Journal articles: 'Membrane receptor-Ligand interactions'

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Langelaan, David N., and Jan K. Rainey. "Membrane catalysis of peptide–receptor bindingThis paper is one of a selection of papers published in this special issue entitled “Canadian Society of Biochemistry, Molecular & Cellular Biology 52nd Annual Meeting — Protein Folding: Principles and Diseases” and has undergone the Journal's usual peer review process." Biochemistry and Cell Biology 88, no. 2 (April 2010): 203–10. http://dx.doi.org/10.1139/o09-129.

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The membrane catalysis hypothesis states that a peptide ligand activates its target receptor after an initial interaction with the surrounding membrane. Upon membrane binding and interaction, the ligand is structured such that receptor binding and activation is encouraged. As evidence for this hypothesis, there are numerous studies concerning the conformation that peptides adopt in membrane mimetic environments. This mini-review analyzes the features of ligand peptides with an available high-resolution membrane-induced structure and a characterized membrane-binding region. At the peptide–membrane interface, both amphipathic helices and turn structures are commonly formed in peptide ligands and both hydrophobic and electrostatic interactions can be responsible for membrane binding. Apelin is the ligand to the G-protein coupled receptor (GPCR) named APJ, with various important physiological effects, which we have recently characterized both in solution and bound to anionic micelles. The structural changes that apelin undergoes when binding to micelles provide strong evidence for membrane catalysis of apelin–APJ interactions.

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Ledig, Matthias M., Fawaz Haj, John L. Bixby, Andrew W. Stoker, and Bernhard K. Mueller. "The Receptor Tyrosine Phosphatase Crypα Promotes Intraretinal Axon Growth." Journal of Cell Biology 147, no. 2 (October 18, 1999): 375–88. http://dx.doi.org/10.1083/jcb.147.2.375.

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Retinal ganglion cell axons grow towards the optic fissure in close contact with the basal membrane, an excellent growth substratum. One of the ligands of receptor tyrosine phosphatase CRYPα is located on the retinal and tectal basal membranes. To analyze the role of this RPTP and its ligand in intraretinal growth and guidance of ganglion cell axons, we disrupted ligand- receptor interactions on the retinal basal membrane in culture. Antibodies against CRYPα strongly reduced retinal axon growth on the basal membrane, and induced a dramatic change in morphology of retinal growth cones, reducing the size of growth cone lamellipodia. A similar effect was observed by blocking the ligand with a CRYPα ectodomain fusion protein. These effects did not occur, or were much reduced, when axons were grown either on laminin-1, on matrigel or on basal membranes with glial endfeet removed. This indicates that a ligand for CRYPα is located on glial endfeet. These results show for the first time in vertebrates that the interaction of a receptor tyrosine phosphatase with its ligand is crucial not only for promotion of retinal axon growth but also for maintenance of retinal growth cone lamellipodia on basal membranes.

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Behling, Ronald W., and Lynn W. Jelinski. "Importance of the membrane in ligand-receptor interactions." Biochemical Pharmacology 40, no. 1 (July 1990): 49–54. http://dx.doi.org/10.1016/0006-2952(90)90177-m.

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KATZ, A., D. RHODES, and L. HERBETTE. "Role of the membrane bilayer in ligand-receptor interactions." Journal of Molecular and Cellular Cardiology 18 (1986): 12. http://dx.doi.org/10.1016/s0022-2828(86)80522-3.

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Wang, Li, Xin-Pu Hou, Angelica Ottova, and H. Ti Tien. "Receptor–ligand interactions in a reconstituted bilayer lipid membrane." Electrochemistry Communications 2, no. 5 (May 2000): 287–89. http://dx.doi.org/10.1016/s1388-2481(00)00008-4.

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Torres, Manuel, Catalina Ana Rosselló, Paula Fernández-García, Victoria Lladó, Or Kakhlon, and Pablo Vicente Escribá. "The Implications for Cells of the Lipid Switches Driven by Protein–Membrane Interactions and the Development of Membrane Lipid Therapy." International Journal of Molecular Sciences 21, no. 7 (March 27, 2020): 2322. http://dx.doi.org/10.3390/ijms21072322.

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The cell membrane contains a variety of receptors that interact with signaling molecules. However, agonist–receptor interactions not always activate a signaling cascade. Amphitropic membrane proteins are required for signal propagation upon ligand-induced receptor activation. These proteins localize to the plasma membrane or internal compartments; however, they are only activated by ligand-receptor complexes when both come into physical contact in membranes. These interactions enable signal propagation. Thus, signals may not propagate into the cell if peripheral proteins do not co-localize with receptors even in the presence of messengers. As the translocation of an amphitropic protein greatly depends on the membrane’s lipid composition, regulation of the lipid bilayer emerges as a novel therapeutic strategy. Some of the signals controlled by proteins non-permanently bound to membranes produce dramatic changes in the cell’s physiology. Indeed, changes in membrane lipids induce translocation of dozens of peripheral signaling proteins from or to the plasma membrane, which controls how cells behave. We called these changes “lipid switches”, as they alter the cell’s status (e.g., proliferation, differentiation, death, etc.) in response to the modulation of membrane lipids. Indeed, this discovery enables therapeutic interventions that modify the bilayer’s lipids, an approach known as membrane-lipid therapy (MLT) or melitherapy.

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Cao, Shengya, Sean M. Peterson, Sören Müller, Mike Reichelt, Christian McRoberts Amador, and Nadia Martinez-Martin. "A membrane protein display platform for receptor interactome discovery." Proceedings of the National Academy of Sciences 118, no. 39 (September 16, 2021): e2025451118. http://dx.doi.org/10.1073/pnas.2025451118.

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Cell surface receptors are critical for cell signaling and constitute a quarter of all human genes. Despite their importance and abundance, receptor interaction networks remain understudied because of difficulties associated with maintaining membrane proteins in their native conformation and their typically weak interactions. To overcome these challenges, we developed an extracellular vesicle-based method for membrane protein display that enables purification-free and high-throughput detection of receptor–ligand interactions in membranes. We demonstrate that this platform is broadly applicable to a variety of membrane proteins, enabling enhanced detection of extracellular interactions over a wide range of binding affinities. We were able to recapitulate and expand the interactome for prominent members of the B7 family of immunoregulatory proteins such as PD-L1/CD274 and B7-H3/CD276. Moreover, when applied to the orphan cancer-associated fibroblast protein, LRRC15, we identified a membrane-dependent interaction with the tumor stroma marker TEM1/CD248. Furthermore, this platform enabled profiling of cellular receptors for target-expressing as well as endogenous extracellular vesicles. Overall, this study presents a sensitive and easy to use screening platform that bypasses membrane protein purification and enables characterization of interactomes for any cell surface–expressed target of interest in its native state.

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Scheel, Andreas A., Bettina Funsch, Michael Busch, Gabriele Gradl, Johannes Pschorr, and Martin J. Lohse. "Receptor-Ligand Interactions Studied with Homogeneous Fluorescence-Based Assays Suitable for Miniaturized Screening." Journal of Biomolecular Screening 6, no. 1 (February 2001): 11–18. http://dx.doi.org/10.1177/108705710100600103.

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Cell membrane receptors play a central role in controlling cellular functions, making them the target of drugs for a wide variety of diseases. This report describes how a recently developed method, fluorescence intensity distribution analysis (FIDA), can be used to develop homogeneous, nonradioactive high throughput screening assays for membrane receptors. With FIDA, free ligand and ligand accumulated on receptor-bearing membrane vesicles can be distinguished on the basis of their particle brightness. This allows the concentration of both bound and free ligand to be determined reliably from a single measurement, without any separation. We demonstrate that ligand affinity, receptor expression level, and potency of inhibitors can be determined using the epidermal growth factor and β2-adrenergic receptors as model systems. Highly focused confocal optics enable single-molecule sensitivity, and sample volumes can thus be reduced to 1,IL without affecting the quality of the fluorescence signal. Our results demonstrate that FIDA is an ideal method for membrane receptor assays offering substantial benefits for assay development and high throughput pharmaceutical screening.

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Yang, Yun-Hee, and Jwa-Min Nam. "Single Nanoparticle Tracking-Based Detection of Membrane Receptor−Ligand Interactions." Analytical Chemistry 81, no. 7 (April 2009): 2564–68. http://dx.doi.org/10.1021/ac802477h.

