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Advanced Healthcare Materials Nov 2023Liposomes play an important role in the field of drug delivery by virtue of their biocompatibility and versatility as carriers. Stealth liposomes, obtained by surface...
Liposomes play an important role in the field of drug delivery by virtue of their biocompatibility and versatility as carriers. Stealth liposomes, obtained by surface decoration with hydrophilic polyethylene glycol (PEG) molecules, represent an important turning point in liposome technology, leading to significant improvements in the pharmacokinetic profile compared to naked liposomes. Nevertheless, the generation of effective targeted liposomes-a central issue for cancer therapy-has faced several difficulties and clinical phase failures. Active targeting remains a challenge for liposomes. In this direction, a new Super Stealth Immunoliposomes (SSIL2) composed of a PEG-bi-phospholipids derivative is designed that stabilizes the polymer shielding over the liposomes. Furthermore, its counterpart, conjugated to the fragment antigen-binding of trastuzumab (Fab' -PEG-bi-phospholipids), is firmly anchored on the liposomes surface and correctly orients outward the targeting moiety. Throughout this study, the performances of SSIL2 are evaluated and compared to classic stealth liposomes and stealth immunoliposomes in vitro in a panel of cell lines and in vivo studies in zebrafish larvae and rodent models. Overall, SSIL2 shows superior in vitro and in vivo outcomes, both in terms of safety and anticancer efficacy, thus representing a step forward in targeted cancer therapy, and valuable for future development.
Topics: Animals; Liposomes; Zebrafish; Drug Delivery Systems; Phospholipids; Neoplasms; Polyethylene Glycols
PubMed: 37590033
DOI: 10.1002/adhm.202301650 -
Handbook of Experimental Pharmacology 2024The transmembrane protein 16 (TMEM16) family consists of Ca-activated chloride channels and phospholipid scramblases. Ten mammalian TMEM16 proteins, TMEM16A-K (with no... (Review)
Review
The transmembrane protein 16 (TMEM16) family consists of Ca-activated chloride channels and phospholipid scramblases. Ten mammalian TMEM16 proteins, TMEM16A-K (with no TMEM16I), and several non-mammalian TMEM16 proteins, such as afTMEM16 and nhTMEM16, have been discovered. All known TMEM16 proteins are homodimeric proteins containing two subunits. Each subunit consists of ten transmembrane helices with Ca-binding sites and a single ion-permeation/phospholipid transport pathway. The ion-permeation pathway and the phospholipid transport pathway of TMEM16 proteins have a wide intracellular vestibule, a narrow neck, and a smaller extracellular vestibule. Interestingly, the lining wall of the ion-permeation/phospholipid transport pathway may be formed, at least partially, by membrane phospholipids, though the degree of pore-wall forming by phospholipids likely varies among TMEM16 proteins. Thus, the biophysical properties and activation mechanisms of TMEM16 proteins could differ from each other accordingly. Here we review the current understanding of the structure and function of TMEM16 molecules.
Topics: Humans; Animals; Chloride Channels; Phospholipid Transfer Proteins; Anoctamins; Ion Transport; Phospholipids; Calcium; Mammals
PubMed: 35792944
DOI: 10.1007/164_2022_595 -
Neuron Jan 2024Astrocytes play crucial roles in regulating neural circuit function by forming a dense network of synapse-associated membrane specializations, but signaling pathways...
Astrocytes play crucial roles in regulating neural circuit function by forming a dense network of synapse-associated membrane specializations, but signaling pathways regulating astrocyte morphogenesis remain poorly defined. Here, we show the Drosophila lipid-binding G protein-coupled receptor (GPCR) Tre1 is required for astrocytes to establish their intricate morphology in vivo. The lipid phosphate phosphatases Wunen/Wunen2 also regulate astrocyte morphology and, via Tre1, mediate astrocyte-astrocyte competition for growth-promoting lipids. Loss of s1pr1, the functional analog of Tre1 in zebrafish, disrupts astrocyte process elaboration, and live imaging and pharmacology demonstrate that S1pr1 balances proper astrocyte process extension/retraction dynamics during growth. Loss of Tre1 in flies or S1pr1 in zebrafish results in defects in simple assays of motor behavior. Tre1 and S1pr1 are thus potent evolutionarily conserved regulators of the elaboration of astrocyte morphological complexity and, ultimately, astrocyte control of behavior.
Topics: Animals; Astrocytes; Drosophila; Drosophila Proteins; Phospholipids; Receptors, G-Protein-Coupled; Sphingosine-1-Phosphate Receptors; Zebrafish
PubMed: 38096817
DOI: 10.1016/j.neuron.2023.11.008 -
Blood Jul 2023The development of pathogenic antibody inhibitors against coagulation factor VIII (FVIII) occurs in ∼30% of patients with congenital hemophilia A receiving FVIII...
