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The International Journal of... Sep 2022Membrane phospholipids are critical components of several signaling pathways. Maintained in a variety of asymmetric distributions, their trafficking across the membrane... (Review)
Review
Membrane phospholipids are critical components of several signaling pathways. Maintained in a variety of asymmetric distributions, their trafficking across the membrane can be induced by intra-, extra-, and intercellular events. A familiar example is the externalization of phosphatidylserine from the inner leaflet to the outer leaflet in apoptosis, inducing phagocytosis of the soma. Recently, it has been recognized that phospholipids in the axonal membrane may be a signal for axonal degeneration, regeneration, or other processes. This review focuses on key recent developments and areas for ongoing investigations. KEY FACTS: Phosphatidylserine externalization propagates along an axon after axonal injury and is delayed in the Wallerian degeneration slow (Wld) mutant. The ATP8A2 flippase mutant has spontaneous axonal degeneration. Microdomains of axonal degeneration in spheroid bodies have differential externalization of phosphatidylserine and phosphatidylethanolamine. Phospholipid trafficking could represent a mechanism for coordinated axonal degeneration and elimination, i.e. axoptosis, analogous to apoptosis of the cell body.
Topics: Axons; Humans; Neurons; Phosphatidylserines; Phospholipids; Wallerian Degeneration
PubMed: 35868612
DOI: 10.1016/j.biocel.2022.106264 -
Theranostics 2020Cancer is a leading cause of mortality and morbidity worldwide. Despite major improvements in current therapeutic methods, ideal therapeutic strategies for improved... (Review)
Review
Cancer is a leading cause of mortality and morbidity worldwide. Despite major improvements in current therapeutic methods, ideal therapeutic strategies for improved tumor elimination are still lacking. Recently, immunotherapy has attracted much attention, and many immune-active agents have been approved for clinical use alone or in combination with other cancer drugs. However, some patients have a poor response to these agents. New agents and strategies are needed to overcome such deficiencies. Phosphatidylserine (PS) is an essential component of bilayer cell membranes and is normally present in the inner leaflet. In the physiological state, PS exposure on the external leaflet not only acts as an engulfment signal for phagocytosis in apoptotic cells but also participates in blood coagulation, myoblast fusion and immune regulation in nonapoptotic cells. In the tumor microenvironment, PS exposure is significantly increased on the surface of tumor cells or tumor cell-derived microvesicles, which have innate immunosuppressive properties and facilitate tumor growth and metastasis. To date, agents targeting PS have been developed, some of which are under investigation in clinical trials as combination drugs for various cancers. However, controversial results are emerging in laboratory research as well as in clinical trials, and the efficiency of PS-targeting agents remains uncertain. In this review, we summarize recent progress in our understanding of the physiological and pathological roles of PS, with a focus on immune suppressive features. In addition, we discuss current drug developments that are based on PS-targeting strategies in both experimental and clinical studies. We hope to provide a future research direction for the development of new agents for cancer therapy.
Topics: Animals; Apoptosis; Cell Membrane; Humans; Immunotherapy; Neoplasms; Phosphatidylserines; Tumor Microenvironment
PubMed: 32802188
DOI: 10.7150/thno.45125 -
Proceedings of the National Academy of... Aug 2023The membrane contact site ER/PM junctions are hubs for signaling pathways, including Ca signaling. Phosphatidylserine (PtdSer) mediates various physiological functions;...
The membrane contact site ER/PM junctions are hubs for signaling pathways, including Ca signaling. Phosphatidylserine (PtdSer) mediates various physiological functions; however, junctional PtdSer composition and the role of PtdSer in Ca signaling and Ca-dependent gene regulation are not understood. Here, we show that STIM1-formed junctions are required for PI(4)P/PtdSer exchange by ORP5 and ORP8, which have reciprocal lipid exchange modes and function as a rheostat that sets the junctional PtdSer/PI(4)P ratio. Targeting the ORP5 and ORP8 and their lipid transfer ORD domains to PM subdomains revealed that ORP5 sets low and ORP8 high junctional PI(4)P/PtdSer ratio that controls STIM1-STIM1 and STIM1-Orai1 interaction and the activity of the SERCA pump to determine the pattern of receptor-evoked Ca oscillations, and consequently translocation of NFAT to the nucleus. Significantly, targeting the ORP5 and ORP8 ORDs to the STIM1 ER subdomain reversed their function. Notably, changing PI(4)P/PtdSer ratio by hydrolysis of PM or ER PtdSer with targeted PtdSer-specific PLA1a1 reproduced the ORPs function. The function of the ORPs is determined both by their differential lipid exchange modes and by privileged localization at the ER/PM subdomains. These findings reveal a role of PtdSer as a signaling lipid that controls the available PM PI(4)P, the unappreciated role of ER PtdSer in cell function, and the diversity of the ER/PM junctions. The effect of PtdSer on the junctional PI(4)P level should have multiple implications in cellular signaling and functions.