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Valenzano, Kenneth J., Wendy Miller, Jared N. Kravitz, Philippe Samama, Dan Fitzpatrick, and Kevin Seeley. "Development of a Fluorescent Ligand-Binding Assay Using the AcroWell Filter Plate." Journal of Biomolecular Screening 5, no. 6 (December 2000): 455–61. http://dx.doi.org/10.1177/108705710000500608.

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One of the most powerful tools for receptor research and drug discovery is the use of receptor-ligand affinity screening of combinatorial libraries. Early work involved the use of radioactive ligands to identify a binding event; however, there are numerous limitations involved in the use of radioactivity for high throughput screening. These limitations have led to the creation of highly sensitive, nonradioactive alternatives to investigate receptor-ligand interactions. Pall Gelman Laboratory has introduced the AcroWell, a patented low-fluorescent-background membrane and sealing process together with a filter plate design that is compatible with robotic systems. Taken together, these allow the AcroWell 96-well filter plate to detect trace quantities of lanthanide-labeled ligands for cell-, bead-, or membrane-based assays using time-resolved fluorescence. Using europium-labeled galanin, we have demonstrated that saturation binding experiments can be performed with low-background fluorescence and signal-to-noise ratios that rival traditional radioisotopic techniques while maintaining biological integrity of the receptor-ligand interaction. In addition, the ability to discriminate between active and inactive compounds in a mock galanin screen is demonstrated with low well-to-well variability, allowing reliable determination of positive hits even for low-affinity interactions.

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Chattopadhyay, Amitabha, Md Jafurulla, and Thomas J. Pucadyil. "Ligand Binding and G-protein Coupling of the Serotonin1A Receptor in Cholesterol-enriched Hippocampal Membranes." Bioscience Reports 26, no. 2 (June 22, 2006): 79–87. http://dx.doi.org/10.1007/s10540-006-9009-9.

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The serotonin1A receptor is the most extensively studied member of the family of seven transmembrane domain G-protein coupled serotonin receptors. Since a large portion of such transmembrane receptors remains in contact with the membrane lipid environment, lipid–protein interactions assume importance in the structure-function analysis of such receptors. We have earlier reported the requirement of cholesterol for serotonin1A receptor function in native hippocampal membranes by specific depletion of cholesterol using methyl- β-cyclodextrin. In this paper, we monitored the serotonin1A receptor function in membranes that are enriched in cholesterol using a complex prepared from cholesterol and methyl-β-cyclodextrin. Our results indicate that ligand binding and receptor/G-protein interaction of the serotonin1A receptor do not exhibit significant difference in native and cholesterol-enriched hippocampal membranes indicating that further enrichment of cholesterol has little functional consequence on the serotonin1A receptor function. These results therefore provide new information on the effect of cholesterol enrichment on the hippocampal serotonin1A receptor function.

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Carbone, Catherine B., Nadja Kern, Ricardo A. Fernandes, Enfu Hui, Xiaolei Su, K. Christopher Garcia, and Ronald D. Vale. "In vitro reconstitution of T cell receptor-mediated segregation of the CD45 phosphatase." Proceedings of the National Academy of Sciences 114, no. 44 (October 17, 2017): E9338—E9345. http://dx.doi.org/10.1073/pnas.1710358114.

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T cell signaling initiates upon the binding of peptide-loaded MHC (pMHC) on an antigen-presenting cell to the T cell receptor (TCR) on a T cell. TCR phosphorylation in response to pMHC binding is accompanied by segregation of the transmembrane phosphatase CD45 away from TCR–pMHC complexes. The kinetic segregation hypothesis proposes that CD45 exclusion shifts the local kinase–phosphatase balance to favor TCR phosphorylation. Spatial partitioning may arise from the size difference between the large CD45 extracellular domain and the smaller TCR–pMHC complex, although parsing potential contributions of extracellular protein size, actin activity, and lipid domains is difficult in living cells. Here, we reconstitute segregation of CD45 from bound receptor–ligand pairs using purified proteins on model membranes. Using a model receptor–ligand pair (FRB–FKBP), we first test physical and computational predictions for protein organization at membrane interfaces. We then show that the TCR–pMHC interaction causes partial exclusion of CD45. Comparing two developmentally regulated isoforms of CD45, the larger RABC variant is excluded more rapidly and efficiently (∼50%) than the smaller R0 isoform (∼20%), suggesting that CD45 isotypes could regulate signaling thresholds in different T cell subtypes. Similar to the sensitivity of T cell signaling, TCR–pMHC interactions with Kds of ≤15 µM were needed to exclude CD45. We further show that the coreceptor PD-1 with its ligand PD-L1, immunotherapy targets that inhibit T cell signaling, also exclude CD45. These results demonstrate that the binding energies of physiological receptor–ligand pairs on the T cell are sufficient to create spatial organization at membrane–membrane interfaces.

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Momin, Noor, Stacey Lee, Avinash K. Gadok, David J. Busch, George D. Bachand, Carl C. Hayden, Jeanne C. Stachowiak, and Darryl Y. Sasaki. "Designing lipids for selective partitioning into liquid ordered membrane domains." Soft Matter 11, no. 16 (2015): 3241–50. http://dx.doi.org/10.1039/c4sm02856b.

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Short PEG spacers effectively decouple headgroup and receptor–ligand interactions from the membrane allowing packing order of the lipid tails to direct partitioning of lipids to specific membrane phases.

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Amin, Divya N., and Gerald L. Hazelbauer. "The Chemoreceptor Dimer Is the Unit of Conformational Coupling and Transmembrane Signaling." Journal of Bacteriology 192, no. 5 (January 8, 2010): 1193–200. http://dx.doi.org/10.1128/jb.01391-09.

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ABSTRACT Transmembrane chemoreceptors are central components in bacterial chemotaxis. Receptors couple ligand binding and adaptational modification to receptor conformation in processes that create transmembrane signaling. Homodimers, the fundamental receptor structural units, associate in trimers and localize in patches of thousands. To what degree do conformational coupling and transmembrane signaling require higher-order interactions among dimers? To what degree are they altered by such interactions? To what degree are they inherent features of homodimers? We addressed these questions using nanodiscs to create membrane environments in which receptor dimers had few or no potential interaction partners. Receptors with many, few, or no interaction partners were tested for conformational changes and transmembrane signaling in response to ligand occupancy and adaptational modification. Conformation was assayed by measuring initial rates of receptor methylation, a parameter independent of receptor-receptor interactions. Coupling of ligand occupancy and adaptational modification to receptor conformation and thus to transmembrane signaling occurred with essentially the same sensitivity and magnitude in isolated dimers as for dimers with many neighbors. Thus, we conclude that the chemoreceptor dimer is the fundamental unit of conformational coupling and transmembrane signaling. This implies that in signaling complexes, coupling and transmembrane signaling occur through individual dimers and that changes between dimers in a receptor trimer or among trimer-based signaling complexes are subsequent steps in signaling.

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Byrne, Patrick O., Kalina Hristova, and Daniel J. Leahy. "EGFR forms ligand-independent oligomers that are distinct from the active state." Journal of Biological Chemistry 295, no. 38 (July 29, 2020): 13353–62. http://dx.doi.org/10.1074/jbc.ra120.012852.

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The human epidermal growth factor receptor (EGFR/ERBB1) is a receptor tyrosine kinase (RTK) that forms activated oligomers in response to ligand. Much evidence indicates that EGFR/ERBB1 also forms oligomers in the absence of ligand, but the structure and physiological role of these ligand-independent oligomers remain unclear. To examine these features, we use fluorescence microscopy to measure the oligomer stability and FRET efficiency for homo- and hetero-oligomers of fluorescent protein-labeled forms of EGFR and its paralog, human epidermal growth factor receptor 2 (HER2/ERBB2) in vesicles derived from mammalian cell membranes. We observe that both receptors form ligand-independent oligomers at physiological plasma membrane concentrations. Mutations introduced in the kinase region at the active state asymmetric kinase dimer interface do not affect the stability of ligand-independent EGFR oligomers. These results indicate that ligand-independent EGFR oligomers form using interactions that are distinct from the EGFR active state.

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Pliska, Vladimir. "Thermodynamic descriptors, profiles and driving forces in membrane receptor-ligand interactions." Journal of Receptors and Signal Transduction 30, no. 6 (October 5, 2010): 454–68. http://dx.doi.org/10.3109/10799893.2010.515594.

