Biology subjects

Shibuya, A.

Publications and source records attributed to Shibuya, A..

11 recordsLinked to original sources

bioRxiv2026

CD300a Blockade attenuates renal fibrosis by enhancing macrophage efferocytosis and neutrophil MMP9 production

Renal fibrosis represents an irreversible endpoint of chronic kidney disease. Although macrophage efferocytosis regulates the response to acute renal injury, the mechanisms by which early efferocytosis confers long-term protection against chronic fibrosis remain elusive. Here, we show that phagocyte-specific deletion of the inhibitory receptor CD300a (Cd300afl/flLyz2-Cre) mitigates obstructive renal injury and subsequent fibrosis. CD300a deficiency enhanced efferocytosis by tissue-resident macrophages during the early phase of injury, thereby suppressing the release of damage-associated molecular patterns (DAMPs) and TGF-{beta} expression. This early attenuation of the inflammatory microenvironment altered the composition of recruited neutrophils, shifting them from profibrotic SiglecF+ toward non-profibrotic SiglecF- subsets. Furthermore, loss of CD300a cell-autonomously primed neutrophils for enhanced matrix metalloproteinase-9 (MMP9) production upon injury, whereas neutrophil depletion abolished the protective effect of CD300a deficiency. Importantly, administration of a neutralizing anti-CD300a antibody to wild-type mice recapitulated these protective effects, promoting the shift toward SiglecF- neutrophils and significantly attenuating renal fibrosis. Together, our findings identify CD300a as a regulator of early macrophage efferocytosis and neutrophil-derived MMP9 production, and establish pharmacological CD300a blockade as a potential therapeutic strategy for progressive renal fibrosis.

immunology↗

bioRxiv2026

SPIN1 selectively silences evolutionarily young transposable elements through chromatin regulation

Mammalian genomes encode multiple transposable element (TE) silencing pathways that distinguish their targets through different molecular features, with KRAB zinc finger proteins recognizing DNA sequence, the HUSH complex sensing intronless transcripts, and the PIWI-piRNA pathway using small RNA guides. How chromatin state itself contributes to TE recognition remains less defined. Here we identify Spindlin1 (SPIN1), a three-Tudor-domain histone reader implicated in germline piRNA-directed DNA methylation, as a transcriptional repressor of evolutionarily young TEs in mouse embryonic stem cells. SPIN1 selectively binds LINE and ERV loci carrying H3K4me3 and H3K9me3, a chromatin signature enriched at young, transcription-permissive elements, and recognition of these marks by Tudor domains 1 and 2 is required for TE targeting. SPIN1 engages SPINDOC as a Tudor 1 and 3-dependent cofactor whose loss phenocopies SPIN1 depletion, and associates with the H3K9 methyltransferases SETDB1 and G9a. SPIN1 loss reduces H3K9me3 and is accompanied by increased chromatin accessibility, without altering DNA methylation. Thus, SPIN1 uses a histone-state-based mechanism to identify and repress young TEs in pluripotent cells, mechanistically distinct from its germline mode in which SPIN1 cooperates with the PIWI-piRNA pathway to promote DNA methylation, illustrating how a single histone reader engages distinct silencing machineries across cellular contexts.

molecular biology↗

bioRxiv2026

Soluble CD155 immune complexes reprogram DNAM-1 signaling to potentiate antitumor immunity

Resistance to immune checkpoint inhibitors (ICIs) remains a major challenge in oncology, yet the mechanisms that selectively disable activating pathways are poorly defined. Here, we identify tumor-derived soluble CD155 (sCD155) as a systemic checkpoint that rewires DNAM-1 signaling to drive immunotherapy resistance. High plasma sCD155 levels correlate with impaired anti-PD-1 responses in patients with non-small cell lung cancer. Mechanistically, sCD155 selectively suppresses DNAM-1-dependent activation of CD8+ T and NK cells, uncoupling ICIs from cytotoxic function. Intriguingly, a selective anti-sCD155 monoclonal antibody does not neutralize the ligand, but rather converts it into an activating scaffold. This complex induces Fc{gamma}R-anchored DNAM-1 microcluster formation and robust downstream signaling, effectively switching the checkpoint into a co-stimulatory signal. This reprogramming restores CTL function, suppresses metastasis, and augments PD-1/TIGIT blockade to achieve durable immunity. Our findings establish antibody-mediated receptor architecture rewiring as a therapeutic principle to overcome cancer immune resistance.

