findings demonstrate the utility of these AAVs in elucidating the functions of individual cell types in brain function and in developing novel, cell-type specific therapies for brain diseases.

neuroscience↗

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↗

bioRxiv2023

Direct photoreception of a pituitary endocrine cell, melanotroph, induces a hormone release

In addition to canonical photoreception by the eye, many other organs express non-visual photoreceptors although their biological significance is mostly unknown. Here, we discovered a novel phenomenon in which the pituitary of medaka directly receives light, which induces hormone release. Ca2+ imaging analysis revealed that a melanotroph, a pituitary endocrine cell secreting melanocyte-stimulating hormone (MSH), robustly increases [Ca2+]i during short-wavelength light irradiation. Moreover, we identified Opn5m as the key molecule of this mechanism. The significance of this phenomenon was suggested to be involved in UV protection because knockout of opn5m significantly reduced the expression of tyrosinase, the rate-limiting enzyme for melanogenesis, in the skin. These results suggest a novel mechanism in which direct reception of short-wavelength light by pituitary endocrine cells triggers the pathway to enhance UV protection. One-Sentence SummaryAn endocrine cell of the pituitary was proven to be a photoreceptive cell that enables autonomous hormone release.

physiology↗

bioRxiv2023

Structure and function of a YeeE-YeeD complex for sophisticated thiosulfate uptake

Uptake of thiosulfate ions as an inorganic sulfur source from the environment is important for bacterial sulfur assimilation. Recently, a selective thiosulfate uptake pathway involving a membrane protein YeeE (TsuA) in Escherichia coli was characterized. YeeE-like proteins are conserved in some bacteria, archaea, and eukaryotes. However, the precise function of YeeE, along with its potential partner protein in the thiosulfate ion uptake pathway, remained unclear. Here, we assessed selective thiosulfate transport via Spirochaeta thermophila YeeE in vitro and characterized E. coli YeeD (TsuB) as an adjacent and essential protein for YeeE-mediated thiosulfate uptake in vivo. We further showed that S. thermophila YeeD possesses thiosulfate decomposition activity and that a conserved cysteine in YeeD was modified to several forms in the presence of thiosulfate. Finally, the crystal structures of S. thermophila YeeE-YeeD fusion proteins at 3.34-[A] and 2.60-[A] resolutions revealed their interactions. The association was evaluated by a binding assay using purified S. thermophila YeeE and YeeD. Based on these results, a model of the sophisticated uptake of thiosulfate ions by YeeE and YeeD is proposed. 125-character sentenceThe structural study of the YeeE-YeeD membrane protein complex provides insight into the thiosulfate uptake and degradation mechanism.

biochemistry↗

bioRxiv2021

A glycosylphosphatidylinositol-anchored α-amylase encoded by amyD contributes to a decrease in the molecular mass of cell wall α-1,3-glucan in Aspergillus nidulans

-1,3-Glucan is one of the main polysaccharides in the cell wall of Aspergillus nidulans. We previously revealed that it plays a role in hyphal aggregation in liquid culture, and that its molecular mass (MM) in an agsA-overexpressing (agsAOE) strain was larger than that in an agsB-overexpressing (agsBOE) strain. The mechanism that regulates the MM of -1,3-glucan is poorly understood. Although the gene amyD, which encodes glycosyl-phosphatidylinositol (GPI)-anchored -amylase (AmyD), is involved in the biosynthesis of -1,3-glucan in A. nidulans, how it regulates this biosynthesis remains unclear. Here we constructed strains with disrupted amyD ({Delta}amyD) or overexpressed amyD (amyDOE) in the genetic background of the ABPU1 (wild-type), agsAOE, or agsBOE strain, and characterized the chemical structure of -1,3-glucans in the cell wall of each strain, focusing on their MM. The MM of -1,3-glucan from the agsBOE amyDOE strain was smaller than that in the parental agsBOE strain. In addition, the MM of -1,3-glucan from the agsAOE {Delta}amyD strain was greater than that in the agsAOE strain. These results suggest that AmyD is involved in decreasing the MM of -1,3-glucan. We also found that the C-terminal GPI-anchoring region is important for these functions.

microbiology↗

bioRxiv2019

Thick perirenal fat predicts the growth pattern of renal cell carcinoma

ObjectiveTo examine the relationship between the direction of renal cell carcinoma growth and the visceral/perirenal fat volume.\n\nPatients and Methods: We retrospectively reviewed computed tomography scans of 153 patients with stage 1 renal cell carcinoma who underwent radical or partial nephrectomy in our hospital between January 2013 and July 2016. We calculated the visceral/subcutaneous/perirenal fat volumes using SYNAPSE VINCENT(R). Of the 60 patients, the perirenal fat was immunohistochemically stained for leptin, adiponectin, COX-2 and UCP-1, and the association with outward tumor protrusion was evaluated.\n\nResultsOf the 153 cases, 88 had confirmed outward expansion (57.5%), 110 were classed as pT1a (52 and 58 with outer and inner expansion, respectively), 43 were classed as pT1b (36 and 7 with outer and inner expansion, respectively; P<0.0001). Multivariate logistic regression model showed a trend toward significance in pT1b (vs pT1a, [OR] 6.033, 95%CI=2.409-15.108, P=0.0001), perirenal fat percentage >1.0 (vs [≤]1.0, [OR] 2.596, 95%CI=1.205-5.591, P=0.014). as independent predictors for outer protrusion. Immunohistochemical staining was positive for UCP-1 expression in 31 out of 41 outgrowth types (75.6%), and all 19 endogenous types (100%; P=0.003).\n\nConclusionsRenal cell carcinoma with thick perirenal fat correlates with an increased likelihood of developing outward tumor protrusion; therefore, fat distribution may affect the development of renal cell carcinoma.

cancer biology↗

bioRxiv2019

Generation of human hepatic progenitor cells with regenerative and metabolic capacities from primary hepatocytes

Hepatocytes are regarded as the only effective cell source for cell transplantation to treat liver diseases; however, their availability is limited due to a donor shortage. Thus, a novel cell source must be developed. We recently reported that mature rodent hepatocytes can be reprogrammed into progenitor-like cells with a repopulative capacity using small molecule inhibitors. Here, we demonstrate that hepatic progenitor cells can be obtained from human infant hepatocytes using the same strategy. These cells, named human chemically induced liver progenitors (hCLiPs), had a significant repopulative capacity in injured mouse livers following transplantation. hCLiPs redifferentiated into mature hepatocytes in vitro upon treatment with hepatic maturation-inducing factors. These redifferentiated cells exhibited cytochrome P450 (CYP) enzymatic activities in response to CYP-inducing molecules and these activities were comparable with those in primary human hepatocytes. These findings will facilitate liver cell transplantation therapy and drug discovery studies.

cell biology↗