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Greene, D’Artagnan, Wesley M. Botello-Smith, Alec Follmer, Li Xiao, Eleftherios Lambros, and Ray Luo. "Modeling Membrane Protein–Ligand Binding Interactions: The Human Purinergic Platelet Receptor." Journal of Physical Chemistry B 120, no. 48 (November 23, 2016): 12293–304. http://dx.doi.org/10.1021/acs.jpcb.6b09535.

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Kim, Keehun, Shayla Paulekas, Fredrik Sadler, Tejas M. Gupte, Michael Ritt, Matthew Dysthe, Nagarajan Vaidehi, and Sivaraj Sivaramakrishnan. "β2-adrenoceptor ligand efficacy is tuned by a two-stage interaction with the Gαs C terminus." Proceedings of the National Academy of Sciences 118, no. 11 (March 8, 2021): e2017201118. http://dx.doi.org/10.1073/pnas.2017201118.

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Classical pharmacological models have incorporated an “intrinsic efficacy” parameter to capture system-independent effects of G protein–coupled receptor (GPCR) ligands. However, the nonlinear serial amplification of downstream signaling limits quantitation of ligand intrinsic efficacy. A recent biophysical study has characterized a ligand “molecular efficacy” that quantifies the influence of ligand-dependent receptor conformation on G protein activation. Nonetheless, the structural translation of ligand molecular efficacy into G protein activation remains unclear and forms the focus of this study. We first establish a robust, accessible, and sensitive assay to probe GPCR interaction with G protein and the Gα C terminus (G-peptide), an established structural determinant of G protein selectivity. We circumvent the need for extensive purification protocols by the single-step incorporation of receptor and G protein elements into giant plasma membrane vesicles (GPMVs). We use previously established SPASM FRET sensors to control the stoichiometry and effective concentration of receptor–G protein interactions. We demonstrate that GPMV-incorporated sensors (v-SPASM sensors) provide enhanced dynamic range, expression-insensitive readout, and a reagent level assay that yields single point measurements of ligand molecular efficacy. Leveraging this technology, we establish the receptor–G-peptide interaction as a sufficient structural determinant of this receptor-level parameter. Combining v-SPASM measurements with molecular dynamics (MD) simulations, we elucidate a two-stage receptor activation mechanism, wherein receptor–G-peptide interactions in an intermediate orientation alter the receptor conformational landscape to facilitate engagement of a fully coupled orientation that tunes G protein activation.

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Mecham, R. P., L. Whitehouse, M. Hay, A. Hinek, and M. P. Sheetz. "Ligand affinity of the 67-kD elastin/laminin binding protein is modulated by the protein's lectin domain: visualization of elastin/laminin-receptor complexes with gold-tagged ligands." Journal of Cell Biology 113, no. 1 (April 1, 1991): 187–94. http://dx.doi.org/10.1083/jcb.113.1.187.

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Video-enhanced microscopy was used to examine the interaction of elastin- or laminin-coated gold particles with elastin binding proteins on the surface of live cells. By visualizing the binding events in real time, it was possible to determine the specificity and avidity of ligand binding as well as to analyze the motion of the receptor-ligand complex in the plane of the plasma membrane. Although it was difficult to interpret the rates of binding and release rigorously because of the possibility for multiple interactions between particles and the cell surface, relative changes in binding have revealed important aspects of the regulation of affinity of ligand-receptor interaction in situ. Both elastin and laminin were found to compete for binding to the cell surface and lactose dramatically decreased the affinity of the receptor(s) for both elastin and laminin. These findings were supported by in vitro studies of the detergent-solubilized receptor. Further, immobilization of the ligand-receptor complexes through binding to the cytoskeleton dramatically decreased the ability of bound particles to leave the receptor. The changes in the kinetics of ligand-coated gold binding to living cells suggest that both laminin and elastin binding is inhibited by lactose and that attachment of receptor to the cytoskeleton increases its affinity for the ligand.

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Fantini, Jacques. "Fundamental Mechanisms in Membrane Receptology: Old Paradigms, New Concepts and Perspectives." Receptors 3, no. 1 (March 18, 2024): 107–21. http://dx.doi.org/10.3390/receptors3010006.

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Receptology, the science of receptors, is a multidimensional field of research which can be dissected into biosynthesis, membrane sorting, ligand binding and signal transduction. Plasma membrane receptors connect the cells with their environment and transmit signals that are translated into biological information. The historical paradigm of ligand–receptor interactions is the lock-and-key model. This model presupposes that both partners have a precise 3D shape that perfectly fits together to form the ligand–receptor complex. However, this simple model suffers from severe limitations due to several levels of simplifications: (i) water molecules and membrane lipids are not considered; (ii) not all ligands have a stable 3D structure; (iii) the ligand-binding pocket of the receptor is often flexible and conformationally rearranged after the initial binding step (induced fit mechanism) and/or subjected to conformational selection by the ligand; (iv) there are signal transduction mechanisms which can be either purely mechanical (conformational change of the receptor induced after binding of the ligand), lipid-assisted (e.g., by raft lipids such as cholesterol or gangliosides), or in some instances of quantic nature (detection of odorant molecules). The aim of the present review is to challenge the old paradigms and present new concepts of membrane receptology that consider the impact of critical parameters such as water molecules, membrane lipids, electrostatic surface potential and quantum mechanisms.

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Lorenz, Bärbel, Rabea Keller, Eva Sunnick, Burkhard Geil, and Andreas Janshoff. "Colloidal probe microscopy of membrane–membrane interactions: From ligand–receptor recognition to fusion events." Biophysical Chemistry 150, no. 1-3 (August 2010): 54–63. http://dx.doi.org/10.1016/j.bpc.2010.02.008.

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Löchte, Sara, Sharon Waichman, Oliver Beutel, Changjiang You, and Jacob Piehler. "Live cell micropatterning reveals the dynamics of signaling complexes at the plasma membrane." Journal of Cell Biology 207, no. 3 (November 10, 2014): 407–18. http://dx.doi.org/10.1083/jcb.201406032.

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Interactions of proteins in the plasma membrane are notoriously challenging to study under physiological conditions. We report in this paper a generic approach for spatial organization of plasma membrane proteins into micropatterns as a tool for visualizing and quantifying interactions with extracellular, intracellular, and transmembrane proteins in live cells. Based on a protein-repellent poly(ethylene glycol) polymer brush, micropatterned surface functionalization with the HaloTag ligand for capturing HaloTag fusion proteins and RGD peptides promoting cell adhesion was devised. Efficient micropatterning of the type I interferon (IFN) receptor subunit IFNAR2 fused to the HaloTag was achieved, and highly specific IFN binding to the receptor was detected. The dynamics of this interaction could be quantified on the single molecule level, and IFN-induced receptor dimerization in micropatterns could be monitored. Assembly of active signaling complexes was confirmed by immunostaining of phosphorylated Janus family kinases, and the interaction dynamics of cytosolic effector proteins recruited to the receptor complex were unambiguously quantified by fluorescence recovery after photobleaching.

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Schrangl, Lukas, Vanessa Mühlgrabner, René Platzer, Florian Kellner, Josephine Wieland, Reinhard Obst, José L. Toca-Herrera, Johannes B. Huppa, Gerhard J. Schütz, and Janett Göhring. "Advanced Quantification of Receptor–Ligand Interaction Lifetimes via Single-Molecule FRET Microscopy." Biomolecules 14, no. 8 (August 13, 2024): 1001. http://dx.doi.org/10.3390/biom14081001.

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Receptor–ligand interactions at cell interfaces initiate signaling cascades essential for cellular communication and effector functions. Specifically, T cell receptor (TCR) interactions with pathogen-derived peptides presented by the major histocompatibility complex (pMHC) molecules on antigen-presenting cells are crucial for T cell activation. The binding duration, or dwell time, of TCR–pMHC interactions correlates with downstream signaling efficacy, with strong agonists exhibiting longer lifetimes compared to weak agonists. Traditional surface plasmon resonance (SPR) methods quantify 3D affinity but lack cellular context and fail to account for factors like membrane fluctuations. In the recent years, single-molecule Förster resonance energy transfer (smFRET) has been applied to measure 2D binding kinetics of TCR–pMHC interactions in a cellular context. Here, we introduce a rigorous mathematical model based on survival analysis to determine exponentially distributed receptor–ligand interaction lifetimes, verified through simulated data. Additionally, we developed a comprehensive analysis pipeline to extract interaction lifetimes from raw microscopy images, demonstrating the model’s accuracy and robustness across multiple TCR–pMHC pairs. Our new software suite automates data processing to enhance throughput and reduce bias. This methodology provides a refined tool for investigating T cell activation mechanisms, offering insights into immune response modulation.