immunology↗

bioRxiv2026

DNAM-1 immunoreceptor integrates innate and adaptive immune programs to drive intestinal inflammation

Innate and adaptive immune responses play critical roles in the pathogenesis of inflammatory bowel disease (IBD), yet the molecular pathways integrating these responses remain elusive. Here, we identify DNAM-1 immunoreceptor as a central driver of colitis through distinct, cell type-specific mechanisms. Transcriptomic analyses of human and murine group 3 innate lymphoid cells (ILC3s) revealed DNAM-1 as a conserved IL-23-responsive surface molecule associated with inflammatory cytokine production. In an innate immune-driven anti-CD40 monoclonal antibody (mAb)-induced colitis model, DNAM-1 expressed on ILC3s promoted intestinal inflammation by enhancing IL-22 and GM-CSF production via the integration of the Akt-mTORC1-HIF-1 signaling pathway. Genetic ablation or antibody-mediated blockade of DNAM-1 attenuated inflammatory cytokine production and disease severity. Paradoxically, in T cell-dependent colitis, DNAM-1 expression on dendritic cells, but not on ILC3s or CD4 T cells, exacerbated disease by promoting dendritic cell activation and pathogenic Th1 and Th17 differentiation. Notably, therapeutic blockade of DNAM-1 ameliorated disease in both colitis models and exerted complementary effects when combined with anti-TNF therapy, accompanied by modulation of immune activation programs distinct from those regulated by TNF inhibition. Collectively, these findings establish DNAM-1 as a pivotal regulator of intestinal inflammation bridging innate and adaptive immunity and identify DNAM-1 blockade as a next-generation therapeutic strategy for IBD. Highlight{blacktriangleright} DNAM-1 is an IL-23-responsive receptor conserved in human and mouse ILC3s. {blacktriangleright}DNAM-1 on ILC3s drives innate colitis via Akt-mTORC1-HIF-1 signaling. {blacktriangleright}DNAM-1 on DCs promotes T cell-dependent colitis by inducing Th1/Th17 cells. {blacktriangleright}DNAM-1 blockade targets immune pathways distinct from TNF inhibition. {blacktriangleright}Combined DNAM-1 and TNF blockade shows additive therapeutic efficacy in colitis.

immunology↗

bioRxiv2026

Two Glu/Asp Residues Cooperatively Mediate an Early Step of ATP Hydrolysis in GHKL ATPases MutL and GyrB

GHKL ATPases share a unique Bergerat ATP-binding fold and regulate diverse biological processes through ATP-dependent conformational changes. An early step of ATP hydrolysis in this family has been attributed to a single highly conserved glutamate residue proposed to function as the general base. However, mutations of this residue impair both the ATPase activity and ATP binding, complicating interpretation of its catalytic role. Re-examination of the high-resolution crystal structures revealed a second conserved acidic residue positioned within a hydrogen-bonding distance from the nucleophilic water molecule. Using Aquifex aeolicus MutL and GyrB as model enzymes, we combined systematic mutagenesis, ATPase and ATP-binding assays, and X-ray crystallography to dissect the roles of these residues. We show that alignment of the nucleophilic water can be maintained as long as the conserved glutamate retains hydrogen-bonding capability, whereas efficient ATP hydrolysis requires proton-accepting capacity at least one of the two acidic residues. These results indicate that the conserved glutamate primarily governs positioning of the nucleophilic water, while activation of this water for catalysis is achieved through cooperative general base function of the two acidic residues. Extending this framework to human MutL homologs, PMS2 and MLH1, we showed that clinically reported variants of uncertain significance in these DNA mismatch repair proteins substantially reduced the ATPase activity, indicating functional impairment. Together, our findings refine the catalytic mechanism of GHKL ATPases and provide a structural and functional framework for interpreting disease-associated variants in GHKL ATPases. Phylogenetic and ancestral state analysis further indicated that the second acidic residue was likely to be present in the common ancestor of major GHKL ATPase lineages but was later modified in a branch including Hsp90, suggesting evolutionary remodeling of the catalytic mechanism in the branch.

