Biology subjects

Takada, Y.

Publications and source records attributed to Takada, Y..

13 recordsLinked to original sources

bioRxiv2026

Mouse oocytes organize mRNA-protein assemblies and ER-associated translation hubs to drive meiosis

Oocyte meiosis in animals is essential for producing fertilizable eggs. As transcription becomes largely quiescent, meiotic progression depends on temporally regulated translation of accumulated maternal mRNAs. However, the underlying mechanisms remain unclear. Here, we identified diverse and highly organized mRNA-protein assemblies in mouse oocytes that coordinate cytoplasmic mRNA regulation and enable spatiotemporal translation to drive oocyte meiosis. Twelve RNA-binding proteins (RBPs), including DDX4/VASA, HDLBP/Vigilin, and TDRD3, were assembled into widely distributed aggregates in germinal vesicle (GV)-stage oocytes. We depleted DDX4 and HDLBP using Trim-Away and found that these RBPs regulate the precise assembly of RNA granules and protein aggregates through distinct mechanisms, promoting temporal protein synthesis at cortical translation factories with the endoplasmic reticulum, termed CORTE. All 12 RBPs formed distinct mRNA-protein assemblies, whose size and distribution changed upon meiotic initiation. These findings reveal the molecular basis of spatiotemporal mRNA-protein-organelle networks underlying animal reproduction and development.

cell biology↗

bioRxiv2026

Ecological drivers of social complexity: the role of predation risk and nesting resource in group-living cichlid

Understanding how social complexity responds to environmental variation remains a longstanding challenge in evolutionary biology. Here, we investigated the drivers of social complexity using intraspecific social variation across seven locations of the obligatory shell-brooding cichlid Neolamprologus meeli in Lake Tanganyika. We quantified the number of subordinate individuals per female territory and examined the effects of predation risk, shell availability, and their interaction. Social complexity increased with shell availability under high predation risk but showed little association under low predation risk. A field manipulative-experiment further demonstrated that increasing shell availability led to higher juvenile retention, indicating a causal effect of territory quality. In addition, removal of subordinates reduced shell availability, suggesting the feedback between group size and territory maintenance. We also assessed genetic population structure based on nuclear SNPs obtained by MIG-seq and found only weak genetic differentiation among localities, suggesting that the observed social variation is unlikely to simply reflect strong genetic subdivision. Together, these results show that predation risk promotes group living, whereas nesting resource availability constrains its extent. Our study highlights that social complexity emerges from the interaction between macro- and micro-ecological factors, providing a mechanistic understanding of the evolution of social complexity and philopatry.

ecology↗

bioRxiv2025

Ivermectin binds to the allosteric site (site 2) and inhibits allosteric integrin activation by pro-inflammatory cytokines.

Ivermectin is known to have anti-inflammatory properties, but the specifics of this action are unknown. We recently showed that multiple pro-inflammatory cytokines (e.g., FGF2, CCL5, CD40L) bind to the allosteric site (site 2) of integrins and activate them. 25-Hydroxycholesterol, a pro-inflammatory lipid mediator, is known to bind to site 2 and induce integrin activation and inflammatory signals (e.g., IL-6 and TNF secretion), suggesting that site 2 is critically involved in inflammation. We showed that TNF, a major pro-inflammatory cytokine, binds to site 2 and induces integrin activation, like other pro-inflammatory cytokines. We recently showed that two anti-inflammatory cytokines (FGF1 and NRG1) bind to site 2 and inhibit integrin activation by inflammatory cytokines. We hypothesized that ivermectin binds to site 2 but inhibits the integrin activation induced by inflammatory cytokines in a mechanism similar to anti-inflammatory cytokines (site 2 antagonists). Consistently, docking simulation predicts that ivermectin binds to site 2. We found that ivermectin suppressed the activation of soluble {beta}3 integrins by multiple pro-inflammatory cytokines (including TNF) in a dose-dependent manner in cell-free conditions, suggesting that ivermectin acts as an antagonist for site 2. This may be a potential mechanism of anti-inflammatory action of ivermectin.

biochemistry↗

bioRxiv2025

The EC2 domains of tetraspanins CD9, CD81, and CD151 bind to the allosteric site of integrins (site 2) and activate integrins αvβ3, α5β1 and α4β1 in a biphasic manner.