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Vallés, Ana Sofía, and Francisco J. Barrantes. "Interactions between the Nicotinic and Endocannabinoid Receptors at the Plasma Membrane." Membranes 12, no. 8 (August 22, 2022): 812. http://dx.doi.org/10.3390/membranes12080812.

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Compartmentalization, together with transbilayer and lateral asymmetries, provide the structural foundation for functional specializations at the cell surface, including the active role of the lipid microenvironment in the modulation of membrane-bound proteins. The chemical synapse, the site where neurotransmitter-coded signals are decoded by neurotransmitter receptors, adds another layer of complexity to the plasma membrane architectural intricacy, mainly due to the need to accommodate a sizeable number of molecules in a minute subcellular compartment with dimensions barely reaching the micrometer. In this review, we discuss how nature has developed suitable adjustments to accommodate different types of membrane-bound receptors and scaffolding proteins via membrane microdomains, and how this “effort-sharing” mechanism has evolved to optimize crosstalk, separation, or coupling, where/when appropriate. We focus on a fast ligand-gated neurotransmitter receptor, the nicotinic acetylcholine receptor, and a second-messenger G-protein coupled receptor, the cannabinoid receptor, as a paradigmatic example.

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Goldsztein, A., S. Babar, M. Voué, J. De Coninck, J. Conti, J. Marchand-Brynaert, S. Devouge, F. Homblé, and E. Goormaghtigh. "Gastric ATPase phosphorylation/dephosphorylation monitored by new FTIR-based BIA–ATR biosensors." Spectroscopy 24, no. 3-4 (2010): 257–60. http://dx.doi.org/10.1155/2010/793594.

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Biosensors are composite devices suitable for the investigation of receptor–ligand interactions. In this paper we present the specific application to a membrane embedded protein of a new sensor device, so-called BIA–ATR, based on Attenuated Total Reflection–Fourier Transform Infrared (ATR–FTIR) spectroscopy. It consists in a functionalised ATR germanium crystal whose surface has been covalently modified to adsorb a biomembrane. Detection of the ligand–receptor interaction is achieved using FTIR spectroscopy. We report the specific detection of the phosphorylation/dephosphorylation of the H+/K+gastric ATPase. The H+, K+-ATPase is a particularly large protein entity. This glycosylated protein contains more than 1300 residues and is embedded in a lipid membrane. Yet we demonstrate that the BIA–ATR sensor is capable of monitoring the binding of a single phosphate on such a large protein entity. Furthermore, we also demonstrate the potential of the approach to monitor the kinetics of binding and dissociation of the ligand.

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Dämgen, Marc A., and Philip C. Biggin. "State-dependent protein-lipid interactions of a pentameric ligand-gated ion channel in a neuronal membrane." PLOS Computational Biology 17, no. 2 (February 11, 2021): e1007856. http://dx.doi.org/10.1371/journal.pcbi.1007856.

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Pentameric ligand-gated ion channels (pLGICs) are receptor proteins that are sensitive to their membrane environment, but the mechanism for how lipids modulate function under physiological conditions in a state dependent manner is not known. The glycine receptor is a pLGIC whose structure has been resolved in different functional states. Using a realistic model of a neuronal membrane coupled with coarse-grained molecular dynamics simulations, we demonstrate that some key lipid-protein interactions are dependent on the receptor state, suggesting that lipids may regulate the receptor’s conformational dynamics. Comparison with existing structural data confirms known lipid binding sites, but we also predict further protein-lipid interactions including a site at the communication interface between the extracellular and transmembrane domain. Moreover, in the active state, cholesterol can bind to the binding site of the positive allosteric modulator ivermectin. These protein-lipid interaction sites could in future be exploited for the rational design of lipid-like allosteric drugs.

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Zhang, Yudie, Long Li, and Jizeng Wang. "Role of Ligand Distribution in the Cytoskeleton-Associated Endocytosis of Ellipsoidal Nanoparticles." Membranes 11, no. 12 (December 19, 2021): 993. http://dx.doi.org/10.3390/membranes11120993.

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Nanoparticle (NP)–cell interaction mediated by receptor–ligand bonds is a crucial phenomenon in pathology, cellular immunity, and drug delivery systems, and relies strongly on the shape of NPs and the stiffness of the cell. Given this significance, a fundamental question is raised on how the ligand distribution may affect the membrane wrapping of non-spherical NPs under the influence of cytoskeleton deformation. To address this issue, in this work we use a coupled elasticity–diffusion model to systematically investigate the role of ligand distribution in the cytoskeleton-associated endocytosis of ellipsoidal NPs for different NP shapes, sizes, cytoskeleton stiffness, and the initial receptor densities. In this model, we have taken into account the effects of receptor diffusion, receptor–ligand binding, cytoskeleton and membrane deformations, and changes in the configuration entropy of receptors. By solving this model, we find that the uptake process can be significantly influenced by the ligand distribution. Additionally, there exists an optimal state of such a distribution, which corresponds to the fastest uptake efficiency and depends on the NP aspect ratio and cytoskeleton stiffness. We also find that the optimal distribution usually needs local ligand density to be sufficiently high at the large curvature region. Furthermore, the optimal state of NP entry into cells can tolerate slight changes to the corresponding optimal distribution of the ligands. The tolerance to such a change is enhanced as the average receptor density and NP size increase. These results may provide guidelines to control NP–cell interactions and improve the efficiency of target drug delivery systems.

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Aymoz-Bressot, Thibaud, Marie Canis, Florian Meurisse, Anne Wijkhuisen, Benoit Favier, Guillaume Mousseau, Anne Dupressoir, Thierry Heidmann, and Agathe Bacquin. "Cell-Int: a cell–cell interaction assay to identify native membrane protein interactions." Life Science Alliance 7, no. 11 (September 5, 2024): e202402844. http://dx.doi.org/10.26508/lsa.202402844.

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Intercellular protein–protein interactions (PPIs) have pivotal roles in biological functions and diseases. Membrane proteins are therefore a major class of drug targets. However, studying such intercellular PPIs is challenging because of the properties of membrane proteins. Current methods commonly use purified or modified proteins that are not physiologically relevant and hence might mischaracterize interactions occurring in vivo. Here, we describe Cell-Int: a cell interaction assay for studying plasma membrane PPIs. The interaction signal is measured through conjugate formation between two populations of cells each expressing either a ligand or a receptor. In these settings, membrane proteins are in their native environment thus being physiologically relevant. Cell-Int has been applied to the study of diverse protein partners, and enables to investigate the inhibitory potential of blocking antibodies, as well as the retargeting of fusion proteins for therapeutic development. The assay was also validated for screening applications and could serve as a platform for identifying new protein interactors.

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Suenaga, Rieko, Mizuki Takemoto, Asuka Inoue, Ryuichiro Ishitani, and Osamu Nureki. "Lateral access mechanism of LPA receptor probed by molecular dynamics simulation." PLOS ONE 17, no. 2 (February 3, 2022): e0263296. http://dx.doi.org/10.1371/journal.pone.0263296.

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G-protein-coupled receptors (GPCR) are a family of membrane receptors that play important roles in the regulation of various physiological phenomena. LPA receptors (LPA1-6) are members of the class A GPCRs, which transduce a lysophosphatidic acid (LPA) signal across the cell membrane and evoke various responses, including cellular survival, proliferation, differentiation, and migration. The crystal structure of LPA6 revealed a gap between its transmembrane helices (TMs), which is opened toward the membrane side. This led to the proposal of the “lateral access model,” in which its lipophilic ligand directly enters the binding pocket through the gap structure at the membrane. In this study, we performed molecular dynamics (MD) simulations and Markov state model (MSM) analyses of LPA6 and LPA, to elucidate the long timescale dynamics of the ligand binding process. The results from the 71.4-μs MD simulation suggested that the flexibility of the TMs constituting the gap structure enables the lateral entrance of the ligand, and the key interactions between the receptor and ligand facilitate the transition state of the ligand binding process.

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Groomes, Patrice V., Usheer Kanjee, and Manoj T. Duraisingh. "RBC membrane biomechanics and Plasmodium falciparum invasion: probing beyond ligand–receptor interactions." Trends in Parasitology 38, no. 4 (April 2022): 302–15. http://dx.doi.org/10.1016/j.pt.2021.12.005.