Topics: Phosphatidylserines; Signal Transduction; Cell Nucleus; Hydrolysis; Mitochondrial Membranes
PubMed: 37607230
DOI: 10.1073/pnas.2301410120 -
Journal For Immunotherapy of Cancer Dec 2023CD1d is a monomorphic major histocompatibility complex class I-like molecule that presents lipid antigens to distinct T-cell subsets and can be expressed by various...
BACKGROUND
CD1d is a monomorphic major histocompatibility complex class I-like molecule that presents lipid antigens to distinct T-cell subsets and can be expressed by various malignancies. Antibody-mediated targeting of CD1d on multiple myeloma cells was reported to induce apoptosis and could therefore constitute a novel therapeutic approach.
METHODS
To determine how a CD1d-specific single-domain antibody (VHH) enhances binding of the early apoptosis marker annexin V to CD1d tumor cells we use in vitro cell-based assays and CRISPR-Cas9-mediated gene editing, and to determine the structure of the VHH1D17-CD1d(endogenous lipid) complex we use X-ray crystallography.
RESULTS
Anti-CD1d VHH1D17 strongly enhances annexin V binding to CD1d tumor cells but this does not reflect induction of apoptosis. Instead, we show that VHH1D17 enhances presentation of phosphatidylserine (PS) in CD1d and that this is saposin dependent. The crystal structure of the VHH1D17-CD1d(endogenous lipid) complex demonstrates that VHH1D17 binds the A'-pocket of CD1d, leaving the lipid headgroup solvent exposed, and has an electro-negatively charged patch which could be involved in the enhanced PS presentation by CD1d. Presentation of PS in CD1d does not trigger phagocytosis but leads to greatly enhanced binding of T-cell immunoglobulin and mucin domain containing molecules (TIM)-1 to TIM-3, TIM-4 and induces TIM-3 signaling.
CONCLUSION
Our findings reveal the existence of an immune modulatory CD1d(PS)-TIM axis with potentially unexpected implications for immune regulation in both physiological and pathological conditions.
Topics: Humans; Hepatitis A Virus Cellular Receptor 2; Single-Domain Antibodies; Phosphatidylserines; Annexin A5; T-Lymphocyte Subsets
PubMed: 38040419
DOI: 10.1136/jitc-2023-007631 -
Cell Reports Nov 2022In "healthy" tumor cells, phosphatidylserine (PS) is predominately localized in the inner plasma membrane leaflet. During apoptosis, PS relocates to the outer leaflet....
In "healthy" tumor cells, phosphatidylserine (PS) is predominately localized in the inner plasma membrane leaflet. During apoptosis, PS relocates to the outer leaflet. Herein, we established PS tumor models with tumor cells lacking PS flippase component CDC50A, constantly exposing PS but alive. PS tumors developed bigger than wild-type (WT) tumors, featuring M2 polarized tumor-associated macrophages (TAMs) and fewer tumor-antigen-specific T cells. The PS receptor TIM-3 is responsible for PS recognition. Employing an opposite tumor model, PS, with tumor cells lacking the PS scramblase Xkr8 and unable to expose PS during otherwise normal apoptosis, we find that the accumulated apoptotic tumor cells produce and release cyclic GAMP (cGAMP) to immune cells to activate the STING pathway, leading to TAM M1 polarization, suppressed interleukin (IL)-10 secretion, and natural killer (NK) cell cytotoxicity. Silencing Xkr8 in vivo by either short hairpin RNA (shRNA) or small interfering RNA (siRNA) to achieve a PS externalization blockade provides robust therapeutic anti-tumor efficiency.
