Biology subjects

Harayama, T.

Publications and source records attributed to Harayama, T..

12 recordsLinked to original sources

bioRxiv2026

Deletion of Ferritin Heavy Chain Limits Tumor Growth and Promotes Iron-Dependent Stress in Medulloblastoma

Iron is essential for tumor proliferation and metabolic adaptation but becomes cytotoxic when unbuffered, creating a potential metabolic vulnerability. Ferritin, a conserved iron-storage complex, limits labile iron and establishes the upper threshold of iron tolerance in cancer cells. Here, we report the first ferritin heavy chain (FTH) knockout in a brain tumor model system. Although FTH loss was tolerated under basal conditions through adaptive remodeling of iron metabolism, it exposed profound vulnerabilities under iron stress. FTH deficiency lowered the threshold for iron toxicity, sensitizing medulloblastoma (MB) cells to both canonical ferroptosis and a mechanistically distinct iron-dependent cell death pathway. Oxidative iron stress impaired tumor growth and prolonged survival in orthotopic xenografts, whereas vitamin C-induced iron reduction triggered a selective, iron-dependent, but non-ferroptotic elimination of MB-like cells in tumor organoids. Notably, sensitivity to iron toxicity correlated strongly with cellular phenotype, with mesenchymal-like cells displaying greater susceptibility than epithelial-like counterparts. Collectively, these findings identify ferritin as a central regulator of iron tolerance in MB and establish iron toxicity, not via iron deprivation, as a therapeutically exploitable vulnerability. More broadly, this work provides a mechanistic framework for targeting iron metabolism through modulation of ferritin-dependent iron buffering and iron redox homeostasis in cancers. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=170 SRC="FIGDIR/small/739635v1_ufig1.gif" ALT="Figure 1000"> View larger version (53K): org.highwire.dtl.DTLVardef@1c499a4org.highwire.dtl.DTLVardef@431c70org.highwire.dtl.DTLVardef@2b5borg.highwire.dtl.DTLVardef@11e9ac1_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2026

MSLipidMapper: a pathway-centered lipidome analysis environment linking lipid class, acyl-chain subsets, and multi-omics data

Lipids exhibit extensive structural diversity arising from variation in lipid classes, subclasses, and acyl-chain compositions, making systematic interpretation of lipidomics data challenging. Although untargeted lipidomics enables the quantification of hundreds to thousands of lipid molecular species, downstream analyses often treat pathway-level summaries, molecular-species visualization, structural subsetting, and multi-omics interpretation as separate steps. Here, we present MSLipidMapper, an R/Shiny-based lipidomics data exploration environment for pathway-centered and structure-aware analysis of annotated lipidomics datasets. MSLipidMapper reconstructs annotated lipid peak tables as Bioconductor SummarizedExperiment objects, thereby organizing quantitative lipid abundance values, sample metadata, lipid subclass annotations, and parsed acyl-chain features within a unified data structure. Lipid molecular species are summarized on static, curated lipid metabolic pathway maps at the subclass level while retaining direct links to the underlying molecular species and acyl-chain annotations. This design enables users to inspect molecular-species patterns underlying each pathway node, define lipid subsets based on structural features such as specific acyl chains, and re-project these subsets onto the same pathway context. Gene or protein expression data can also be overlaid on pathway-associated reactions to support multi-layer interpretation of lipid metabolism. The program is showcased using publicly available aging lipidome datasets of mice, illustrating how subclass-level pathway summaries can be connected to molecular-species heatmaps, acyl-chain-defined subsets, and transcriptome or proteome information.