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Uebler, Susanne, and Thomas Dresselhaus. "Identifying plant cell-surface receptors: combining ‘classical’ techniques with novel methods." Biochemical Society Transactions 42, no. 2 (March 20, 2014): 395–400. http://dx.doi.org/10.1042/bst20130251.

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Cell–cell communication during development and reproduction in plants depends largely on a few phytohormones and many diverse classes of polymorphic secreted peptides. The peptide ligands are bound at the cell surface of target cells by their membranous interaction partners representing, in most cases, either receptor-like kinases or ion channels. Although knowledge of both the extracellular ligand and its corresponding receptor(s) is necessary to describe the downstream signalling pathway(s), to date only a few ligand–receptor pairs have been identified. Several methods, such as affinity purification and yeast two-hybrid screens, have been used very successfully to elucidate interactions between soluble proteins, but most of these methods cannot be applied to membranous proteins. Experimental obstacles such as low concentration and poor solubility of membrane receptors, as well as instable transient interactions, often hamper the use of these ‘classical’ approaches. However, over the last few years, a lot of progress has been made to overcome these problems by combining classical techniques with new methodologies. In the present article, we review the most promising recent methods in identifying cell-surface receptor interactions, with an emphasis on success stories outside the field of plant research.

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Rogers, Cheryl, and Simon Lemaire. "Characterization of [3H] desmethylimipramine binding in bovine adrenal medulla: interactions with σ- and (or) phencyclidine-receptor ligands." Canadian Journal of Physiology and Pharmacology 70, no. 11 (November 1, 1992): 1508–14. http://dx.doi.org/10.1139/y92-214.

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High-affinity binding sites (apparent KD 2.87 nM) for [3H]desmethylimipramine ([3H]DMI), have been demonstrated and characterized in membrane preparations of bovine adrenal medulla. The binding of [3H]DMI improved upon pretreatment of the membrane with KCl and was saturable, sodium dependent, and potently inhibited by nisoxetine and imipramine. [3H]DMI binding was also inhibited by various phencyclidine (PCP)- and (or) σ-receptor ligands, with the following order of potency: haloperidol > rimcazole > (−)-butaclamol > dextromethorphan > MK-801 > (+)-3-(3-hydroxyphenyl)-N-(1-propyl)piperidine((+)-3-PPP) > PCP > N-(2-thienyl)cyclohexyl-3,4-piperidine (TCP) > (+)-SKF-10047 > (−)-SKF-10047. The inhibition produced by σ ligands was not attributed to stimulation of either σ1- or σ2-receptors, owing to inactivity of the selective σ-receptor ligands (+)-pentazocine and 1,3-di(2-tolyl)guanidine (DTG). The inhibition of [3H]DMI binding by σ- and PCP-receptor ligands was not attributed to PCP1- or PCP2-receptor stimulation, owing to the decreased potency (100-fold) of these ligands in [3H]DMI assays compared with the affinity for brain PCP1 sites, and the ineffectiveness of the PCP2-ligand N-(1-(2-benzo(b)thiophenyl)cyclohexyl)piperidine (BTCP). Scatchard analysis of the inhibition by the σ-ligands haloperidol and (+)-3-PPP, as well as the PCP1 receptor ligand MK-801, demonstrated noncompetitive interaction with the site bound by [3H]DMI. These studies indicate that bovine adrenomedullary membranes possess a specific receptor for the noradrenaline uptake inhibitor [3H]DMI, which is sensitive to allosteric modulation produced by PCP and σ-ligands.Key words: desmethylimipramine, σ-receptor, phencyclidine, noradrenaline uptake, adrenal medulla.

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Gross, Catharina C., Emily Martinez, and Eric O. Long. "Control of NK cell activation by distribution and mobility of ligands on target cells (134.16)." Journal of Immunology 182, no. 1_Supplement (April 1, 2009): 134.16. http://dx.doi.org/10.4049/jimmunol.182.supp.134.16.

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Abstract Natural killer (NK) cells are large granular lymphocytes that comprise 5-15% of peripheral blood mononuclear cells and function through cytokine production or cytotoxicity. The response of NK cells to contact with other cells is regulated through many different receptor-ligand interactions. However, it is not clear yet to what extent the distribution and mobility of ligands anchored into the plasma membrane of target cells influence NK cell activation and immune synapse formation during NK-target cell interactions. To study the role of the distribution and mobility of ligands on target cells, and to avoid the use of toxic inhibitors such as cyclodextrin, cholesterol-auxotroph Drosophila cell line S2 was transfected with GPI-linked CD48 (the ligand of NK cell receptor 2B4) and ULBP1 (the ligand of NK cell receptor NKG2D) and grown in serum-free conditions to eliminate cholesterol-enriched domains. 2B4- and NKG2D-dependent activation of NK cells was altered when the ligands where expressed in cholesterol-free S2 cells, suggesting a role of target cell membrane micro-domains in the response of NK cells. Experiments are underway to test how the mobility of ligands on the target cell may influence recognition by NK cells.

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Strauss, Mike, David J. Filman, David M. Belnap, Naiqian Cheng, Roane T. Noel, and James M. Hogle. "Nectin-Like Interactions between Poliovirus and Its Receptor Trigger Conformational Changes Associated with Cell Entry." Journal of Virology 89, no. 8 (January 28, 2015): 4143–57. http://dx.doi.org/10.1128/jvi.03101-14.

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ABSTRACTPoliovirus infection is initiated by attachment to a receptor on the cell surface called Pvr or CD155. At physiological temperatures, the receptor catalyzes an irreversible expansion of the virus to form an expanded form of the capsid called the 135S particle. This expansion results in the externalization of the myristoylated capsid protein VP4 and the N-terminal extension of the capsid protein VP1, both of which become inserted into the cell membrane. Structures of the expanded forms of poliovirus and of several related viruses have recently been reported. However, until now, it has been unclear how receptor binding triggers viral expansion at physiological temperature. Here, we report poliovirus in complex with an enzymatically partially deglycosylated form of the 3-domain ectodomain of Pvr at a 4-Å resolution, as determined by cryo-electron microscopy. The interaction of the receptor with the virus in this structure is reminiscent of the interactions of Pvr with its natural ligands. At a low temperature, the receptor induces very few changes in the structure of the virus, with the largest changes occurring within the footprint of the receptor, and in a loop of the internal protein VP4. Changes in the vicinity of the receptor include the displacement of a natural lipid ligand (called “pocket factor”), demonstrating that the loss of this ligand, alone, is not sufficient to induce particle expansion. Finally, analogies with naturally occurring ligand binding in the nectin family suggest which specific structural rearrangements in the virus-receptor complex could help to trigger the irreversible expansion of the capsid.IMPORTANCEThe cell-surface receptor (Pvr) catalyzes a large structural change in the virus that exposes membrane-binding protein chains. We fitted known atomic models of the virus and Pvr into three-dimensional experimental maps of the receptor-virus complex. The molecular interactions we see between poliovirus and its receptor are reminiscent of the nectin family, by involving the burying of otherwise-exposed hydrophobic groups. Importantly, poliovirus expansion is regulated by the binding of a lipid molecule within the viral capsid. We show that receptor binding either causes this molecule to be expelled or requires it, but that its loss is not sufficient to trigger irreversible expansion. Based on our model, we propose testable hypotheses to explain how the viral shell becomes destabilized, leading to RNA uncoating. These findings give us a better understanding of how poliovirus has evolved to exploit a natural process of its host to penetrate the membrane barrier.

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Wilmes, Stephan, Maximillian Hafer, Joni Vuorio, Julie A. Tucker, Hauke Winkelmann, Sara Löchte, Tess A. Stanly, et al. "Mechanism of homodimeric cytokine receptor activation and dysregulation by oncogenic mutations." Science 367, no. 6478 (February 6, 2020): 643–52. http://dx.doi.org/10.1126/science.aaw3242.

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Homodimeric class I cytokine receptors are assumed to exist as preformed dimers that are activated by ligand-induced conformational changes. We quantified the dimerization of three prototypic class I cytokine receptors in the plasma membrane of living cells by single-molecule fluorescence microscopy. Spatial and spatiotemporal correlation of individual receptor subunits showed ligand-induced dimerization and revealed that the associated Janus kinase 2 (JAK2) dimerizes through its pseudokinase domain. Oncogenic receptor and hyperactive JAK2 mutants promoted ligand-independent dimerization, highlighting the formation of receptor dimers as the switch responsible for signal activation. Atomistic modeling and molecular dynamics simulations based on a detailed energetic analysis of the interactions involved in dimerization yielded a mechanistic blueprint for homodimeric class I cytokine receptor activation and its dysregulation by individual mutations.

