Optimization of heterologous protein production process using genetically engineered Ogataea minuta toward industrial-scale manufacturing.

We have developed the methylotrophic yeast Ogataea minuta as a useful host for producing heterologous proteins. In this study, a double mutant that lacks the Prb1 protease and alcohol oxidase was generated and applied for heterologous protein production. Upon our optimization of the fermentation conditions, such as feeding of carbon and nitrogen sources and pH control, this mutant showed increased production of human serum albumin, resulting in a yield of approximately 7.

Single-cell glycome and transcriptome profiling uncovers the glycan signature of each cell subpopulation of human iPSC-derived neurons.

Human induced pluripotent stem cell (iPSC)-derived neurons are often heterogeneous, posing challenges for disease modeling and cell therapy. We previously developed single-cell glycan and RNA sequencing (scGR-seq) to analyze the glycome and transcriptome simultaneously. Here, we applied scGR-seq to examine heterogeneous populations of human iPSC-derived neurons.

Gut mucin fucosylation dictates the entry of botulinum toxin complexes.

Botulinum toxins (BoNTs) are the most potent known bacterial toxins. The BoNT complex from B-Okra (large progenitor toxin complex (L-PTC)/B, hyper-oral-toxic) exerts at least 80-fold higher oral toxicity in mice compared with that from serotype A1 (L-PTC/A, non-hyper-oral-toxic). Here, we showed that L-PTC/B was predominantly absorbed through enterocytes, whereas L-PTC/A targeted intestinal microfold cells.

Tumor-expressed GPNMB orchestrates Siglec-9 TAM polarization and EMT to promote metastasis in triple-negative breast cancer.

Metastasis remains the leading cause of cancer-related mortality, driven by complex interactions within the tumor microenvironment (TME). Tumor-associated macrophages (TAMs) play a pivotal role in metastatic progression, yet their molecular diversity and upstream regulators remain poorly defined. Glycoprotein nonmetastatic melanoma protein B (GPNMB), overexpressed in subsets of tumors including triple-negative breast cancer (TNBC), is implicated in epithelial-mesenchymal transition (EMT) and cancer stemness.

Induction of intercellular interaction and cell fusion by cell-penetrating peptide-conjugated lipids.

Cell fusion is widely used in biomedical applications. However, conventional cell fusion methods, including polyethylene glycol (PEG)-based methods, often exhibit low efficiency. In this study, we investigated an alternative strategy to induce cell fusion with higher efficiency using transactivator of transcription peptide-conjugated PEG-lipids (Tat-PEG-lipids) with lauroyl chains (C12).

Development of a monomeric recombinant Butea monosperma agglutinin as a diagnostic and prognostic biomarker for cholangiocarcinoma.

Native Butea monosperma agglutinin (nBMA), is a lectin isolated from the seeds of the Butea monosperma plant, which binds specifically to galactose, N-acetylgalactosamine, and lactose. This study developed a recombinant β-chain of BMA (rBMA) expressed in Escherichia coli. The rBMA exists in a monomeric form, retains native structure and sugar-binding capacity without exhibiting hemagglutination activity.

Effects of pyruvate administration on mRNA expression of inflammatory cytokines in adipose tissue and whole-body glucose metabolism in male mice.

Pyruvate administration leads to the accumulation of intracellular lactate in adipocytes and affects inflammatory cytokine production and whole-body glucose metabolism. Therefore, the purpose of this study was to determine whether pyruvate administration improves the dysfunction of glucose metabolism induced by high-fat diet (HFD) intake. In an acute experiment, intraperitoneal injection of male mice with sodium pyruvate (1 g/kg body weight) increased pyruvate and lactate concentrations in blood and epididymal white adipose tissue (eWAT).

Analytical dissection of minor glycoforms and glycoprotein associations in rAAV preparations by multimodal glycoproteomics.

Accurate glycan analysis of viral vectors is essential for evaluating pharmaceutical quality. Recent advances in mass spectrometry-based analytical technologies have achieved glycosylation detection in adeno-associated viruses (AAVs). However, because only a minor subpopulation (< 1%) of recombinant AAV (rAAV) particles may carry glycans or associate with glycoproteins, distinguishing genuine AAV glycosylation from that of co-purified glycoproteins remains technically challenging, highlighting the need for analytical strategies that minimize glycan misassignment and reliably identify glycoprotein interactions.