bioinformatics↗

bioRxiv2026

Ether lipid remodeling during neuronal differentiation prevents ferroptosis

Ferroptosis is a form of cell death driven by iron-dependent lipid peroxidation, with specific lipid species playing key roles in modulating susceptibility. Among these, ether lipids have shown conflicting effects, being linked to both protection and sensitization. Here, we dissect the relationship between lipid structure and ferroptosis sensitivity and explain how ether lipids exert context-dependent effects. Ether lipids can promote ferroptosis through a metabolic bias towards the accumulation of polyunsaturated acyl chains and ethanolamine head groups, whereas this pro-ferroptotic tendency is counterbalanced by the anti-ferroptotic vinyl ether moiety introduced by plasmanylethanolamine desaturase 1. We show that this protective effect is critical for preventing ferroptosis in hiPSC-derived neurons, which accumulate otherwise pro-ferroptotic ether lipids during differentiation. This effect is not solely due to its antioxidant properties but also stems from the reprogramming of mitochondrial respiration. The lack of vinyl ether bonds leads to multiple mitochondrial defects, including increased mitochondrial reactive oxygen species (ROS), lower membrane potential, and abnormal cristae structures. These findings indicate that vinyl ether bonds in ether lipids offer dual ferroptosis resistance by scavenging ROS and minimizing its production at the mitochondrial level. The disruption of this system in Caenorhabditis elegans leads to iron-induced death and impaired motility. Thus, our study reveals ether lipid structural remodeling as a key regulator of ferroptosis sensitivity in neurons.

cell biology↗

bioRxiv2026

Selective Editing and Functionalization of the Mammalian Lipidome

Lipids exhibit extraordinary molecular diversity, yet tools to selectively manipulate defined lipid classes in living cells are lacking. Here we show that lipid tail structure biases metabolic fate, enabling the design of synthetic lipid analogs with programmable metabolic selectivity. This approach enables selective cellular production of distinct lipid species or subclasses, including types of neutral lipids, phospholipids, sphingolipids, and ether lipids, without genetic or enzymatic perturbation. We further couple metabolic selectivity to chemical functionalization using bifunctional lipids, in which one modification directs metabolic flux and a second enables bioorthogonal tagging. Using this strategy, we achieve selective in situ labeling of different lipid pools in living cells. Together, our work establishes a chemical biology strategy that enables unprecedented precision in modulating, functionalizing, and rewiring the mammalian lipidome.

cell biology↗

bioRxiv2025

Retrospective metabolomics via dual-dimensional deconvolution using ZT Scan DIA 2.0

We present a scanning data-independent acquisition (DIA) strategy, ZT Scan DIA, combined with dual-dimensional tandem mass spectrometry spectral filtering and deconvolution along both the quadrupole and retention time axes to reconstruct compound-specific MS2 spectra from complex mixtures. This approach is particularly effective for hydrophilic metabolomics data, where spectral similarity-based annotation is widely used, increasing annotation rates by 119-193% compared with conventional data-dependent acquisition (DDA) and window-based DIA methods. In lipidomics, deconvolution improved annotation precision by removing contaminant product ions and enabled separate quantification of co-eluting isomers using MS2 chromatograms, although common diagnostic ions could also be erroneously removed. Nevertheless, optimization of analysis parameters minimized this negative effect. Furthermore, we developed a practical data processing pipeline in which raw ZT Scan DIA-MS2 chromatograms are directly used for isomer separation and MS2-based quantification, covering 1,393 and 3,020 molecules for human plasma and mouse liver tissues, respectively. All data processing steps, including direct import of vendor raw data, are supported in MS-DIAL.

bioengineering↗

bioRxiv2025

Very long chain fatty acids drive 1-deoxy-Sphingolipid toxicity

1-deoxy-sphingolipids (1-deoxySLs) are atypical sphingolipids synthesized by the serine palmitoyltransferase (SPT) when L-alanine is used instead of its canonical substrate L-serine. Increased 1-deoxySLs are associated with sensory neuropathies such as Hereditary Sensory and Autonomic Neuropathy type 1 (HSAN1) and diabetic polyneuropathy (DPN). Despite their known cellular, mitochondrial, and neurotoxic effects, the mechanisms underlying their toxicity remain poorly understood. Using a CRISPR interference (CRISPRi) screening approach, we identified CERS2, ELOVL1, ACACA, HSD17B12, and PTPLB as key mediators of 1-deoxySL-induced toxicity. All genes are integral to the biosynthesis of very long-chain (VLC) fatty acids and VLC-ceramides. We validated these findings through genetic knockdown experiments, cytotoxicity assays, and stable isotope-resolved lipidomics via LC-MS/MS. Pharmacological inhibition of ELOVL1 using a preclinical tested compound alleviated the cellular, mitochondrial, and neuronal toxicity induced by 1-deoxySLs. Supplementation experiments combining 1-deoxySLs with various VLC fatty acids revealed that 1-deoxyDHceramide conjugated to nervonic acid (m18:0/24:1) is the principal toxic specie. Further mechanistic studies showed that m18:0/24:1 induces apoptosis through the mitochondrial permeability transition pore (mPTP) formation. Inhibition of BAX or blocking mPTP formation with cyclosporin A effectively prevented toxicity. In conclusion, our findings demonstrate that 1-deoxyDHCeramides conjugated to nervonic acid are the primary mediators of 1-deoxySL toxicity, acting through mitochondrial dysfunction and BAX-dependent apoptosis.