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Bennasroune, Amar, Maria Fickova, Anne Gardin, Sylvie Dirrig-Grosch, Dominique Aunis, Gérard Crémel, and Pierre Hubert. "Transmembrane Peptides as Inhibitors of ErbB Receptor Signaling." Molecular Biology of the Cell 15, no. 7 (July 2004): 3464–74. http://dx.doi.org/10.1091/mbc.e03-10-0753.

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Receptor tyrosine kinases have a single transmembrane (TM) segment that is usually assumed to play a passive role in ligand-induced dimerization and activation of the receptor. However, mutations within some of these receptors, and recent studies with the epidermal growth factor (EGF) and ErbB2 receptors have indicated that interactions between TM domains do contribute to stabilization of ligand-independent and/or ligand-induced receptor dimerization and activation. One consequence of the importance of these interactions is that short hydrophobic peptides corresponding to these domains should act as specific inhibitors. To test this hypothesis, we constructed expression vectors encoding short fusion peptides encompassing native or mutated TM domains of the EGF, ErbB2, and insulin receptors. In human cell lines overexpressing the wild-type EGF receptor or ErbB2, we observed that the peptides are expressed at the cell surface and that they inhibit specifically the autophosphorylation and signaling pathway of their cognate receptor. Identical results were obtained with peptides chemically synthesized. Mechanism of action involves inhibition of dimerization of the receptors as shown by the lack of effects of mutant nondimerizing sequences, completed by density centrifugation and covalent cross-linking experiments. Our findings stress the role of TM domain interactions in ErbB receptor function, and possibly for other single-spanning membrane proteins.

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Dao, Long, Qingnan Zhao, Jiemiao Hu, Xueqing Xia, Qing Yang, and Shulin Li. "A microfluidics-based method for isolation and visualization of cells based on receptor-ligand interactions." PLOS ONE 17, no. 10 (October 6, 2022): e0274601. http://dx.doi.org/10.1371/journal.pone.0274601.

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Receptor-ligand binding has been analyzed at the protein level using isothermal titration calorimetry and surface plasmon resonance and at the cellular level using interaction-associated downstream gene induction/suppression. However, no currently available technique can characterize this interaction directly through visualization. In addition, all available assays require a large pool of cells; no assay capable of analyzing receptor-ligand interactions at the single-cell level is publicly available. Here, we describe a new microfluidic chip–based technique for analyzing and visualizing these interactions at the single-cell level. First, a protein is immobilized on a glass slide and a low-flow-rate pump is used to isolate cells that express receptors that bind to the immobilized ligand. Specifically, we demonstrate the efficacy of this technique by immobilizing biotin-conjugated FGL2 on an avidin-coated slide chip and passing a mixture of GFP-labeled wild-type T cells and RFP-labeled FcγRIIB-knockout T cells through the chip. Using automated scanning and counting, we found a large number of GFP+ T cells with binding activity but significantly fewer RFP+ FcγRIIB-knockout T cells. We further isolated T cells expressing a membrane-anchored, tumor-targeted IL-12 based on the receptor’s affinity to vimentin to confirm the versatility of our technique. This protocol allows researchers to isolate receptor-expressing cells in about 4 hours for further downstream processing.

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38

Káňová, E., I. Jiménez-Munguía, Ľ. Čomor, Z. Tkáčová, I. Širochmanová, K. Bhide, and M. Bhide. "The Role of Meningococcal Porin B in Protein-Protein Interactions with Host Cells." Folia Veterinaria 62, no. 1 (March 1, 2018): 52–58. http://dx.doi.org/10.2478/fv-2018-0008.

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Abstract Neisseria meningitidis is a Gram-negative diplococcus responsible for bacterial meningitis and fatal sepsis. Ligand-receptor interactions are one of the main steps in the development of neuroinvasion. Porin B (PorB), neisserial outer membrane protein (ligand), binds to host receptors and triggers many cell signalling cascades allowing the meningococcus to damage the host cells or induce immune cells responses via the TLR2-dependent mechanisms. In this paper, we present a brief review of the structure and function of PorB.

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Sebald, Walter, Joachim Nickel, Jin-Li Zhang, and Thomas D. Mueller. "Molecular recognition in bone morphogenetic protein (BMP)/receptor interaction." Biological Chemistry 385, no. 8 (August 1, 2004): 697–710. http://dx.doi.org/10.1515/bc.2004.086.

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AbstractBone morphogenetic proteins (BMPs) and other members of the TGF-β superfamily are secreted signalling proteins determining the development, maintenance and regeneration of tissues and organs. These dimeric proteins bind, via multiple epitopes, two types of signalling receptor chains and numerous extracellular modulator proteins that stringently control their activity. Crystal structures of free ligands and of complexes with type I and type II receptor extracellular domains and with the modulator protein Noggin reveal structural epitopes that determine the affinity and specificity of the interactions. Modelling of a ternary complex BMP/(BMPR-IAEC)2/(ActR-IIEC)2suggests a mechanism of receptor activation that does not rely on direct contacts between extracellular domains of the receptors. Mutational and interaction analyses indicate that the large hydrophobic core of the interface of BMP-2 (wrist epitope) with the type I receptor does not provide a hydrophobic hot spot for binding. Instead, main chain amide and carbonyl groups that are completely buried in the contact region represent major binding determinants. The affinity between ligand and receptor chains is probably strongly increased by two-fold interactions of the dimeric ligand and receptor chains that exist as homodimers in the membrane (avidity effects). BMP muteins with disrupted epitopes for receptor chains or modulator proteins provide clues for drug design and development.

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Zhang, Luhao, Fei Wang, Qian Li, Lihua Wang, Chunhai Fan, Jiang Li, and Ying Zhu. "Classifying Cell Types with DNA-Encoded Ligand–Receptor Interactions on the Cell Membrane." Nano Letters 20, no. 5 (March 30, 2020): 3521–27. http://dx.doi.org/10.1021/acs.nanolett.0c00445.

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Periyasamy, Sankaridrug, and Pitambar Somani. "Pretreatment of human platelet membranes with trypsin abolishes GTP but not Na+ effects on α2-adrenoreceptor–agonist interactions." Canadian Journal of Physiology and Pharmacology 65, no. 5 (May 1, 1987): 778–84. http://dx.doi.org/10.1139/y87-125.

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The affinity of many types of membrane-bound receptors coupled negatively to adenylate cyclase is regulated by divalent and monovalent cations and by guanine nucleotides (GTP). We used α2-adrenoreceptors of human platelets as a model system to find out the effect of limited proteolysis with trypsin on the regulation of the α2-adrenoreceptor–agonist interactions by GTP and Na+. We found that partial proteolysis of the membranes with trypsin for 3 min at 35 °C reduced specific [3H]yohimbine binding to platelet membranes to 40–50% of control. The following characteristics of the receptors remaining after proteolysis were similar to those of untreated membranes: affinity for the agonist and antagonists, stereospecificity, and kinetic properties. Trypsin also did not modify the ability of the receptor's change from a high to low affinity state in the presence of Na+. These findings suggested that the capability of the receptors to recognize the ligand and their ability to undergo a conformational change in the presence of the agonist were retained despite a reduction in the total number of receptors by trypsin. However, the modulation of the receptor–agonist interactions by GTP or Mg2+ was lost in the trypsin-pretreated membranes, while the modulation by Na+ remained intact. It is suggested that the loss of GTP or Mg2+ effects on receptor–ligand interactions produced by trypsin may be due to trypsin-induced disruption of subunits (αi, βγ) interactions of Gi protein.

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Wei, Ying, Daniel Simon, David Waltz, and Harold Chapman. "Role of Urokinase Receptor and Caveolin in Regulation of Integrin Signaling." Thrombosis and Haemostasis 82, no. 08 (1999): 291–97. http://dx.doi.org/10.1055/s-0037-1615845.