biochemistry↗

bioRxiv2022

Liver type 1 innate lymphoid cells lacking IL-7 receptor are a native killer cell subset fostered by parenchymal niches

Group 1 innate lymphoid cells (G1-ILCs), including circulating natural killer (NK) cells and tissue-resident type 1 ILCs (ILC1s), are innate immune sentinels critical for responses against infection and cancer. In contrast to relatively uniform NK cells through the body, diverse ILC1 subsets have been characterized across and within tissues in mice, but their developmental and functional heterogeneity remain unsolved. Here, using multimodal in vivo approaches including fate-mapping and targeting of the interleukin 15 (IL-15)-producing microenvironment, we demonstrate that liver parenchymal niches support the development of a cytotoxic ILC1 subset lacking IL-7 receptor (7R- ILC1s). During ontogeny, fetal liver (FL) G1-ILCs arise perivascularly and then differentiate into 7R- ILC1s within sinusoids. Hepatocyte-derived IL-15 supports parenchymal development of FL G1-ILCs to maintain adult pool of 7R- ILC1s. IL-7R+ (7R+) ILC1s in the liver, candidate precursors for 7R- ILC1s, are not essential for 7R- ILC1 development in physiological conditions. Functionally, 7R- ILC1s exhibit killing activity at steady state through granzyme B expression, which is underpinned by constitutive mTOR activity, unlike NK cells with exogenous stimulation-dependent cytotoxicity. Our study reveals the unique ontogeny and functions of liver-specific ILC1s, providing a detailed interpretation of ILC1 heterogeneity.

immunology↗

bioRxiv2022

G307S DNAM-1 mutation exacerbates autoimmune encephalomyelitis via enhancing CD4+ T cell activation

Although rs763361, which causes a non-synonymous glycine-to-serine mutation at residue 307 (G307S mutation) of the DNAM-1 immunoreceptor, is a single-nucleotide polymorphism (SNP) associated with autoimmune disease susceptibility, little is known about how the SNP is involved in pathogenesis. Here, we established CD4+ T cells expressing wild-type or G307S DNAM-1 and showed that the costimulatory signal from G307S DNAM-1 induced greater pro-inflammatory cytokine production and cell proliferation than that from wild-type DNAM-1. The G307S mutation also enhanced the recruitment of the tyrosine kinase Lck and augmented tyrosine phosphorylation at residue 322 (Tyr322) of DNAM-1. However, conversion of Tyr322 of DNAM-1 to phenylalanine (Y322F mutation) canceled the enhanced activation of CD4+ T cell transfectants expressing G307S DNAM-1. Adoptive transfer of myelin-antigen-specific CD4+ T cells expressing G307S DNAM-1 into mice exacerbated experimental autoimmune encephalomyelitis compared with the transfer of cells expressing wild-type DNAM-1. These findings suggest that rs763361 is a gain-of-function mutation that enhances DNAM-1-mediated costimulatory signaling for proinflammatory responses.

immunology↗

bioRxiv2022

Mod(mdg4) variants repress telomeric retrotransposonHeT-A by blocking subtelomeric enhancers

Telomeres in Drosophila are composed of sequential non-LTR retrotransposons: HeT-A, TART, and TAHRE. Although they are repressed by the piRNA pathway in the germline, how these retrotransposons are regulated in somatic cells is poorly understood. Here, we show that specific splice variants of Mod(mdg4) repress HeT-A by blocking subtelomeric enhancers in ovarian somatic cells. We found that the Mod(mdg4)-N variant represses HeT-A expression most efficiently among the variants. Mod(mdg4)-N mutant flies show elevated HeT-A expression and female sterility. Mod(mdg4)-N-binding subtelomeric sequences exhibit enhancer-blocking activity, and recruitment of RNA polymerase II (Pol II) on subtelomeres by Mod(mdg4)-N is essential for this enhancer-blocking. Moreover, Mod(mdg4)-N functions to form chromatin boundaries of higher-order chromatin conformation but this mechanism is independent of its Pol II recruitment activity at telomeres/subtelomeres. This study provides a link between enhancer-blocking and telomere regulation, and two different molecular mechanisms exhibited by an insulator protein to orchestrate precise gene expression.