biochemistry↗

bioRxiv2024

Seipin Regulates Caveolin-1 Trafficking and Organelle Crosstalk

Caveolin-1 (CAV1), the main structural component of caveolae, is essential in various biological processes, including mechanotransduction, lipid metabolism, and endocytosis1-4. Deregulation of CAV1 dynamics is linked to various pathologies, including cellular senescence, cancer, insulin resistance, and lipodystrophy5-9. However, mechanisms regulating CAV1 trafficking and function remain poorly understood. Here, we show that seipin, a crucial lipid droplet (LD) biogenesis factor10, modulates CAV1 trafficking. Deletion of seipin resulted in the accumulation of saturated lipids, leading to ceramide and sphingomyelin overproduction, which disrupted the membrane order of the trans-Golgi network (TGN). In seipin deficiency, CAV1 location to the plasma membrane (PM) was impaired, reducing caveolae. Instead, CAV1 accumulated in TGN and late endosome compartments, which fused with LDs and delivered the protein. In wild-type (WT) cells, this process was minimal but significantly enhanced by treatment with palmitate, ceramide, or Stearoyl-CoA desaturase-1 (SCD1) inhibition. Conversely, in seipin-deficient cells, inhibiting Fatty Acid Synthase (FASN) or overexpressing SCD1 restored CAV1 localization to the PM and reduced its accumulation in LDs. Our findings reveal that seipin controls the funneling of palmitate toward glycerolipids synthesis and storage in LDs versus conversion to ceramides in the ER. This balance is crucial to cellular protein trafficking by controlling the TGN membrane order. Therefore, our study identifies seipin as a critical regulator of cellular lipid metabolism, protein trafficking, and organelle homeostasis. These findings shed light on the processes regulating CAV1 trafficking and show that convergent pathophysiological mechanisms associated with defects in CAV1 and seipin contribute to metabolic disorders, including insulin resistance and lipodystrophies11-14.

cell biology↗

bioRxiv2024

MS-DIAL 5 multimodal mass spectrometry data mining unveils lipidome complexities

Lipidomics and metabolomics communities comprise various informatics tools; however, software programs that can handle multimodal mass spectrometry (MS) data with structural annotations guided by the Lipidomics Standards Initiative are limited. Here, we provide MS-DIAL 5 to facilitate the in-depth structural elucidation of lipids through electron-activated dissociation (EAD)-based tandem MS, as well as determine their molecular localization through MS imaging (MSI) data using a species/tissue-specific lipidome database containing the predicted collision-cross section (CCS) values. With the optimized EAD settings using 14 eV kinetic energy conditions, the program correctly delineated the lipid structures based on EAD-MS/MS data from 96.4% of authentic standards. Our workflow was showcased by annotating the sn- and double-bond positions of eye-specific phosphatidylcholine molecules containing very-long-chain polyunsaturated fatty acids (VLC-PUFAs), characterized as PC n-3-VLC-PUFA/FA. Using MSI data from the eye and HeLa cells supplemented with n-3-VLC-PUFA, we identified glycerol 3-phosphate (G3P) acyltransferase (GPAT) as an enzyme candidate responsible for incorporating n-3 VLC-PUFAs into the sn-1 position of phospholipids in mammalian cells, which was confirmed using recombinant proteins in a cell-free system. Therefore, the MS-DIAL 5 environment, combined with optimized MS data acquisition methods, facilitates a better understanding of lipid structures and their localization, offering novel insights into lipid biology.

bioinformatics↗

bioRxiv2024

Acyltransferases in the first step of glycerophospholipid synthesis have redundant and non-redundant roles in sn-1 acyl chain regulation, ether lipid levels, and cell survival