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Introduction: Integrin-Associated ProteinsIntegrins are a group of heterodimeric adhesion receptors that mediate attachment of cells to extracellular matrices and other cells. These receptors serve as a major site of information flow from the immediate pericellular environment to the cellular interior and, in reverse, as effectors of the cellular responses to such information in biological processes as diverse as inflammation, tissue remodeling, growth, and tumorigenesis.1-3 The binding specificities of integrin receptors are determined by interactions of ligands with both the α and β chains that comprise integrins. Diversity of ligand binding is promoted by the fact that a single α chain may partner with numerous distinct β chains, and cells may express more than one and, sometimes, many α and β chains. Thus, as a family, integrins have the capacity to interact with a large set of cellular and extracellular matrix ligands. The structural features of integrin heterodimers that underlie their interactive potential and ligand specificity have been the subject of several recent reviews and will not be discussed here.4,5 A fundamental feature of integrins is that their function is determined not simply by their expression on the cell surface but by their dynamic regulation through activating events that amplify, sometimes only transiently, the adhesive capacity of integrins for their counter-ligands and the signals that follow.1-3,6,7 These dynamic aspects of integrin function are dependent on integrin interactions with its neighbors in the cell membrane and inside the cell, a structural consequence of the fact that integrins contain only short cytoplasmic tails without intrinsic signaling capacity. Three major sites of dynamic regulation of integrin function, indicated schematically in Figure 1, are as follows: ligand binding, association of cytoplasmic signaling elements with integrin cytoplasmic tails, and regulation of integrin binding and signaling by association with non-integrin “membrane adaptors.” Ligand binding by integrins is a function of their conformational state (Fig. 1a). Recent molecular modeling suggests that integrin α and β chains may exist in a relatively weak binding or “inactive state,” in which α and β chains extensively overlap and the β chain obscures binding sites on the α chain.5,8 In this model “activation” of integrins results from an altered conformation in which there is less overlap and more extensive availability of binding sites for ligand engagement. Such changes in conformational state may occur as a consequence of integrin clustering (following initially weak ligand binding).The capacity of integrins to cluster and connect with the cytoskeleton (Fig. 1b) in a manner promoting adhesion and migration is also regulated by complex interactions of kinases, phosphatases, and various structural proteins that bind to and assemble around integrin cytoplasmic tails. Signals derived from G-protein-coupled chemotactic receptors or generated by engagement of integrin extracellular ligand domains and clustering promote the activation of Src-family kinases, generation of lipid mediators, and in some cases, Ca2+ transients. These membrane proximate signals initiate and then propagate the accumulation of kinases and structural proteins surrounding a cluster of integrins. These events may also, in effect, be “insideout signals” affecting the conformational state of integrin extracellular domains1 (Fig. 1a). The assembly of cytoplasmic signaling elements on or near integrin cytoplasmic tails appears fundamental to the capacity of integrins to mediate adhesion and to impart, by signaling to the cellular interior, the nature of the immediate extracellular milieu.More recently, evidence has emerged that integrin function is also regulated by non-integrin membrane proteins that associate with integrins outside or within the plasma membrane (Fig. 1c). These include the tetraspan family of membrane proteins (CD9, CD63, CD81, CD82, and others), integrin-associated protein (CD47), and CD98.9-11 In addition, we and others have previously reported that β1 integrin function is regulated by its association with the glycosylphosphatidyl-inositol (GPI)-linked non-integrin receptor, the urokinase receptor (u-PAR, CD87), and a cholesterol-binding membrane protein called caveolin.12,13 As with ligand binding, structural features of the integrin heterodimers appear to dictate their association with these different membrane adaptors. For example, CD47 is reported to specifically interact with and promote the signaling function of αv/β3.10 The tetraspan family of proteins predominantly interacts with β1 integrins, but reportedly, only with certain α chain/β1 pairs.9 The functional effects of tetraspan proteins on integrin function is uncertain, though it is remarkable that at least two of these proteins (CD81 and 82) have been reported to inhibit tumor cell motility and act as tumor suppressors.14, 15 Caveolin and u-PAR also exhibit preferences for integrin interactions. Wary et al reported α-chain specificity for the association of β1 integrins with caveolin-1.12 The u-PAR receptor physically binds to the purified β2 integrin Mac-1 (CD11b/CD18), though no binding is observed in similar experiments with the β2 integrin CD11a/CD18 (LFA-1).16 Thus, like extracellular ligand interactions and intracellular cytoplasmic interactions, the capacity of integrins to interact with non-integrin membrane adaptors appears to be determined by amino acid sequences within the integrin heterodimers. The exact molecular basis for these interactions remains uncertain and is being actively investigated. This brief review will focus on the functional consequences of interaction of β1integrins with two of these membrane adaptors, u-PAR and caveolin.

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Hay, Debbie L., Christopher S. Walker, Joseph J. Gingell, Graham Ladds, Christopher A. Reynolds, and David R. Poyner. "Receptor activity-modifying proteins; multifunctional G protein-coupled receptor accessory proteins." Biochemical Society Transactions 44, no. 2 (April 11, 2016): 568–73. http://dx.doi.org/10.1042/bst20150237.

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Receptor activity-modifying proteins (RAMPs) are single pass membrane proteins initially identified by their ability to determine the pharmacology of the calcitonin receptor-like receptor (CLR), a family B G protein-coupled receptor (GPCR). It is now known that RAMPs can interact with a much wider range of GPCRs. This review considers recent developments on the structure of the complexes formed between the extracellular domains (ECDs) of CLR and RAMP1 or RAMP2 as these provide insights as to how the RAMPs direct ligand binding. The range of RAMP interactions is also considered; RAMPs can interact with numerous family B GPCRs as well as examples of family A and family C GPCRs. They influence receptor expression at the cell surface, trafficking, ligand binding and G protein coupling. The GPCR–RAMP interface offers opportunities for drug targeting, illustrated by examples of drugs developed for migraine.

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Frazzette, Nicholas, Anthony C. Cruz, Xufeng Wu, John A. Hammer, Jennifer Lippincott-Schwartz, Richard M. Siegel, and Prabuddha Sengupta. "Super-Resolution Imaging of Fas/CD95 Reorganization Induced by Membrane-Bound Fas Ligand Reveals Nanoscale Clustering Upstream of FADD Recruitment." Cells 11, no. 12 (June 12, 2022): 1908. http://dx.doi.org/10.3390/cells11121908.

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Signaling through the TNF-family receptor Fas/CD95 can trigger apoptosis or non-apoptotic cellular responses and is essential for protection from autoimmunity. Receptor clustering has been observed following interaction with Fas ligand (FasL), but the stoichiometry of Fas, particularly when triggered by membrane-bound FasL, the only form of FasL competent at inducing programmed cell death, is not known. Here we used super-resolution microscopy to study the behavior of single molecules of Fas/CD95 on the plasma membrane after interaction of Fas with FasL on planar lipid bilayers. We observed rapid formation of Fas protein superclusters containing more than 20 receptors after interactions with membrane-bound FasL. Fluorescence correlation imaging demonstrated recruitment of FADD dependent on an intact Fas death domain, with lipid raft association playing a secondary role. Flow-cytometric FRET analysis confirmed these results, and also showed that some Fas clustering can occur in the absence of FADD and caspase-8. Point mutations in the Fas death domain associated with autoimmune lymphoproliferative syndrome (ALPS) completely disrupted Fas reorganization and FADD recruitment, confirming structure-based predictions of the critical role that these residues play in Fas–Fas and Fas–FADD interactions. Finally, we showed that induction of apoptosis correlated with the ability to form superclusters and recruit FADD.

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Żuk, Justyna, Damian Bartuzi, Przemysław Miszta, and Agnieszka A. Kaczor. "The Role of Lipids in Allosteric Modulation of Dopamine D2 Receptor—In Silico Study." Molecules 27, no. 4 (February 16, 2022): 1335. http://dx.doi.org/10.3390/molecules27041335.

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The dopamine D2 receptor, belonging to the class A G protein-coupled receptors (GPCRs), is an important drug target for several diseases, including schizophrenia and Parkinson’s disease. The D2 receptor can be activated by the natural neurotransmitter dopamine or by synthetic ligands, which in both cases leads to the receptor coupling with a G protein. In addition to receptor modulation by orthosteric or allosteric ligands, it has been shown that lipids may affect the behaviour of membrane proteins. We constructed a model of a D2 receptor with a long intracellular loop (ICL3) coupled with Giα1 or Giα2 proteins, embedded in a complex asymmetric membrane, and simulated it in complex with positive, negative or neutral allosteric ligands. In this study, we focused on the influence of ligand binding and G protein coupling on the membrane–receptor interactions. We show that there is a noticeable interplay between the cell membrane, G proteins, D2 receptor and its modulators.

