Inhibition of rat ventricular automaticity by adenosine

This study was designed to characterize adenosine's negative chronotropic effect on ventricular pacemakers. The spontaneous beating rate of isolated, isovolumic rat ventricular preparations perfused with Krebs-Henseleit solution decreased as the adenosine concentration was increased [log M effective concentration 50% (EC50) = -5.22 +/- 0.17]. The lack of effect of propranolol or atropine on this adenosine response eliminates the involvement of endogenous neurotransmitters. Support for the involvement of an external cell surface receptor was provided by findings that theophylline and 8-(4-sulfophenyl)theophylline, an analogue thought to act solely at the cell surface, significantly increased the adenosine log M EC50 to -3.94 +/- 0.22 and -3.61 +/- 0.22, respectively. An increase in spontaneous beating rate induced by theophylline, but not by its analogue, was blocked by the addition of propranolol. The relative chronotropic potency of the adenosine analogues R-PIA, S-PIA, and NECA suggests that the cell surface receptors may be of the Ri type. The negative chronotropic effects of adenosine and its analogues occurred at concentrations that had no effect on the developed pressure of the paced preparation. Electrocardiographic evaluations indicate that at high agonist concentrations, there was an abrupt alteration in electrical properties of the preparation, which could be blocked by theophylline and its analogue.

Download Full-text

Topological and Structural Plasticity of the Single Ig Fold and the Double Ig Fold Present in CD19

Biomolecules ◽

10.3390/biom11091290 ◽

2021 ◽

Vol 11 (9) ◽

pp. 1290 ◽

Cited By ~ 1

Author(s):

Philippe Youkharibache

Keyword(s):

Cell Surface ◽

3D Structure ◽

Cell Surface Receptor ◽

Cell Surface Receptors ◽

Structural Domain ◽

Surface Receptors ◽

Surface Receptor ◽

In Trans ◽

In Cis ◽

Ig Domain

The Ig fold has had a remarkable success in vertebrate evolution, with a presence in over 2% of human genes. The Ig fold is not just the elementary structural domain of antibodies and TCRs, it is also at the heart of a staggering 30% of immunologic cell surface receptors, making it a major orchestrator of cell–cell interactions. While BCRs, TCRs, and numerous Ig-based cell surface receptors form homo- or heterodimers on the same cell surface (in cis), many of them interface as ligand-receptors (checkpoints) on interacting cells (in trans) through their Ig domains. New Ig-Ig interfaces are still being discovered between Ig-based cell surface receptors, even in well-known families such as B7. What is largely ignored, however, is that the Ig fold itself is pseudosymmetric, a property that makes the Ig domain a versatile self-associative 3D structure and may, in part, explain its success in evolution, especially through its ability to bind in cis or in trans in the context of cell surface receptor–ligand interactions. In this paper, we review the Ig domains’ tertiary and quaternary pseudosymmetries, with particular attention to the newly identified double Ig fold in the solved CD19 molecular structure to highlight the underlying fundamental folding elements of Ig domains, i.e., Ig protodomains. This pseudosymmetric property of Ig domains gives us a decoding frame of reference to understand the fold, relate all Ig domain forms, single or double, and suggest new protein engineering avenues.

Download Full-text

Identifying plant cell-surface receptors: combining ‘classical’ techniques with novel methods

Biochemical Society Transactions ◽

10.1042/bst20130251 ◽

2014 ◽

Vol 42 (2) ◽

pp. 395-400 ◽

Cited By ~ 4

Author(s):

Susanne Uebler ◽

Thomas Dresselhaus

Keyword(s):

Cell Surface ◽

Affinity Purification ◽

Cell Communication ◽

Membrane Receptors ◽

Cell Surface Receptor ◽

Target Cells ◽

Surface Receptors ◽

Success Stories ◽

Surface Receptor ◽

Secreted Peptides

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.

Download Full-text

Intracellular movement of cell surface receptors after endocytosis: resialylation of asialo-transferrin receptor in human erythroleukemia cells.

The Journal of Cell Biology ◽

10.1083/jcb.100.3.826 ◽

1985 ◽

Vol 100 (3) ◽

pp. 826-834 ◽

Cited By ~ 142

Author(s):

M D Snider ◽

O C Rogers

Keyword(s):

Sialic Acid ◽

Cell Surface ◽

Golgi Complex ◽

Transferrin Receptor ◽

Cell Surface Receptor ◽

Surface Receptors ◽

Erythroleukemia Cells ◽

Intracellular Movement ◽

Surface Receptor ◽

A Cell

The intracellular movement of cell surface transferrin receptor (TfR) after internalization was studied in K562 cultured human erythroleukemia cells. The sialic acid residues of the TfR glycoprotein were used to monitor transport to the Golgi complex, the site of sialyltransferases. Surface-labeled cells were treated with neuraminidase, and readdition of sialic acid residues, monitored by isoelectric focusing of immunoprecipitated TfR, was used to assess the movement of receptor to sialyltransferase-containing compartments. Asialo-TfR was resialylated by the cells with a half-time of 2-3 h. Resialylation occurred in an intracellular organelle, since it was inhibited by treatments that allow internalization of surface components but block transfer out of the endosomal compartment. Moreover, roughly half of the resialylated molecules were cleaved when cells were retreated with neuraminidase after culturing, indicating that this fraction of the molecules had returned to the cell surface. These results suggest that TfR is transported from the cell surface to the Golgi complex, the intracellular site of sialyltransferases, and then returns to the cell surface. This pathway, which has not been previously described for a cell surface receptor, may be different from the route followed by TfR in iron uptake, since reported rates of transferrin uptake and release are significantly more rapid than the resialylation of asialo-TfR.