The development of pathogenic antibody inhibitors against coagulation factor VIII (FVIII) occurs in ∼30% of patients with congenital hemophilia A receiving FVIII replacement therapy, as well as in all cases of acquired hemophilia A. KM33 is an anti-C1 domain antibody inhibitor previously isolated from a patient with severe hemophilia A. In addition to potently blocking FVIII binding to von Willebrand factor and phospholipid surfaces, KM33 disrupts FVIII binding to lipoprotein receptor-related protein 1 (LRP1), which drives FVIII hepatic clearance and antigen presentation in dendritic cells. Here, we report on the structure of FVIII bound to NB33, a recombinant derivative of KM33, via single-particle cryo-electron microscopy. Structural analysis revealed that the NB33 epitope localizes to the FVIII residues R2090-S2094 and I2158-R2159, which constitute membrane-binding loops in the C1 domain. Further analysis revealed that multiple FVIII lysine and arginine residues, previously shown to mediate binding to LRP1, dock onto an acidic cleft at the NB33 variable domain interface, thus blocking a putative LRP1 binding site. Together, these results demonstrate a novel mechanism of FVIII inhibition by a patient-derived antibody inhibitor and provide structural evidence for engineering FVIII with reduced LRP1-mediated clearance.
Topics: Humans; Factor VIII; Hemophilia A; Cryoelectron Microscopy; Protein Domains; von Willebrand Factor; Hemostatics
PubMed: 37192299
DOI: 10.1182/blood.2023020181 -
EMBO Reports Oct 2023SLC25A51 is a member of the mitochondrial carrier family (MCF) but lacks key residues that contribute to the mechanism of other nucleotide MCF transporters. Thus, how...
SLC25A51 is a member of the mitochondrial carrier family (MCF) but lacks key residues that contribute to the mechanism of other nucleotide MCF transporters. Thus, how SLC25A51 transports NAD across the inner mitochondrial membrane remains unclear. To elucidate its mechanism, we use Molecular Dynamics simulations to reconstitute SLC25A51 homology models into lipid bilayers and to generate hypotheses to test. We observe spontaneous binding of cardiolipin phospholipids to three distinct sites on the exterior of SLC25A51's central pore and find that mutation of these sites impairs cardiolipin binding and transporter activity. We also observe that stable formation of the required matrix gate is controlled by a single salt bridge. We identify binding sites in SLC25A51 for NAD and show that its selectivity for NAD is guided by an electrostatic interaction between the charged nicotinamide ring in the ligand and a negatively charged patch in the pore. In turn, interaction of NAD with interior residue E132 guides the ligand to dynamically engage and weaken the salt bridge gate, representing a ligand-induced initiation of transport.
Topics: Cardiolipins; Ligands; Mitochondria; Mitochondrial Membranes; NAD; Humans
PubMed: 37575034
DOI: 10.15252/embr.202256596 -
BioRxiv : the Preprint Server For... Aug 2023ATG2 proteins facilitate bulk lipid transport between membranes. ATG2 is an essential autophagy protein, but ATG2 also localizes to lipid droplets (LDs), and genetic...
ATG2 proteins facilitate bulk lipid transport between membranes. ATG2 is an essential autophagy protein, but ATG2 also localizes to lipid droplets (LDs), and genetic depletion of ATG2 increases LD numbers while impairing fatty acid transport from LDs to mitochondria. How ATG2 supports LD homeostasis and whether lipid transport regulates this homeostasis remains unknown. Here we demonstrate that ATG2 is preferentially recruited to phospholipid monolayers such as those surrounding LDs rather than to phospholipid bilayers. , ATG2 can drive phospholipid transport from artificial LDs with rates that correlate with the binding affinities, such that phospholipids are moved much more efficiently when one of the ATG2-interacting structures is an artificial LD. ATG2 is thought to exhibit 'bridge-like" lipid transport, with lipids flowing across the protein between membranes. We mutated key amino acids within the bridge to form a transport-dead ATG2 mutant (TD-ATG2A) which we show specifically blocks bridge-like, but not shuttle-like, lipid transport . TD-ATG2A still localizes to LDs, but is unable to rescue LD accumulation in ATG2 knockout cells. Thus, ATG2 has a natural affinity for, and an enhanced activity upon LD surfaces and uses bridge-like lipid transport to support LD dynamics in cells.
PubMed: 37645754
DOI: 10.1101/2023.08.14.553257 -
Nature Structural & Molecular Biology Apr 2024Class C G-protein-coupled receptors (GPCRs) are activated through binding of agonists to the large extracellular domain (ECD) followed by rearrangement of the...
Class C G-protein-coupled receptors (GPCRs) are activated through binding of agonists to the large extracellular domain (ECD) followed by rearrangement of the transmembrane domains (TMDs). GPR156, a class C orphan GPCR, is unique because it lacks an ECD and exhibits constitutive activity. Impaired GPR156-G signaling contributes to loss of hearing. Here we present the cryo-electron microscopy structures of human GPR156 in the G-free and G-coupled states. We found that an endogenous phospholipid molecule is located within each TMD of the GPR156 dimer. Asymmetric binding of Gα to the phospholipid-bound GPR156 dimer restructures the first and second intracellular loops and the carboxy-terminal part of the elongated transmembrane 7 (TM7) without altering dimer conformation. Our findings reveal that GPR156 is a transducer for phospholipid signaling. Constant binding of abundant phospholipid molecules and the G-protein-induced reshaping of the cytoplasmic face provide a basis for the constitutive activation of GPR156.