Previous studies showed that tetraspanins activate integrins, but the mechanism of this action is unclear. We previously showed that the extracellular-2 (EC2) domains of CD9, CD81, and CD151 bind to the classical RGD-binding site (site 1) of integrin v{beta}3, suggesting that they are integrin ligands. We showed that several inflammatory cytokines (e.g., CX3CL1, CXCL12, CCL5, and CD40L) bind to the allosteric site (site 2) of integrins, which is distinct from site 1, and activate integrins (allosteric activation). 25-hydroxycholesterol, a major inflammatory lipid mediator, is known to bind to site 2 and induce inflammatory signals, suggesting that site 2 plays a role in inflammatory signaling. We hypothesized that the EC2 domains activate integrins by binding to site 2. Here we describe that docking simulation predicted that CD81 EC2 binds to site 2 of v{beta}3 and more strongly to site 2 of 5{beta}1. Peptide from site 2 bound to isolated EC2 domains, suggesting that the EC2 domains bind to site 2. The EC2 domains only weakly activated v{beta}3 but more efficiently activated cell surface integrins 5{beta}1 and 4{beta}1 on the cell surface. These results are consistent with the previous findings that these tetraspanins preferentially interact with {beta}1 integrins. The integrin activation by the EC2 domains was increased at low EC2 concentrations and reduced as EC2 concentrations increased (biphasic), which is consistent with the findings that the EC2 domains bind to two sites (site 1 and 2). We propose that the EC2 binding to site 2 is a novel target for drug discovery.

biochemistry↗

bioRxiv2024

Blue light toxicity drives the gut microbiota-mediated lipid accumulation in Drosophila

Human agriculture has always raced against insect adaptation, requiring updated pest control methods and knowledge of evolutionary processes. Excessive exposure to blue light (BL) kills a wide range of insect species and has attracted attention as an alternative to chemical pesticides. Here, to understand how insects adapt to BL toxicity, we investigated evolutionary responses to BL toxicity in Drosophila melanogaster over 70 generations using laboratory selection experiments. The selected line exhibited an obese phenotype accompanied by midgut elongation, with BL tolerance dependent on gut microbiota- mediated lipid accumulation. Whole-genome and transcriptome analyses consistently highlighted interactions between the microbiota and host lipid metabolism-related genes. Remarkably, manipulating genes associated with lipid accumulation conferred BL tolerance even in the absence of selection. We suggest that the acquisition of BL tolerance occurs through adaptive obesity. Our study introduces a mechanism of evolutionary adaptation of insects against BL-based selective pressure by maximising the benefits from the gut microbiota via midgut elongation. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=141 SRC="FIGDIR/small/608892v5_ufig1.gif" ALT="Figure 1"> View larger version (62K): org.highwire.dtl.DTLVardef@111c9d5org.highwire.dtl.DTLVardef@d9bf7forg.highwire.dtl.DTLVardef@1ab46fborg.highwire.dtl.DTLVardef@1c111cd_HPS_FORMAT_FIGEXP M_FIG C_FIG Brief summaryLaboratory selection for tolerance to blue light toxicity in Drosophila revealed that gut microbiota-mediated obesity, marked by midgut elongation and increased beneficial bacteria, is a key evolutionary adaptation.

evolutionary biology↗

bioRxiv2024

FGF2 binds to the allosteric site (site 2) and activates αvβ3 integrin and FGF1 binds to site 2 but suppresses integrin activation by FGF2: a potential mechanism of anti-inflammatory action of FGF1

