Cell reports最新文献

Bacillus methanolicus, a unique plasmid-dependent and thermophilic methylotroph, is an ideal chassis for one-carbon (C1) biomanufacturing. Despite its evolutionary uniqueness and industrial promise, the synthetic biology toolkit remains limited in comparison to that of conventional model microorganisms. Here, we present a comprehensive toolkit comprising a high-efficiency electroporation protocol, a CRISPR-Cas9 method enabling robust and multiplex genome editing, diverse neutral loci for gene integration, and a cloud-based genome-scale metabolic model iBM822 for user-friendly biodesign. Leveraging this toolkit, we systematically dissected plasmid-dependent methylotrophy, restriction-modification machinery, and the functional significance of chromosomal methylotrophic genes. To address plasmid loss-induced strain degeneration, we integrated the large endogenous plasmid pBM19 into the chromosome for stable and intact methylotrophic growth. Finally, by integrating metabolic modeling with CRISPR-Cas9 editing, we engineered L-arginine feedback regulation to achieve L-arginine overproduction from methanol. This study establishes a synthetic biology framework for B. methanolicus, promoting mechanistic exploration of methylotrophy and C1 biomanufacturing.

TRPV1, a member of the transient receptor potential vanilloid subfamily, mediates nociception and thermoregulation. TRPV1-targeting analgesics frequently induce hyperthermia, underscoring the need for structural insights to guide the development of safer compounds. Here, we determined the structures of rat TRPV1 bound to the clinical candidate analgesics AMG517, AMG9810, and SB366791. AMG517 and AMG9810 are deeply situated within the S3-S4 interface of the vanilloid pocket, where they interact with residues from the S3-S6 helices, as well as the S4-S5 linker. These interactions induce local deformations in the TRP-box and lower S6 helix, accompanied by a modest rotation of the S1-S4 bundle, leading to partial dilation of the lower gate. The distinct allosteric changes of AMG517 and AMG9810, compared with the non-hyperthermic ligand SB366791, suggest a structural basis by which TRPV1-targeting analgesics influence thermoregulation and provide insights for designing safer analogs.

Type 1 conventional dendritic cells (cDC1s) are infrequent immune cells with an essential role in orchestrating immune responses to malignancies or infections. Despite their significance in regulating adaptive immunity, the absence of efficient manufacturing techniques to produce sufficient cDC1s hinders their therapeutic application. Here, we show that interleukin 4 (IL-4) markedly increases the yield of cDC1 cells derived from Flt3 ligand cultures of mouse and human progenitors, while concurrently inhibiting plasmacytoid DCs development. IL-4 predominantly acts on DC progenitors, and its activity requires cell-intrinsic IL4-RA or STAT6 signaling. Both in vitro and in vivo, IL-4-stimulated cDC1s efficiently prime cytotoxic CD8⁺ T cells. Transcriptomic analyses reveal that IL-4 promotes cDC1 proliferation, providing a mechanistic basis for the enhanced output. Together, these findings uncover an unexpected role for IL-4 in driving the development and scalable production of bona fide cDC1s, facilitating mechanistic studies and supporting their future therapeutic application in human disease.

Neoantigens are crucial for antitumor immunity and immune checkpoint inhibitor (ICI) efficacy by triggering strong immune responses. However, conventional methods for identifying neoantigens, such as whole-exon sequencing and short-read RNA sequencing (RNA-seq), appear to be insufficient, and the tumor mutational burden cannot sufficiently predict ICI efficacy. In this study, we employed a proteogenomic approach using long-read RNA-seq with Pacific Biosciences Single-Molecule Real-Time Sequencing technology to analyze full-length transcripts in combination with the human leukocyte antigen ligandome. As a result, many neoantigen candidates were identified, which were unregistered in a comprehensive database, including those from non-coding regions. Additionally, we validated the responses of specific T cell receptors (TCRs) to these candidates and identified several pairs of TCRs and neoantigens. These findings highlight the presence of more diverse neoantigens than expected that cannot be identified by conventional methods.

Proteins are the most abundant source of amino acids in body fluids. However, the potential contribution of extracellular protein catabolism to the regulation of T cell immunity remains poorly understood. In this study, we show that endocytosed extracellular proteins function as an amino acid source in activated T cells, maintaining mTORC1 activity and sustaining cytokine production following T cell activation. Genetic ablation of Tfe3 impairs the activation-induced upregulation of lysosomal genes and disrupts extracellular protein catabolism, resulting in attenuated mTORC1 signaling and compromised anti-viral and anti-tumor T cell responses. The TFE3-protein-mTORC1 signaling axis demonstrates clinical relevance. CD8⁺PD-1⁺ tumor-infiltrating T cells from older patients with lung cancer display reduced lysosomal degradation capacity and impaired cytokine secretion compared to their middle-aged counterparts. This functional defect is rescued by treatment with Vismodegib, a TFE3-inducing drug. Our findings reveal lysosome-mediated extracellular protein catabolism as an important metabolic pathway supporting T cell immunity.

