Stay in touch with the endoplasmic reticulum

Science China Life Sciences, Год журнала: 2024, Номер 67(2), С. 230 - 257

Опубликована: Янв. 4, 2024

Язык: Английский

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Cell membranes sustain phospholipid imbalance via cholesterol asymmetry

Cell, Год журнала: 2025, Номер unknown

Опубликована: Апрель 1, 2025

Язык: Английский

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Last step in the path of LDL cholesterol from lysosome to plasma membrane to ER is governed by phosphatidylserine

Proceedings of the National Academy of Sciences, Год журнала: 2020, Номер 117(31), С. 18521 - 18529

Опубликована: Июль 20, 2020

Significance Feedback control of cholesterol metabolism is essential for cell viability and prevention heart attacks. Cells acquire from receptor-mediated uptake low-density lipoprotein, which delivers to lysosomes. To exert feedback control, must reach the endoplasmic reticulum (ER). Here we use a CRISPR screen show that lysosome-derived moves first plasma membrane then ER. The last movement requires an enzyme produces phosphatidylserine. This demonstrates transmembrane one lipid (cholesterol) another (phosphatidylserine). Our results explain how organelle (ER) monitors content (plasma membrane), thereby maintaining integrity ensuring survival.

Язык: Английский

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Molecular basis of accessible plasma membrane cholesterol recognition by the GRAM domain of GRAMD1b

The EMBO Journal, Год журнала: 2021, Номер 40(6)