FGF1 is known as an anti-inflammatory and has suppresses insulin resistance. Its homologue FGF2 is pro-inflammatory. Mechanism of FGF1s anti-inflammatory action and FGF2s pro-inflammatory action are unknown. Several inflammatory cytokines (e.g., CX3CL1, CCL5, and CXCL12, and CD40L) bind to the classical ligand (RGD)-binding site (site 1) of integrin v{beta}3. In addition, they bind to the allosteric site (site 2) of v{beta}3, which is distinct from site 1, and allosterically activate v{beta}3. Site 2 is involved in inflammatory signals since inflammatory lipid mediator 5-hydroxycholesterol binds to site 2 and induces integrin activation and inflammatory signals (e.g., TNF and IL-6 secretion). We thus hypothesized that FGF1 and FGF2 bind to site 2 and affect activation status of integrins. Here we describe that FGF2 bound to site 2 and allosterically activated v{beta}3 integrin. Point mutations in the site 2-binding interface of FGF2 suppressed this activation, indicating that FGF2 binding to site 2 is required for inducing integrin activation. In contrast, FGF1 bound to site 2 but did not activate v{beta}3, and instead suppressed integrin activation induced by FGF2, indicating that FGF1 acts as an antagonist of site 2. These findings suggest that FGF1s anti-inflammatory action is mediated by blocking site 2. FGF1 has potential as an anti-inflammatory agent, but is not appropriate for long-term use since it is potent mitogen. A non-mitogenic FGF1 mutant (R50E), which is defective in binding to site 1 of v{beta}3, suppressed v{beta}3 activation by FGF2 as effectively as WT FGF1. We propose that FGF1 R50E has therapeutic potential for inflammatory diseases.

biochemistry↗

bioRxiv2024

FGF2 binds to the allosteric site (site 2) and activates integrin αIIbβ3 and FGF1 binds to site 2 but suppresses integrin activation by FGF2: A potential mechanism of anti-thrombotic action of FGF1.

It has been believed that platelet integrin IIb{beta}3 recognizes fibrinogen and several ECM proteins, and we recently showed that IIb{beta}3 binds to several inflammatory cytokines (e.g., CCL5, and CXCL12), which are stored in platelet granules. These ligands bind to the classical ligand (RGD)-binding site (site 1) of integrin IIb{beta}3. Also, they bind to the allosteric site (site 2) of IIb{beta}3, which is distinct from site 1, and allosterically activate IIb{beta}3. Site 2 is known to be involved in allosteric integrin activation and inflammatory signaling. FGF2 is also stored in platelet granules and known to be pro-thrombotic, but it is unclear if FGF2 binds to IIb{beta}3. We studied if FGF2 and its homologue FGF1 bind to IIb{beta}3 and induce allosteric activation. FGF1 (not stored in platelet granules) is known to be anti-thrombotic. Mechanism of FGF1s anti-thrombotic action is unknown. Here we describe that FGF1 and FGF2 bound to site 1 of IIb{beta}3, indicating that IIb{beta}3 is a new receptor for FGF1/2. Notably, FGF2 bound to site 2 and allosterically activated IIb{beta}3. Point mutations in the site 2-binding interface of FGF2 suppressed this activation, indicating that FGF2 binding to site 2 is required for activation (FGF2 is an agonist to site 2). In contrast, FGF1 bound to site 2 but did not activate IIb{beta}3, and instead suppressed integrin activation induced by FGF2, indicating that FGF1 acts as an antagonist of site 2. A non-mitogenic FGF1 mutant (R50E), which is defective in binding to site 1 of v{beta}3, suppressed IIb{beta}3 activation by FGF2 as effectively as WT FGF1. We propose that FGF1 R50E has therapeutic potential for anti-thrombosis.