The increased use of lipidomics analyses in biomedical research made it crucial to understand correctly how lipid compositions are regulated. Lipid acyl chains are important regulators of membrane physicochemical properties, and are incorporated by various acyltransferases. The acyltransferases implicated in the first step of glycerophospholipid synthesis, the GPATs and GNPAT, have distinct localizations and substrate preferences. This suggests that they have distinct roles on lipid regulation, but a complete understanding of their redundant and non-redundant functions is missing. We report here a comprehensive analysis of cells having mutations in GPATs and GNPAT, either alone or in combinations. Our results suggest that the balance between GPATs and GNPAT affect the levels of ether lipids, together with glycerophospholipid acyl chain length and unsaturation. In addition, we found that lipid synthesis initiated at peroxisomes, but not mitochondria, is sufficient to provide the lipid synthesis flux required for normal cell growth. Our study unveils the multifaceted roles of the first step of de novo glycerophospholipid synthesis, thus leading to a better understanding of how lipidomes are shaped.

cell biology↗

bioRxiv2022

Optical Control of Membrane Fluidity Modulates Protein Secretion

The lipid composition of cellular membranes is dynamic and undergoes remodelling affecting biophysical properties, such as membrane fluidity, which are critical to biological function. Here, we introduce an optical approach to manipulate membrane fluidity based on exogenous synthetic fatty acid with an azobenzene photoswitch, termed FAAzo4. Cells rapidly incorporate FAAzo4 into phosphatidylcholine (PC), the major phospholipid in mammalian cells, in a concentration- and cell type-dependent manner. This generates photoswitchable PC analogs (AzoPC), which are predominantly located in the endoplasmic reticulum (ER). Irradiation causes a rapid photoisomerization that increases membrane fluidity with high spatiotemporal precision. We use these PhotoCells to study the impact of membrane mechanics on protein export from the ER and demonstrate that this two-step process has distinct membrane fluidity requirements. Our approach represents an unprecedented way of manipulating membrane fluidity in cellulo and opens novel avenues to probe roles of fluidity in a wide variety of biological processes.

cell biology↗

bioRxiv2020

Establishment of a highly efficient gene disruption strategy to analyze and manipulate lipid co-regulatory networks

Gene disruption has been dramatically facilitated by genome editing tools. Despite improvements in gene disruption rates in cultured cells, clone isolation remains routinely performed to obtain mutants, potentially leading to artifacts due to clonal variation in cellular phenotypes. Here we report GENF, a highly efficient strategy to disrupt genes without isolating clones, which can be multiplexed. Using it, we obtained reliable lipidomics datasets from mutant cells without being affected by variances related to clone isolation. Through this, we found that an enzyme involved in congenital generalized lipodystrophy regulates glycerophospholipids with specific acyl-chains. We also demonstrate the possibility to dissect complex lipid co-regulatory mechanisms, explaining cell adaptations to altered lipid metabolism. With its simplicity and the avoidance of cloning-related artifacts, GENF is likely to contribute to many cell biology studies, especially those involving -omics approaches.

genetics↗

bioRxiv2020

Mutagenesis and homology modeling reveal a predicted pocket of lysophosphatidylcholine acyltransferase 2 to catch Acyl-CoA.

Platelet-activating factor (PAF) is a potent proinflammatory phospholipid mediator that elicits various cellular functions and promotes several pathological conditions, including anaphylaxis and neuropathic pain. PAF is biosynthesized by two types of lyso-PAF acetyltransferases: lysophosphatidylcholine acyltransferase 1 (LPCAT1) and LPCAT2, which are constitutive and inducible forms of lyso-PAF acetyltransferase, respectively. Because LPCAT2 mainly produces PAF under inflammatory conditions, understanding the structure of LPCAT2 is important for developing specific drugs against PAF-related inflammatory diseases. Although the structure of LPCAT2 has not been determined, the crystal structure was reported for Thermotoga maritima PlsC, an enzyme in the same enzyme family as LPCAT2. In this study, we identified residues in mouse LPCAT2 essential for its enzymatic activity and a potential acyl-coenzyme A (CoA)-binding pocket, based on homology modeling of mouse LPCAT2 with PlsC. We also found that Ala115 of mouse LPCAT2 was important for acyl-CoA selectivity. In conclusion, these results predict the structure of mouse LPCAT2. Our findings have implications for the future development of new drugs against PAF-related diseases.

biochemistry↗