Topics: Humans; Phosphatidylserines; Phospholipids; Cell Membrane; Apoptosis; Neoplasms; Immunotherapy
PubMed: 36323258
DOI: 10.1016/j.celrep.2022.111582 -
Methods in Molecular Biology (Clifton,... 2021Recently, urinary extracellular vesicles (EVs) have garnered interest as a potential source of noninvasive biomarkers of diseases related to urinary organs (kidney,...
Recently, urinary extracellular vesicles (EVs) have garnered interest as a potential source of noninvasive biomarkers of diseases related to urinary organs (kidney, bladder, urethra, and prostate).Ultracentrifugation is considered the gold standard method for isolation of EVs. However, the precipitates after ultracentrifugation steps are usually contaminated with soluble proteins, such as the Tamm-Horsfall protein (uromodulin).Therefore, ultracentrifugation on a sucrose-deuterium oxide (DO) cushion for purer EV isolation is performed to remove these proteins. In addition, as a nonultracentrifugation method for EV isolation, we have also adopted the phosphatidylserine (PS) affinity method, which is a novel method for EV purification using the T-cell immunoglobulin domain and the mucin domain-containing protein 4 (Tim4).Here, we describe an ultracentrifugation protocol based on a sucrose-DO cushion and the PS affinity method protocol for the isolation of urinary EVs.
Topics: Biomarkers; Deuterium Oxide; Extracellular Vesicles; Humans; Phosphatidylserines; Sucrose; Ultracentrifugation; Urinalysis; Urine Specimen Collection
PubMed: 33651361
DOI: 10.1007/978-1-0716-1354-2_15 -
Cell Communication and Signaling : CCS Feb 2020Immunotherapy for cancer is making impressive strides at improving survival of a subset of cancer patients. To increase the breadth of patients that benefit from... (Review)
Review
Immunotherapy for cancer is making impressive strides at improving survival of a subset of cancer patients. To increase the breadth of patients that benefit from immunotherapy, new strategies that combat the immunosuppressive microenvironment of tumors are needed. Phosphatidylserine (PS) signaling is exploited by tumors to enhance tumor immune evasion and thus strategies to inhibit PS-mediated immune suppression have potential to increase the efficacy of immunotherapy. PS is a membrane lipid that flips to the outer surface of the cell membrane during apoptosis and/or cell stress. Externalized PS can drive efferocytosis or engage PS receptors (PSRs) to promote local immune suppression. In the tumor microenvironment (TME) PS-mediated immune suppression is often termed apoptotic mimicry. Monoclonal antibodies (mAbs) targeting PS or PSRs have been developed and are in preclinical and clinical testing. The TIM (T-cell/transmembrane, immunoglobulin, and mucin) and TAM (Tyro3, AXL, and MerTK) family of receptors are PSRs that have been shown to drive PS-mediated immune suppression in tumors. This review will highlight the development of mAbs targeting PS, TIM-3 and the TAM receptors. Video Abstract.
Topics: Animals; Antineoplastic Agents, Immunological; Humans; Immunotherapy; Neoplasms; Phosphatidylserines; Receptors, Cell Surface; Tumor Microenvironment
PubMed: 32087708
DOI: 10.1186/s12964-020-0521-5 -
Molecular Cancer May 2016Unlike normal cells, cancer cells express high levels of phosphatidylserine on the extracellular leaflet of their cell membrane. Exploiting this characteristic, our lab... (Review)
Review
Unlike normal cells, cancer cells express high levels of phosphatidylserine on the extracellular leaflet of their cell membrane. Exploiting this characteristic, our lab developed a therapeutic agent that consists of the fusogenic protein, saposin C (SapC) which is embedded in dioleoylphosphatidylserine (DOPS) vesicles. These nanovesicles selectively target cancer cells and induce apoptosis. Here we review the data supporting use of SapC-DOPS to locate tumors for surgical resection or for treatment. In addition, there is important evidence suggesting that SapC-DOPS may also prove to be an effective novel cancer therapeutic reagent. Given that SapC-DOPS is easily labeled with lipophilic dyes, it has been combined with the far-red fluorescent dye, CellVue Maroon (CVM), for tumor targeting studies. We also have used contrast agents incorporated in the SapC-DOPS nanovesicles for computed tomography and magnetic resonance imaging, and review that data here. Administered intravenously, the fluorescently labeled SapC-DOPS traversed the blood-brain tumor barrier enabling identification of brain tumors. SapC-DOPS-CVM also detected a variety of other mouse tumors in vivo, rendering them observable by optical imaging using IVIS and multi-angle rotational optical imaging. Dye is detected within 30 min and remains within tumor for at least 7 days, whereas non-tumor tissues were unstained (some dye observed in the liver was transient, likely representing degradation products). Additionally, labeled SapC-DOPS ex vivo delineated tumors in human histological specimens. SapC-DOPS can also be labeled with contrast reagents for computed tomography or magnetic resonance imaging. In conclusion, labeled SapC-DOPS provides a convenient, specific, and nontoxic method for detecting tumors while concurrently offering a therapeutic benefit.