biochemistry↗

bioRxiv2022

Diagnosis of type 2 Diabetes Mellitus (T2DM) using Paired microRNAs

Type 2 Diabetes mellitus (T2DM) is one of the most common diseases in the world and its prevalence ratio is still increasing. Patients with T2DM have diverse pathophysiological changes like as macrovascular, microvascular diseases, cancers as well as abnormal glucose metabolism. Thus, there are urgent needs to develop relevant biomarkers for the broad range of pathophysiology in patients with T2DM. We analyzed the signatures of serum miRNAs with the miRNA array analysis and reverse-transcription based quantitative polymerase chain reaction (RT-qPCR) in 50 patients with type 2 DM (T2DM) and 15 normal subjects. Array analysis showed that 19 miRNAs were up-regulated more than 2-fold and 71 miRNAs were down-regulated less than 0.5 in T2DM in comparison with normal subjects. Top 5 of up-regulated miRNAs were miR-3619-3p, miR-557, miR-6850-5p, miR-3648, miR-4730, and 5 of most down-regulated miRNAs were miR-5100, miR-4454, miR-1260b, miR-7975, miR-6131. We selected 4 miRNAs for validation analysis with RT-qPCR based on the abundance enough for reliable analyses and disease-specificities reported in previous reports. Serum miR-126-3p was down-regulated (3.21-fold, p<0.05) in T2DM, and miR-10a up-regulated (1.94-fold, p<0.05). However, none of single miRNA had significant correlation with clinical data and state. Data of the paired miRNAs: miR-10a and miR-200c, or miR-126 and miR-10a, clearly differentiated T2DM patients from normal subjects (p<0.05). Our study showed the paired-miRNA analyses as the more effective diagnostics for T2DM than the single miRNA analysis.

molecular biology↗

bioRxiv2021

Tdrd3 regulates the progression of meiosis II through translational control of Emi2 mRNA in mouse oocytes

After completion of meiosis I, the oocyte immediately enters meiosis II and forms a metaphase II (MII) spindle without an interphase, which is fundamental for generating a haploid gamete. Here, we identify tudor domain-containing protein 3 (Tdrd3) as a novel regulator of oocyte meiosis. Although early mitotic inhibitor 2 (Emi2) protein has been shown to ensure the meiosis I to II transition and the subsequent MII spindle formation by inhibiting the anaphase-promoting complex/cyclosome (APC/C), how it accumulates after meiosis I has remained unresolved. We isolated Tdrd3 as a protein directly binding to Emi2 mRNA. In GV-stage mouse oocytes, Emi2 mRNA assembled into RNA granules containing Tdrd3, while cyclin B1 mRNA, which was translated in early meiosis I, formed different granules. Knockdown of Tdrd3 attenuated Emi2 synthesis in meiosis II without affecting cyclin B1 synthesis in meiosis I. Moreover, Tdrd3-deficient oocytes entered interphase and failed to form an MII spindle after completion of meiosis I. Taken together, our results indicate the importance of Tdrd3-mediated translational control of Emi2 mRNA, which promotes Emi2 synthesis in meiosis II, for the progression of meiosis.

developmental biology↗

bioRxiv2020

Ethanolamine phosphate on the second mannose as bridge in GPI anchored proteins: Towards understanding inherited PIGG deficiency

Glycosylphosphatidylinositols (GPIs) are glycolipids that anchor many proteins (GPI-APs) on the cell surface. The core glycan of GPI precursor has three mannoses, which in mammals, are all modified by ethanolamine-phosphate (EthN-P). It is postulated that EthN-P on the third mannose (EthN-P-Man3) is the bridge between GPI to the protein and the second (EthN-P-Man2) is removed after GPI-protein attachment. However, EthN-P-Man2 may not be always transient, as mutations of PIGG, the enzyme that transfers EthN-P to Man2, result in inherited GPI deficiencies (IGDs), characterized by neuronal dysfunctions. Here, we show EthN-P on Man2 is the preferential bridge in some GPI-APs, among them, the ect-5-nucleotidase and netrin G2. We found that CD59, a GPI-AP, is attached via EthN-P-Man2 both in PIGB-knockout cells, in which GPI lacks Man3 and with a small fraction, in wild type cells. Our findings modify the current view of GPI anchoring and provide mechanistic bases of IGDs caused by PIGG mutations.