Topics: Humans; Cryoelectron Microscopy; Receptors, G-Protein-Coupled; Signal Transduction; GTP-Binding Proteins; Phospholipids
PubMed: 38332368
DOI: 10.1038/s41594-024-01224-7 -
Experimental & Molecular Medicine Aug 2023Intracellular calcium (Ca) and phosphoinositides (PIPs) are crucial for regulating cellular activities such as metabolism and cell survival. Cells maintain precise... (Review)
Review
Intracellular calcium (Ca) and phosphoinositides (PIPs) are crucial for regulating cellular activities such as metabolism and cell survival. Cells maintain precise intracellular Ca and PIP levels via the actions of a complex system of Ca channels, transporters, Ca ATPases, and signaling effectors, including specific lipid kinases, phosphatases, and phospholipases. Recent research has shed light on the complex interplay between Ca and PIP signaling, suggesting that elevated intracellular Ca levels negatively regulate PIP signaling by inhibiting the membrane localization of PIP-binding proteins carrying specific domains, such as the pleckstrin homology (PH) and Ca-independent C2 domains. This dysregulation is often associated with cancer and metabolic diseases. PIPs recruit various proteins with PH domains to the plasma membrane in response to growth hormones, which activate signaling pathways regulating metabolism, cell survival, and growth. However, abnormal PIP signaling in cancer cells triggers consistent membrane localization and activation of PIP-binding proteins. In the context of obesity, an excessive intracellular Ca level prevents the membrane localization of the PIP-binding proteins AKT, IRS1, and PLCδ via Ca-PIPs, contributing to insulin resistance and other metabolic diseases. Furthermore, an excessive intracellular Ca level can cause functional defects in subcellular organelles such as the endoplasmic reticulum (ER), lysosomes, and mitochondria, causing metabolic diseases. This review explores how intracellular Ca overload negatively regulates the membrane localization of PIP-binding proteins.
Topics: Humans; Phosphatidylinositols; Calcium; Calcium Signaling; Cell Membrane; Insulin Resistance
PubMed: 37524877
DOI: 10.1038/s12276-023-01067-0 -
BioEssays : News and Reviews in... Sep 2023The photocycle of visual opsins is essential to maintain the light sensitivity of the retina. The early physical observations of the rhodopsin photocycle by Böll and... (Review)
Review
The photocycle of visual opsins is essential to maintain the light sensitivity of the retina. The early physical observations of the rhodopsin photocycle by Böll and Kühne in the 1870s inspired over a century's worth of investigations on rhodopsin biochemistry. A single photon isomerizes the Schiff-base linked 11-cis-retinylidene chromophore of rhodopsin, converting it to the all-trans agonist to elicit phototransduction through photoactivated rhodopsin (Rho*). Schiff base hydrolysis of the agonist is a key step in the photocycle, not only diminishing ongoing phototransduction but also allowing for entry and binding of fresh 11-cis chromophore to regenerate the rhodopsin pigment and maintain light sensitivity. Many challenges have been encountered in measuring the rate of this hydrolysis, but recent advancements have facilitated studies of the hydrolysis within the native membrane environment of rhodopsin. These techniques can now be applied to study hydrolysis of agonist in other opsin proteins that mediate phototransduction or chromophore turnover. In this review, we discuss the progress that has been made in characterizing the rhodopsin photocycle and the journey to characterize the hydrolysis of its all-trans-retinylidene agonist.
Topics: Humans; Rhodopsin; Photophobia; Retinaldehyde; Retina
PubMed: 37454357
DOI: 10.1002/bies.202300068 -
PLoS Pathogens Jul 2023L. pneumophila propagates in eukaryotic cells within a specialized niche, the Legionella-containing vacuole (LCV). The infection process is controlled by over 330...
L. pneumophila propagates in eukaryotic cells within a specialized niche, the Legionella-containing vacuole (LCV). The infection process is controlled by over 330 effector proteins delivered through the type IV secretion system. In this study, we report that the Legionella MavH effector localizes to endosomes and remodels host actin cytoskeleton in a phosphatidylinositol 3-phosphate (PI(3)P) dependent manner when ectopically expressed. We show that MavH recruits host actin capping protein (CP) and actin to the endosome via its CP-interacting (CPI) motif and WH2-like actin-binding domain, respectively. In vitro assays revealed that MavH stimulates actin assembly on PI(3)P-containing liposomes causing their tubulation. In addition, the recruitment of CP by MavH negatively regulates F-actin density at the membrane. We further show that, in L. pneumophila-infected cells, MavH appears around the LCV at the very early stage of infection and facilitates bacterium entry into the host. Together, our results reveal a novel mechanism of membrane tubulation induced by membrane-dependent actin polymerization catalyzed by MavH that contributes to the early stage of L. pneumophila infection by regulating host actin dynamics.
Topics: Legionella pneumophila; Actins; Polymerization; Phosphatidylinositol Phosphates; Vacuoles; Bacterial Proteins
PubMed: 37463171
DOI: 10.1371/journal.ppat.1011512