Topics: Animals; Cell Membrane; Contrast Media; Fluorescent Dyes; Humans; Models, Animal; Molecular Imaging; Multimodal Imaging; Nanoparticles; Neoplasms; Phosphatidylserines; Protein Binding; Saposins
PubMed: 27160923
DOI: 10.1186/s12943-016-0519-1 -
Thrombosis and Haemostasis Dec 2023Although thrombosis events are the leading complication of uremia, their mechanism is largely unknown. The interaction between endothelial cells (ECs) and red blood...
BACKGROUND
Although thrombosis events are the leading complication of uremia, their mechanism is largely unknown. The interaction between endothelial cells (ECs) and red blood cells (RBCs) in uremic solutes and its prothrombotic role need to be investigated.
METHODS AND RESULTS
Here, we established an in vitro co-incubation model of uremic RBC and EC as well as a uremic rat model induced by adenine. Using flow cytometry, confocal microscopy, and electron microscopy, we found increased erythrophagocytosis by EC accompanied by increased reactive oxygen species, lipid peroxidation, and impairment of mitochondria, indicating that ECs undergo ferroptosis. Further investigations showed increased proteins' expression of heme oxygenase-1 and ferritin and labile iron pool accumulation in EC, which could be suppressed by deferoxamine (DFO). The ferroptosis-negative regulators glutathione peroxidase 4 and SLC7A11 were decreased in our erythrophagocytosis model and could be enhanced by ferrostatin-1 or DFO. In vivo, we observed that vascular EC phagocytosed RBC and underwent ferroptosis in the kidney of the uremic rat, which could be inhibited by blocking the phagocytic pathway or inhibiting ferroptosis. Next, we found that the high tendency of thrombus formation was accompanied by erythrophagocytosis-induced ferroptosis in vitro and in vivo. Importantly, we further revealed that upregulated TMEM16F expression mediated phosphatidylserine externalization on ferroptotic EC, which contributed to a uremia-associated hypercoagulable state.
CONCLUSION
Our results indicate that erythrophagocytosis-triggered ferroptosis followed by phosphatidylserine exposure of EC may play a key role in uremic thrombotic complications, which may be a promising target to prevent thrombogenesis of uremia.
Topics: Rats; Animals; Ferroptosis; Endothelial Cells; Phosphatidylserines; Erythrocytes; Thrombosis; Uremia
PubMed: 37364609
DOI: 10.1055/a-2117-7890 -
Nature Communications Jun 2019Various studies have demonstrated that the two leaflets of cellular membranes interact, potentially through so-called interdigitation between the fatty acyl groups.... (Review)
Review
Various studies have demonstrated that the two leaflets of cellular membranes interact, potentially through so-called interdigitation between the fatty acyl groups. While the molecular mechanism underlying interleaflet coupling remains to be fully understood, recent results suggest interactions between the very-long-chain sphingolipids in the outer leaflet, and phosphatidylserine PS18:0/18:1 in the inner leaflet, and an important role for cholesterol for these interactions. Here we review the evidence that cross-linking of sphingolipids may result in clustering of phosphatidylserine and transfer of signals to the cytosol. Although much remains to be uncovered, the molecular properties and abundance of PS 18:0/18:1 suggest a unique role for this lipid.
Topics: Animals; Cell Line; Cell Membrane; Cholesterol; Cross-Linking Reagents; Cytosol; Humans; Lipid Bilayers; Phosphatidylserines; Sphingolipids
PubMed: 31227693
DOI: 10.1038/s41467-019-10711-1