molecular biology↗

bioRxiv2020

Tumor-derived extracellular vesicles regulate tumor-infiltrating regulatory T cells via the immunoreceptor CD300a

Although tumor-infiltrating regulatory T (Treg) cells play a pivotal role in tumor immunity, how Treg cell activation are regulated in tumor microenvironments remains unclear. Here, we found that mice deficient in the inhibitory immunoreceptor CD300a on their dendritic cells (DCs) have increased numbers of Treg cells in tumors and greater tumor growth compared with wild-type mice after transplantation of B16 melanoma. Pharmacological impairment of extracellular vesicle (EV) release decreased Treg cell numbers in CD300a-deficient mice. Coculture of DCs with tumor-derived EV (TEV) induced the internalization of CD300a and the incorporation of EVs into endosomes, in which CD300a inhibited TEV-mediated TLR3-TRIF signaling for activation of the IFN-{beta}-Treg cells axis. We also show that higher expression of CD300A was associated with decreased tumor-infiltrating Treg cells and longer survival time in patients with melanoma. Our findings reveal the role of TEV and CD300a on DCs in Treg cell activation in the tumor microenvironment.

cancer biology↗

bioRxiv2019

Nuclear RNA export factor variant initiates piRNA-guided co-transcriptional silencing

The PIWI-interacting RNA (piRNA) pathway preserves genomic integrity by repressing transposable elements (TEs) in animal germ cells. Among PIWI-clade proteins in Drosophila, Piwi transcriptionally silences its targets through interactions with cofactors, including Panoramix (Panx) and forms heterochromatin characterized by H3K9me3 and H1. Here, we identified Nxf2, a nuclear RNA export factor (NXF) variant, as a protein that forms complexes with Piwi, Panx, and p15. Panx-Nxf2-p15 complex formation is necessary in the silencing by stabilizing protein levels of Nxf2 and Panx. Notably, ectopic targeting of Nxf2 initiates co-transcriptional repression of the target reporter in a manner independent of H3K9me3 marks or H1. However, continuous silencing requires HP1a and H1. In addition, Nxf2 directly interacts with target TE transcripts in a Piwi-dependent manner. These findings suggest a model in which the Nxf2-Panx-p15 complex enforces the association of Piwi with target transcripts to trigger co-transcriptional repression, prior to heterochromatin formation in the nuclear piRNA pathway.\n\nHighlightsO_LINxf2 plays an essential role in the Piwi-piRNA pathway\nC_LIO_LIFormation of Piwi-Panx-Nxf2-p15 (PPNP) complexes stabilizes both Panx and Nxf2\nC_LIO_LIThe PPNP complex triggers transcriptional silencing before heterochromatin formation\nC_LIO_LINxf2 directly binds to target transcripts in a Piwi-dependent manner\nC_LI

molecular biology↗

bioRxiv2018

A long-chain fatty acid elongase Elovl6 regulates mechanical damage-induced keratinocyte death and skin inflammation

Mechanical damage on the skin not only affect the barrier function but also induce various immune responses, which trigger or exacerbate the inflammation in healthy individuals and patients with inflammatory skin diseases. However, how mechanical damage-induced skin inflammation is regulated remains largely unknown. Here, we show that mechanical damage due to tape stripping triggered keratinocyte death and release of danger-associated molecular patterns (DAMPs) such as high-mobility group box 1 protein (HMGB-1) and IL-1, which induced production of proinflammatory cytokines and chemokines IL-1{beta} and CXCL-1 by keratinocytes in mice. We also show that a long-chain fatty acid elongase Elovl6 is expressed in keratinocytes. Mice deficient in Elovl6 had increased epidermal levels of cis-vaccenic acid (CVA); this accelerated keratinocyte death triggered by tape stripping and release of DAMPs and exacerbated skin inflammation. Our results demonstrate that Elovl6 regulates mechanical damage-triggered keratinocyte death and skin inflammation.

immunology↗