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Kilpatrick, Laura E., and Stephen J. Hill. "The use of fluorescence correlation spectroscopy to characterize the molecular mobility of fluorescently labelled G protein-coupled receptors." Biochemical Society Transactions 44, no. 2 (April 11, 2016): 624–29. http://dx.doi.org/10.1042/bst20150285.

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The membranes of living cells have been shown to be highly organized into distinct microdomains, which has spatial and temporal consequences for the interaction of membrane bound receptors and their signalling partners as complexes. Fluorescence correlation spectroscopy (FCS) is a technique with single cell sensitivity that sheds light on the molecular dynamics of fluorescently labelled receptors, ligands or signalling complexes within small plasma membrane regions of living cells. This review provides an overview of the use of FCS to probe the real time quantification of the diffusion and concentration of G protein-coupled receptors (GPCRs), primarily to gain insights into ligand–receptor interactions and the molecular composition of signalling complexes. In addition we document the use of photon counting histogram (PCH) analysis to investigate how changes in molecular brightness (ε) can be a sensitive indicator of changes in molecular mass of fluorescently labelled moieties.

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Bean, J. W., D. F. Sargent, and R. Schwyzer. "Ligand/Receptor Interactions —The Influence of the Microenvironment on Macroscopic Properties. Electrostatic Interactions with the Membrane Phase." Journal of Receptor Research 8, no. 1-4 (January 1988): 375–89. http://dx.doi.org/10.3109/10799898809048999.

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Mani, Maheswaran, Shivkumar Venkatasubrahmanyam, Yujun Yang, Mark Krampf, Jing Huang, Atul Butte, Thomas Jahn, and Kenneth I. Weinberg. "Synergy Between Kit Ligand (KL) and IL-4 In Mast Cells Is Mediated by Cross-Receptor Interactions In Lipid Rafts." Blood 116, no. 21 (November 19, 2010): 1564. http://dx.doi.org/10.1182/blood.v116.21.1564.1564.

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Abstract Abstract 1564 Studies of erythroid and thymic differentiation have shown that cross-receptor interactions between the stem cell factor receptor Kit, a receptor tyrosine kinase (RTK), and tissue-restricted Type I cytokine receptors (EpoR and IL-7R, respectively) are necessary for normal development of each lineage. To determine whether these Kit-Type I cytokine receptor interactions are a ubiquitous phenomena in Kit (+) hematopoietic cells, we studied murine bone marrow derived mast cells (BMMC), which express Kit and IL-4R, and are responsive to both cognate ligands. Both KL and IL-4 were individually mitogenic; combinations of KL and IL-4 synergistically promoted BMMC proliferation, even at suboptimal concentrations of each ligand (Synergy Index 2.7 for SCF 20ng/ml + IL-4 5ng/ml). Similar to the results seen previously with Kit and EpoR or IL-7R, activation of Kit by KL resulted in cross-receptor tyrosine phosphorylation of the IL-4Rα and γc subunits of IL-4R, even in the absence of their cognate ligand, IL-4. Each subunit of the IL-4R was independently phosphorylated by activated Kit, in the absence of Jak3. Furthermore, in the malignant mast cell line HMC-1, inhibition of oncogenic Kit by imatinib also reversed constitutive phosphorylation of IL-4R. Previous studies have shown that STAT 1α, STAT 5A, and STAT 5B, but not Stat6, are bound to and directly phosphorylated by activated Kit. In contrast, STAT6 is activated by engagement of IL-4R by its cognate ligand. Cross-receptor phosphorylation of IL-4R by activated Kit in BMMC induced STAT6 phosphorylation, with the same apparent pI as after activation of IL-4R by cognate ligand. Subcellular fractionation showed that activated Kit, γc, Jak3, and STATs were localized in lipid raft fractions upon KL stimulation. Inhibition of lipid raft formation by MβCD resulted in loss of both cross-receptor tyrosine phosphorylation of IL-4R by Kit and synergistic proliferation, but not proliferation induced by each cognate ligand. Gene expression analyses of KL stimulated BMMC from wt and IL-4Rα/γc deficient mice demonstrated that over 30% of the Kit gene signature was dependent on the presence of the IL-4R. Together, the data indicate that the synergy of KL and IL-4 in BMMC is mediated by cross-receptor interactions between Kit and IL-4R in raft membrane microdomains. The Kit-IL-4R interaction is the third cross-receptor interaction described between the Kit RTK and a tissue-restricted Type I cytokine receptor. Besides RBC, thymocytes, and mast cells, such cross-receptor interactions are likely to be a general mechanism for synergistic and tissue-specific pleiotropic signaling by Kit in hematopoiesis and possibly other cell types, e.g., various Kit (+) stem cell populations and cancers. Disclosures: No relevant conflicts of interest to declare.

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Weth, Oliver, Simone Haeberlein, Martin Haimann, Yinjie Zhang, and Christoph G. Grevelding. "Towards deorphanizing G protein-coupled receptors of Schistosoma mansoni using the MALAR yeast two-hybrid system." Parasitology 147, no. 8 (December 16, 2019): 865–72. http://dx.doi.org/10.1017/s0031182019001756.

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AbstractSchistosomiasis is an acute and chronic disease caused by parasitic worms of the genus Schistosoma. Treatment is solely dependent on praziquantel. In the face of the worldwide dimension, projects have been initiated to develop new chemotherapies. Due to their proven druggability, G protein-coupled receptors (GPCRs) are promising targets for anthelmintics. However, to identify candidate receptors, a deeper understanding of GPCR signalling in schistosome biology is essential. Comparative transcriptomics of paired and unpaired worms and their gonads revealed 59 differentially regulated GPCR-coding genes putatively involved in neuronal processes. In general, the diversity among GPCRs and their integral membrane topology make it difficult to characterize and deorphanize these receptors. To overcome existing limitations, we performed a pilot approach and utilized the innovative Membrane-Anchored Ligand And Receptor yeast two-hybrid system (MALAR-Y2H) to associate potential neuropeptide ligands with their cognate receptors. Here, we demonstrated the ability to express full-length GPCRs of Schistosoma mansoni in a heterologous yeast-based system. Additionally, we localized GPCRs and chimeras of neuropeptides fused to the WBP1 transmembrane domain of yeast to the plasma membrane of yeast cells. Reporter gene assays indicated ligand-receptor binding, which allowed us to identify certain neuropeptides as potential ligands for two GPCRs, which had been found before to be differentially expressed in schistosomes in a pairing-dependent manner. Thus, the MALAR-Y2H system appears suitable to unravel schistosome GPCR–ligand interactions. Besides its relevance for understanding schistosome biology, identifying and characterizing GPCR–ligand interaction will also contribute to applied research aspects.

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Betancourt, Miguel, Yvonne Ducolomb, Irma Jiménez, Eduardo Casas, Edmundo Bonilla, and Trish Berger. "Sperm plasma membrane receptors for the porcine oocyte plasma membrane." Zygote 6, no. 2 (May 1998): 155–58. http://dx.doi.org/10.1017/s0967199498000082.

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In vitro fertilisation (IVF) was used to assess the ability of solubilised sperm plasma membrane (PM) proteins to inhibit the interaction of intact gametes. This is a first step before evaluating the ability of individual isolated proteins to competitively inhibit sperm-oocyte interaction as part of the process of studying the molecular events of fertilisation. Porcine oocytes were aspirated from ovaries, matured for 48 h in Medium 199, and the zona pellucida (ZP) was removed by exposure to acid Tyrode's solution. ZP-free matured oocytes were exposed to 200–800 μg/ml sperm PM protein for 1 h prior to insemination and during gamete co-incubation. Twenty-four hours after insemination with 5 × 105 capacitated sperm/ml, the oocytes were fixed, stained and examined. Sperm PM protein clearly inhibited IVF in a concentration-dependent manner (r = −0.87). The inhibition index (I50%), representing the sperm PM protein concentration necessary to inhibit IVF to 50% of the control value, was 310 µg/ml. These results demonstrate that solubilised sperm PM protein inhibits the interaction of intact gametes as one might expect for receptor-ligand interactions. Furthermore, the complement of sperm PM proteins appeared maximally effective at a calculated concentration of 690 µm/ml, providing a foundation for further studies with individual proteins.

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Journal articles on the topic 'Membrane receptor-Ligand interactions'

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