Опубликована: Фев. 19, 2021

Article19 February 2021Open Access Source DataTransparent process Molecular basis of accessible plasma membrane cholesterol recognition by the GRAM domain GRAMD1b Bilge Ercan orcid.org/0000-0002-3780-7946 Lee Kong Chian School Medicine, Nanyang Technological University, SingaporeThese authors contributed equally to this work Search for more papers author Tomoki Naito orcid.org/0000-0002-8393-3601 Dylan Hong Zheng Koh orcid.org/0000-0002-5252-6309 Singapore Dennis Dharmawan Yasunori Saheki Corresponding Author [email protected] orcid.org/0000-0002-1229-6668 Institute Resource Development and Analysis, Kumamoto Kumamoto, Japan Information Ercan1, Naito1, Koh1, Dharmawan1 *,1,2 1Lee 2Institute *Corresponding author. Tel: +65 6592 3996; Fax: 6515 0417; E-mail: The EMBO Journal (2021)40:e106524https://doi.org/10.15252/embj.2020106524 PDFDownload PDF article text main figures. Peer ReviewDownload a summary editorial decision including letters, reviewer comments responses feedback. ToolsAdd favoritesDownload CitationsTrack CitationsPermissions ShareFacebookTwitterLinked InMendeleyWechatReddit Figures & Info Abstract Cholesterol is essential cell physiology. Transport "accessible" pool from (PM) endoplasmic reticulum (ER) ER-localized GRAMD1 proteins (GRAMD1a/1b/1c) contributes homeostasis. However, how cells detect within PM remains unclear. We show that GRAMD1b, coincidence detector anionic lipids, phosphatidylserine (PS), cholesterol, possesses distinct but synergistic sites sensing lipids. find mutation associated with intellectual disability in humans specifically impairs sensing. In addition, we identified another point enhances sensitivity without altering its PS sensitivity. Cell-free reconstitution cell-based assays revealed ability sense regulates tethering determines rate transport GRAMD1b. Thus, codistribution lipids fine-tune non-vesicular ER via GRAMD1s. SYNOPSIS are (ER)-anchored sterol transporters facilitate extraction ER. Analysis molecular determinants lipid reveals which it modulates ER-PM transport. their co-distribution PM. R189W, humans, recognize cholesterol. A single glycine residue (G187) critical determining newly (G187L) makes hypersensitive may serve as tool tracking Introduction Sterol serves major building block cellular membranes eukaryotes. metazoans, represents approximately 20% total therefore structural integrity physiology, regulation signaling pathways (van Meer et al, 2008; Vance, 2015). Cells either synthesize de novo or acquire external sources, primarily through receptor-mediated endocytosis low-density lipoproteins (LDLs) (Goldstein Brown, Up 90% concentrated (PM), where almost half bilayer (Ray 1969; Duve, 1971; Lange 1989; Maxfield Mondal, 2006; Ikonen, van 2008). must constantly monitor levels adjust biosynthesis uptake maintain This mediated, at least part, regulated between ER, regulatory element binding (SREBPs) network controls rates LDL located 1990; Brown 2018). Sterols, transported various transport, independent trafficking (Urbani Simoni, Heino 2000; Hao 2002; Baumann 2005; Normally, majority forms complexes other sphingomyelin phospholipids, sequestered inaccessible intracellular (also known "chemically inactive") (Radhakrishnan McConnell, Ohvo-Rekila McConnell Radhakrishnan, 2003; 2004; Sokolov 2010; 2013; Das 2014; Gay 2015; Chakrabarti 2017). small fraction (~15% lipids) unsequestered active") (Das 2014). Increases level beyond certain threshold (or liberation sequestration) result transient expansions respond rapidly transporting expanded suppress activities SREBPs, shutting down uptake, thereby avoiding overaccumulation maintaining (Slotte Bierman, 1988; Steck, 1997; Scheek Infante Artificially trapping results constitutive activation SREBPs dysregulated metabolism (Infante 2017; Johnson 2019). Despite importance, accessibility regulate still poorly understood. Identifying will provide important insights into metabolism. extends throughout cytoplasm, forming physical contacts virtually all compartments, (Phillips Voeltz, 2016; Wu These contact play roles many aspects exchange/delivery specific (e.g., cholesterol) transfer (Elbaz Schuldiner, 2011; Lev, 2012; Drin, Holthuis Menon, Gatta Murley De Camilli, 2017a,b; Antonny 2018; Luo Jeyasimman Saheki, 2019; Nishimura Stefan, Petrungaro Kornmann, Wong Lees Reinisch, 2020; Meng Prinz 2020). others recently found family evolutionarily conserved ER-anchored proteins, GRAMD1s/Asters (GRAMD1a/Aster-A, GRAMD1b/Aster-B, GRAMD1c/Aster-C) (the Lam/Ltc yeast), mediate sites, contributing homeostasis (Gatta Sandhu Marek GRAMD1s possess an N-terminal domain, PH-like (Begley Tong 2018), cholesterol-harboring StART-like C-terminal transmembrane anchors protein Importantly, acts unique detector, detecting presence both (Naito 2019), include (PS) (a acidic phospholipid PM) (Leventis Grinstein, 2010). Further, facilitates cholesterol-dependent recruitment when transiently expands then extract can directly capture across cytoplasm Horenkamp Jentsch Khelashvili absence GRAMD1s, inefficiently resulting chronic expansion Ferrari Interestingly, recent human genetic studies, genome-wide association have links neurodevelopmental disorders such schizophrenia (Schizophrenia Working Group Psychiatric Genomics C, Reuter Santos-Cortez Thyme Notably, missense (R189W) was consanguineous moderate disability, indicating importance function (Reuter precise mechanism domains impact