molecular biology↗

bioRxiv2020

Changes in subcellular structures and states of Pumilio1 regulate the translation of target Mad2 and Cyclin B1 mRNAs

Temporal and spatial control of mRNA translation has emerged as a major mechanism for promoting diverse biological processes. However, the molecular nature of temporal control of translation remains unclear. In oocytes, many mRNAs are deposited as a translationally repressed form and are translated at appropriate timings to promote the progression of meiosis and development. Here, we show that changes in structures and states of the RNA-binding protein Pumilio1 regulate the translation of target mRNAs and progression of oocyte maturation. Pumilio1 was shown to bind to Mad2 and Cyclin B1 mRNAs, assemble highly clustered solid-like aggregates, and surround Mad2 and Cyclin B1 RNA granules in mouse oocytes. These Pumilio1 aggregates were dissolved by phosphorylation prior to the translational activation of target mRNAs. Stabilization of Pumilio1 aggregates prevented the translational activation of target mRNAs and oocyte maturation. Together, our results provide an aggregation-dissolution model for the temporal and spatial control of translation.

cell biology↗

bioRxiv2020

A meiosis-specific factor MRM/C19orf57 modulates localization of RAD51 and DMC1 recombinases to DSBs in mouse meiotic recombination

Meiotic recombination is critical for genetic exchange and generation of chiasmata that ensures faithful chromosome segregation during meiosis I. Meiotic recombination is initiated by DNA double-strand break (DSB) followed by multiple processes of DNA repair. The exact mechanisms how recombinases localize to DSB remained elusive. Here we show that C19orf57/4930432K21Rik/BRME1 is a new player for meiotic recombination in mice. C19orf57/4930432K21Rik/BRME1 associates with ssDNA binding proteins, BRCA2 and MEILB2/HSF2BP, critical recruiters of recombinases onto DSB sites. Disruption of C19orf57/4930432K21Rik/BRME1 shows severe impact on DSB repair and male fertility. Remarkably, removal of single stranded DNA (ssDNA) binding proteins from DSB sites is delayed, and reciprocally the loading of RAD51 and DMC1 onto resected ssDNA is impaired in Brme1 KO spermatocytes. We propose that C19orf57/4930432K21Rik/BRME1 modulates localization of recombinases to meiotic DSB sites through the interaction with the BRCA2-MEILB2/HSF2BP complex during meiotic recombination.

molecular biology↗

bioRxiv2019

Cross-talks of glycosylphosphatidylinositol biosynthesis with glycosphingolipid biosynthesis and ER-associated degradation

Glycosylphosphatidylinositol (GPI)-anchored proteins and glycosphingolipids interact with each other in the mammalian plasma membranes, forming dynamic microdomains. How their interaction starts in the cells has been unclear. Here, based on a genome-wide CRISPR-Cas9 genetic screen for genes required for GPI side-chain modification by galactose in the Golgi apparatus, we report that {beta}1,3-galactosyltransferase 4 (B3GALT4), also called GM1 ganglioside synthase, additionally functions in transferring galactose to the N-acetylgalactosamine side-chain of GPI. Furthermore, B3GALT4 requires lactosylceramide for the efficient GPI side-chain galactosylation. Thus, our work demonstrates previously unexpected evolutionary and functional relationships between GPI-anchored proteins and glycosphingolipids in the Golgi. Through the same screening, we also show that GPI biosynthesis in the endoplasmic reticulum (ER) is severely suppressed by ER-associated degradation to prevent GPI accumulation when the transfer of synthesized GPI to proteins is defective. Our data demonstrates cross-talks of GPI biosynthesis with glycosphingolipid biosynthesis and the ER quality control system.

cell biology↗