ARID1A mutation drives gastric tumorigenesis via activating type 2 immune dominant microenvironment.

is a frequently mutated gene in gastric cancers (GCs), particularly in those associated with the Epstein-Barr virus (EBV), which also often shows mutations and silencing. However, the role of these alterations in the development of GC remains unclear. Here, using Cre; ; ; LSL- mice (APP mice), we found that deletion alone promoted a type 2 immune microenvironment marked by the infiltration of type 2 innate lymphoid cells (ILC2s), eosinophils, mast cells, and M2 macrophages via triggering aberrant IL-33-expressing pit lineage differentiation in stem/progenitor cells.

The sulfation pattern of glycosaminoglycans in human brain development and neurological disorders such as Alzheimer's disease.

Glycosaminoglycans (GAGs) are modified by various sulfotransferases and endosulfatases. The resulting sulfation patterns are formed, influencing numerous functions. Sulfation leads to a strong negative charge on GAGs, inducing specific interactions with proteins such as signaling ligands and pathogenicity factors, impacting cellular functions and disease onset.

Targeting Siglec-10/α3β1 Integrin Interactions Enhances Macrophage-Mediated Phagocytosis of Pancreatic Cancer.

Tumor-associated macrophages (TAMs) in the pancreatic ductal adenocarcinoma (PDAC) tumor microenvironment (TME) exhibit immunosuppressive phenotypes and impaired phagocytic activity, facilitating tumor progression and immune evasion. Here, we identify integrin α3β1, composed of ITGA3 and ITGB1 subunits, as a sialylated glycoprotein ligand for Siglec-10, an inhibitory glyco-immune checkpoint receptor highly expressed on TAMs in PDAC. The interaction between Siglec-10 on TAMs and α3β1 on PDAC cells suppresses macrophage-mediated phagocytosis, thereby promoting immune evasion.

RaptGen-UI: an integrated platform for exploring and analyzing the sequence landscape of HT-SELEX experiments.

Summary: RaptGen-UI provides intuitive graphical user-interface of the system exploring and analyzing the sequence landscape of high-throughput (HT)-SELEX (Systematic Evolution of Ligands by EXponential enrichment) experiments through machine learning-driven visualization with optimization capabilities. This software enables wet-lab researchers to efficiently analyze HT-SELEX dataset and optimize RNA aptamers without requiring extensive computational expertise. The containerized architecture ensures secure local deployment and supports both of high-performance Graphics Processing Unit (GPU) acceleration and CPU-only environments, making it suitable for various research settings.

Glycophenotyping of mutants of by lectin microarray.

We previously identified a gene cluster of strain Shirota (YIT 9029) for cell surface long-chain polysaccharides (LCPS-1) biosynthesis, which modulates YIT 9029 activity to induce cytokine production in immune cells, and showed that a lectin microarray can be useful for distinguishing the profile of bacterial cell-surface polysaccharide (PS) structures. Therefore, we isolated disruptive mutant strains of 51 genes predicted to be involved in cell wall PS biosynthesis in YIT 9029. Their binding profiles to lectins in conjunction with their binding abilities to YIT 9029-specific monoclonal antibody (MAb) were compared.

Time-of-day effect of high-intensity muscle contraction on mTOR signaling and protein synthesis in mice.

Resistance exercise promotes muscle protein synthesis by activating mechanistic target of rapamycin (mTOR). The magnitude of muscle hypertrophy might differ depending on the timing of resistance exercise, but the molecular mechanism remains unclear. We aimed to define whether the time of day when muscles are contracted affects mTOR signaling and muscle protein synthesis.

The E3 ubiquitin ligase, RNF219, suppresses CNOT6L expression to exhibit antiproliferative activity.

Despite the increasing evidence of the role of CCR4-NOT complex in posttranscriptional gene regulation, relatively little is known about its mode of action. In a search for novel CCR4-NOT interacting partners, we carried out mass spectrometry analysis of immunoprecipitates with antibodies against four different CCR4-NOT subunits and identified RNF219, ring finger protein 219. A pull-down assay revealed that the C-terminal part of RNF219 directly binds to the CNOT1 DUF3819 domain and is associated with ubiquitin ligase activity.

Investigation on the Substrate Specificity of Serine Protease Neuropsin by Molecular Dynamics Simulation and Marmoset Gene Atlas (MGA).