disability-associated has on remain unknown. originally membrane-associated myotubularin phosphatases (MTMs) (Doerks 2000). Crystal structures myotubularin-related MTMR2 Begley 2006) Lam6/Ltc1, homolog yeast (Tong together high degree similarity PH often binds phosphoinositides, membranes. overall structure comprised typical β-sandwich fold, consists two seven-stranded antiparallel β-sheets followed α-helix additional short case Lam6/Ltc1 been reported bind specificity PI(3,5)P2 PI5P, (Berger Tsujita Lorenzo 2005), PI(4,5)P2 PI4P, GRAMD2 paralog does not contain domain) (Besprozvannaya these currently It also unclear most efficiently Because conserved, elucidating key mechanisms sensed study, explore detects GRAMD1-mediated Our physically proximal lipid-sensing one dedicated Together, they close proximity, detection PS, becomes prominent during increases Remarkably, (R189W), reduces affecting affinity toward dramatically impairing established cell-free assay using purified near full-length artificial used demonstrate R189W failed tether less compared wild-type (WT) proteins. Accordingly, re-expression WT mutant triple knockout restore proper SREBP-2 abnormal accumulation Finally, performed mini-mutagenesis screen position 187 Converting hydrophobic leucine made enhanced GRAMD1b-dependent cholesterol-sensing property GRAMD1-dependent Collectively, our extent codistributed inner leaflet use information Results phospholipids sphingomyelin. formation sequestration making (GRAM1b) enriched PM), 2019) suggests some codistributes being membrane. Such platform GRAM1b amount To confirm codistribution, Förster resonance energy (FRET) assay. Liposomes containing amounts were incubated ECFP-D4H (an biosensor) (Maekawa Fairn, 2015) mVenus-Lact-C2 (Yeung 2008), FRET ECFP mVenus measured 525 nm. Close proximity (1–10 nm) liposomes would increase signal excited 430 nm (Fig 1A). Strikingly, strong observed (50%) (20%) simultaneously incorporated [i.e. condition allows robustly interact EV1A)] 1B C). Additional incorporation (25%) reduced signals increased fluorescence (Figs EV1B As control, only phosphatidylcholine (PC), (50%), examined; no any conditions C), demonstrating signals. consistent be modulated sphingomyelin, normally sequesters pools Figure 1. Schematic vitro mixed biosensor (ECFP-D4H) (mVenus-Lact-C2). emission due recorded Representative spectra mixtures ECFP-D4H, mVenus-Lact-C2, indicated compositions Note (DOPS) decreased (SM) additionally liposomes. Asterisks indicate positions maximum (*477 (**525 nm). DOPC, (1,2-dioleoyl-sn-glycero-3-phosphocholine); DOPS, (1,2-dioleoyl-sn-glycero-3-phospho-L-serine); Chol., cholesterol; SM, (N-oleoyl-D-erythro-sphingosylphosphorylcholine). Quantification ratio 480 (FRET/ECFP) (see Materials Methods) shown (B) (mean ± SEM, n = 4 experiments; Dunnett's multiple comparisons test, **P < 0.0001). Left: Time course normalized mCherry signal, assessed TIRF microscopy, (TKO) HeLa expressing mCherry-tagged (mCherry-GRAM1b). pre-incubated control imaging buffer (for control) (3 µM) masking 30 min 37°C imaged same conditions. Sphingomyelinase (SMase) treatment (100 mU/ml) indicated. Right: Values ΔF/F0 corresponding end experiment arrow [mean 32 (Control), 35 (Pre-incubation w/ ECFP-D4H); data pooled experiments each condition; two-tailed unpaired Student's t-test, 0.0001]. EGFP EGFP-tagged (EGFP-GRAM1b) mCherry-Lact-C2 masking. SMase 53 (mCherry), (mCherry-Lact-C2 OE); four Schematics showing interaction before after sphingomyelinase treatment. At rest, subthreshold limited PM, sufficient induce Liberation (SM)-sequestered leads outer leaflets (as flip-flop leaflets), concomitant inducing Download figure PowerPoint Click here expand figure. EV1. Liposome sedimentation (accessible (phosphatidylserine biosensor). mole% shown. Bound [pellet, (P)] separated unbound [supernatant, (S)], run SDS–PAGE visualized colloidal blue staining 3 experiments). affect cholesterol-containing pool, decrease Fig 1B. (20%), (DOPC) (30%), untagged Lact-C2 upon addition increasing mixture. (C) conditions). D4H (E) concentration-dependent manner EV1-EV5). If occurs should affected masking/trapping incubation overexpression (Raghupathy EV2. basic patch A. Domain organization comparison homolog, Lam6/Ltc1. B. Sequence alignment secondary prediction selected homologs. ESPript 3.0 align amino acid sequences annotate information. Secondary based crystal (PDB: 5YQR). K161 R189 residues red asterisks. White letters background: strict identity; Red letters: group; Blue frames: groups. D. logos graphical representations sequence homologs regions around (D) (n 106 aligned sequences). Error bars approximate Bayesian 95% confidence interval. Black: residues; Green: neutral Blue: Red: residues. conservation different species. EV3. Cholesterol-sensing TKO stably expressed (EGFP-GRAMD1b) constructs loading [the cholesterol/methyl-β-cyclodextrin (MCD) complex (200 µM)] (EGFP), 31 (EGFP-GRAMD1b), 27 [EGFP-GRAMD1b (R189W)], 26 (R189W/R191A)], See Movie Confocal images live (control) transfected biosensor, treated mU/ml 1 h 37°C) imaging. Insets higher magnification white dashed boxes. very weak mCherry-GRAM1b cells. Scale bars, 10 µm. EV4. G187L versions EGFP-GRAM1b methyl-β-cyclodextrin (5 mM) value 75 time point. ΔF/F75 70 41 (WT), 40 (G187L), C. confocal image (WT). cultured medium supplemented 10% lipoprotein-deficient serum (LPDS) mevastatin (50 16 An inset shows region box. recruitment. D–F. (S