Hyperactivation of the cell cycle in cancer cells suppresses antitumor immunity. The endogenous cyclin-dependent kinase inhibitor p57 is an important tumor suppressor and a potential therapeutic target for hepatocellular carcinoma (HCC). However, the immunomodulatory role of p57 remains unclear. Using samples from patients with HCC, we found that p57 expression correlated with an improved response to immune checkpoint inhibitors (ICIs) and increased CD8⁺ T cell infiltration. Mechanistically, p57 induced chromosomal instability and subsequently stimulated cGAS-STING-type I IFN signaling, leading to upregulation of the chemokines CCL5 and CXCL10, which promoted CD8⁺ T cell infiltration. Meanwhile, p57 also increased the expression of PD-L1 on the surface of HCC cells. Moreover, combining p57 overexpression with anti-PD-1 treatment synergistically inhibited tumor growth in vivo. Our studies demonstrated that p57 may serve as a biomarker for ICI efficacy, and increasing p57 expression is a potential therapeutic strategy to increase the efficacy of immunotherapy.

Impaired intestinal barrier function is a major feature of Crohn's disease, leading to exacerbated inflammation in response to the microbiota. In this context, the translocation of intestinal bacteriophages (phages) and their effects on the host haven't been fully investigated. We use phage fluorescence imaging coupled with ex vivo, in vitro, and in vivo models that mimic physiological and inflammatory conditions and find that phages can translocate across the intestinal barrier without disrupting its integrity. While changes in the epithelial barrier integrity selectively impact phage translocation rates, alterations in the permeability of the vascular endothelium do not affect phage crossing. Virome analysis confirms that viral sequences shared between the blood and fecal samples of patients with Crohn's disease are more abundant than in healthy subjects, suggesting that a barrier defect facilitates intestinal phage translocation.

During flowering plant reproduction, microspores develop a prominent vacuole before entering pollen mitosis I (PMI), a critical checkpoint where defective microspores are selectively eliminated via programmed cell death (PCD). However, the mechanism governing the vacuole-driven PCD remains enigmatic. Here, we identify CELL DEATH1 (CED1), a fasciclin I (FAS1)-domain-containing protein, as a suppressor of premature microspore PCD. CED1 is predominantly expressed in pollen mother cells (PMCs) and tetrads, with the encoded protein localizing to the plasma membrane and endomembranes. ced1 microspores exhibit defective vacuole biogenesis, vesicle deacidification, nuclear arrest, and cell clearance with hallmarks of PCD. CED1 physically interacts with the subunit c of vacuolar H⁺-ATPase (VHA-c), where it sustains V-ATPase activity, and the overexpression of VHA-c decreases pollen viability. Our work reveals that CED1 safeguards pollen development and may suppress vacuole-driven PCD in association with VHA regulation, elucidating a quality control mechanism in pollen development.

Maternal diet is critical in shaping neonatal metabolism and long-term health by governing breast milk composition. Although bile acids are present in breast milk, their functional role in infant development is not well understood. We identify enteromammary trafficking as the primary source of milk bile acids and show that this pool is modifiable by maternal diet. We also find that maternal bile acids regulate infant growth and levels of the growth-promoting hormone insulin-like growth factor 1 (IGF-1). Remarkably, maternal bile acid sequestration completely prevents excess weight gain in offspring nursed by dams on a high-fat diet. Supplementation with a bile acid or an agonist of the bile acid receptor TGR5 restores growth. Furthermore, TGR5-deficient pups phenocopy the maternal sequestration phenotype, supporting the model that maternal milk bile acids activate neonatal TGR5 to promote infant growth. Altogether, these findings reveal milk bile acids as active metabolic signals with potential for nutritional intervention in early-life programming.

Browning of white adipose tissue (WAT) contributes to the sustained hypermetabolism observed in patients with burns. How glycogen metabolism in WAT is linked to burn-induced hypermetabolism remains unknown. We discover that burn-induced UCP1 expression in subcutaneous WAT is accompanied by elevation of glycogen synthase 1 (GYS1). Adipose tissue-specific deletion of Gys1 suppresses burn-induced UCP1 expression. Gys1 deletion inhibits WAT lipolysis and mitigates hepatic steatosis. Mechanistically, the effects of Gys1 deletion on burn-induced hypermetabolism are mediated by an increase in uridine diphosphate glucose (UDPG), the substrate of GYS1. Both Gys1 deletion and UDPG administration attenuate signaling of interleukin-6. UDPG directly interacts with JAK2 and inhibits STAT3 phosphorylation. Administration of MZ-101, a small-molecule inhibitor of GYS1, suppresses post-burn hypermetabolism and improves the survival rate of mice. Our findings uncover the regulatory role of the GYS1-UDPG-JAK2-STAT3 cascade in WAT during post-burn hypermetabolism and underscore the potential of GYS1 inhibition as a therapeutic strategy for burn injury.