Serine protease neuropsin is highly expressed in the central nervous system, which regulates brain cognitive function through its proteolytic activity. Neuropsin cleaves heparan sulfate proteoglycan bound to neuregulin-1 (NRG-1) at three specific sites, liberating the ligand domain of NRG-1, which, in turn, induces activation of ErbB4 to release GABA from parvalbumin-expressing interneurons. Little, however, is known about the selective substrate-cleavage mechanism of neuropsin.

A virtual system-coupled molecular dynamics simulation free from experimental knowledge on binding sites: Application to RNA-ligand binding free-energy landscape.

Ligand-receptor docking simulation is difficult when the biomolecules have high intrinsic flexibility. If some knowledge on the ligand-receptor complex structure or inter-molecular contact sites are presented in advance, the difficulty of docking problem considerably decreases. This paper proposes a generalized-ensemble method "cartesian-space division mD-VcMD" (or CSD-mD-VcMD), which calculates stable complex structures without assist of experimental knowledge on the complex structure.

LM-GlycoRepo Version 1.0: A Novel Repository System for Mouse Tissue Glycome Mapping Data.

Lectin microarray (LMA) is a high-sensitive profiling method of protein glycosylation. The increasing use of this method in many studies has led to a growing demand for a repository system that meets the FAIR data principles (Findable, Accessible, Interoperable, and Reusable). Herein, we present a novel repository system, "LM-GlycoRepo," for lectin-based multimodal (LM) data, including LMA data, in accordance with the international guideline MIRAGE (Minimum Information Required for a Glycomics Experiment).

Characterization of glycoside hydrolases involved in xyloglucan degradation in the thermophilic bacterium Thermotoga maritima.

Xyloglucan, a plant hemicellulosic polysaccharide, has a β-glucan main chain and complex side chains composed of sugars such as xylose, galactose and fucose. In this study, we identified xyloglucan degradation-related enzymes in the thermophilic bacterium Thermotoga maritima. TmCel74, belonging to glycoside hydrolase (GH) family 74, was found to be an endo-processive-type xyloglucanase that degraded xyloglucan into xyloglucan oligosaccharides and was able to cleave the β-glucan main chain at both unbranched and xylosylated glucosyl residues.

Isoflurane activates the type 1 ryanodine receptor to induce anesthesia in mice.

Inhaled anesthetics were first introduced into clinical use in the 1840s. Molecular and transgenic animal studies indicate that inhaled anesthetics act through several ion channels, including γ-aminobutyric acid type A receptors (GABAARs) and two-pore domain K+ (K2P) channels, but other targets may mediate anesthetic effects. Mutations in the type 1 ryanodine receptor (RyR1), which is a calcium release channel on the endoplasmic reticulum membrane, are relevant to malignant hyperthermia, a condition that can be induced by inhaled anesthetics.

Photo-controllable cytotoxicity in cell culture using a diarylethene photoswitch.

For diarylethene (DAE) derivatives, whose dark cytotoxicity varies greatly depending on their isomerization state, we observe that the non-toxic open-ring isomer taken up by cells is converted to a toxic closed-ring isomer after only 10 seconds of UV light irradiation. This enables these derivatives to cause pronounced cell death after 1 day. Furthermore, when the closed-ring isomer is detoxified by opening the ring with green light irradiation after several intervals, cell damage increases with the period that DAE remains in the closed-ring state.

Development of on-chip cell domes using Ca-alginate hydrogel shells for non-adherent cell studies.

Cell domes are hemispherical microstructures comprising hydrogel shells that enclose cells within their cavities. They are approximately 500 and 300 μm in radius and height, respectively. Multiple domes can be fabricated in an array on a single glass plate to facilitate optical observations and provide a localised and stable environment for non-adherent cell studies.

RT-4M: Real-Time Mosaicing Manager for Manual Microscopy System.

The creation of virtual slides, i.e., high-resolution digital images of biological samples, is expensive, and existing manual methods often suffer from stitching errors and additional reimaging costs.

CrypTothML: An Integrated Mixed-Solvent Molecular Dynamics Simulation and Machine Learning Approach for Cryptic Site Prediction.

Cryptic sites, which are transient binding sites that emerge through protein conformational changes upon ligand binding, are valuable targets for drug discovery, particularly for allosteric modulators. However, identifying these sites remains challenging because they are often discovered serendipitously when both ligand-binding (holo) and ligand-free (apo) states are experimentally determined. Here, we introduce CrypTothML, a novel framework that integrates mixed-solvent molecular dynamics (MSMD) simulations and machine learning to predict cryptic sites accurately.