Язык: Английский

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New Insights Into Targeting Membrane Lipids for Cancer Therapy

Frontiers in Cell and Developmental Biology, Год журнала: 2020, Номер 8

Опубликована: Сен. 2, 2020

Modulation of membrane lipid composition and organization is currently developing as an effective therapeutic strategy against a wide range diseases, including cancer. This field, known membrane-lipid therapy, has risen from new discoveries on the complex lipids between proteins in plasma membranes. Membrane microdomains present all eukaryotic cells, rafts, have been recognized important concentrating platform for protein receptors involved regulation intracellular signaling, apoptosis, redox balance immune response. The difference cellular membranes healthy cells tumor allows development novel therapies based targeting cancer to increase sensitivity chemotherapeutic agents consequently defeat multidrug resistance. In current manuscript strategies influencing cholesterol/sphingolipids content will be presented together with innovative ones, more focused changing biophysical properties bilayer without affecting its constituents.

Язык: Английский

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Cholesterol transport between cellular membranes: A balancing act between interconnected lipid fluxes

Developmental Cell, Год журнала: 2021, Номер 56(10), С. 1430 - 1436

Опубликована: Май 1, 2021

Язык: Английский

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Intracellular Cholesterol Synthesis and Transport

Frontiers in Cell and Developmental Biology, Год журнала: 2022, Номер 10

Опубликована: Март 21, 2022

Cholesterol homeostasis is related to multiple diseases in humans, including cardiovascular disease, cancer, and neurodegenerative hepatic diseases. The cholesterol levels cells are balanced dynamically by uptake, biosynthesis, transport, distribution, esterification, export. In this review, we focus on de novo synthesis, synthesis regulation, intracellular trafficking. addition, the progression of lipid transfer proteins (LTPs) at contact sites between organelles considered.

Язык: Английский

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Hepatic nonvesicular cholesterol transport is critical for systemic lipid homeostasis

Nature Metabolism, Год журнала: 2023, Номер 5(1), С. 165 - 181

Опубликована: Янв. 16, 2023

Язык: Английский

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Aster-dependent nonvesicular transport facilitates dietary cholesterol uptake

Science, Год журнала: 2023, Номер 382(6671)

Опубликована: Ноя. 9, 2023

Intestinal absorption is an important contributor to systemic cholesterol homeostasis. Niemann-Pick C1 Like 1 (NPC1L1) assists in the initial step of dietary uptake, but how moves downstream NPC1L1 unknown. We show that Aster-B and Aster-C are critical for nonvesicular movement enterocytes. Loss diminishes accessible plasma membrane (PM) abolishes Aster recruitment intestinal brush border. Enterocytes lacking Asters accumulate PM endoplasmic reticulum depletion. Aster-deficient mice have impaired protected against diet-induced hypercholesterolemia. Finally, pathway can be targeted with a small-molecule inhibitor manipulate uptake. These findings identify as physiologically pharmacologically tractable node lipid absorption.

Язык: Английский

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Cell Membranes Sustain Phospholipid Imbalance Via Cholesterol Asymmetry

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2023, Номер unknown

Опубликована: Июль 31, 2023

Membranes are molecular interfaces that compartmentalize cells to control the flow of nutrients and information. These functions facilitated by diverse collections lipids, nearly all which distributed asymmetrically between two bilayer leaflets. Most models biomembrane structure function often include implicit assumption these leaflets have similar abundances phospholipids. Here, we show this is generally invalid investigate consequences lipid abundance imbalances in mammalian plasma membranes (PM). Using quantitative lipidomics, discovered cytoplasmic human erythrocyte >50% overabundance phospholipids compared exoplasmic This imbalance enabled an asymmetric interleaflet distribution cholesterol, regulates cellular cholesterol homeostasis. features produce unique functional characteristics, including low PM permeability resting tension leaflet protein localization. largely overlooked aspects membrane asymmetry represent evolution classic paradigms physiology.

Язык: Английский

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