Download Full-text

SUN-LB130 Identification of IGSF1 Ligands

Journal of the Endocrine Society ◽

10.1210/jendso/bvaa046.2113 ◽

2020 ◽

Vol 4 (Supplement_1) ◽

Author(s):

Courtney L Smith ◽

Andrew N Bayne ◽

Jean-François Trempe ◽

Daniel J Bernard

Keyword(s):

Cell Surface ◽

Transmembrane Domain ◽

Protein Complexes ◽

Protein A ◽

Negative Control ◽

Cell Surface Receptor ◽

Immunoglobulin Superfamily ◽

Central Hypothyroidism ◽

Surface Receptors ◽

Surface Receptor

Abstract Immunoglobulin superfamily, member 1 (IGSF1), is an X-linked, type 1 transmembrane glycoprotein that is highly expressed in the anterior pituitary gland and testes. Mutations in the IGSF1 gene cause congenital central hypothyroidism, variable hypoprolactinemia, growth hormone dysregulation, and macroorchidism. Igsf1 knockout mice exhibit reduced pituitary TRH receptor (Trhr1) expression with an associated impairment in TRH-stimulated TSH secretion. The mechanism through which IGSF1 loss leads to reductions in Trhr1 levels is unresolved, at least in part because IGSF1’s cellular functions are unknown. The mature form of the IGSF1 protein consists of seven extracellular Ig loops, a single transmembrane domain containing a positively charged arginine, and a short intracellular carboxy-tail devoid of known functional motifs. Recently, IGSF1 was argued to be a member of the leukocyte receptor cluster (LRC) family. LRC proteins act as cell surface receptors for soluble or membrane-bound proteins. We therefore hypothesized that IGSF1 is a cell surface receptor for a presently undescribed ligand that regulates Trhr1 expression in pituitary thyrotrope cells. To identify candidate IGSF1 ligands, we implemented a new ligand trapping method, Ecto-Fc MS. We fused the extracellular (Ecto) domain of IGSF1 to the fragment crystallizable (Fc) region of human IgG, creating an Ecto-Fc fusion protein. Secreted IGSF1-Fc was purified and used as a ligand trap for bait proteins extracted from rat testes. The protein complexes were affinity purified with protein A beads, trypsin digested into peptides, subjected to orthogonal high-pH fractionation, and identified by tandem LC-MS/MS. More than 700 proteins were enriched in IGSF1-Fc preparations compared to an Fc-only negative control. Several secreted ligands and plasma-membrane proteins were identified, many of which are also expressed in pituitary thyrotrope cells. Identifying the ligand or ligands will enable us to determine IGSF1 function, and may lead to the discovery of novel causes of central hypothyroidism and macroorchidism.

Download Full-text

Identification of Amino Acid Residues of Anthrax Protective Antigen Involved in Binding with Lethal Factor

Infection and Immunity ◽

10.1128/iai.70.8.4477-4484.2002 ◽

2002 ◽

Vol 70 (8) ◽

pp. 4477-4484 ◽

Cited By ~ 18

Author(s):

Vibha Chauhan ◽

Rakesh Bhatnagar

Keyword(s):

Cell Surface ◽

Protective Antigen ◽

Lethal Factor ◽

Lethal Toxin ◽

Cell Surface Receptor ◽

Amino Acid Residues ◽

Anthrax Lethal Toxin ◽

Mutant Proteins ◽

Surface Receptors ◽

Surface Receptor

ABSTRACT Protective antigen (PA) and lethal factor (LF) are the two components of anthrax lethal toxin. PA is responsible for the translocation of LF to the cytosol. The binding of LF to cell surface receptor-bound PA is a prerequisite for the formation of lethal toxin. It has been hypothesized that hydrophobic residues P184, L187, F202, L203, P205, I207, I210, W226, and F236 of domain 1b of PA play an important role in the binding of PA to LF. These residues are normally buried in the 83-kDA version of PA, PA83, as determined by the crystal structure of PA. However, they become exposed due to the conformational change brought about by the cleavage of PA83 to PA63 by a cell surface protease. Mutation of the above-mentioned residues to alanine resulted in mutant proteins that were able to bind to the cell surface receptors and also to be specifically cleaved by the cellular proteases. All the mutant proteins except the F202A, L203A, P205A, and I207A mutants were able to bind to LF and were also toxic to macrophage cells in combination with LF. It was concluded that residues 202, 203, 205, and 207 of PA are essential for the binding of LF to PA.

Download Full-text

Heterogeneity in VEGF Receptor-2 mobility and organization on the endothelial cell surface leads to diverse activation models by VEGF

10.1101/800946 ◽

2019 ◽

Author(s):

Bruno da Rocha-Azevedo ◽

Sungsoo Lee ◽

Aparajita Dasgupta ◽

Anthony R. Vega ◽

Luciana R. de Oliveira ◽

...

Keyword(s):

Cell Surface ◽

Single Molecule ◽

Receptor Activation ◽

Cell Surface Receptor ◽

Vegf Receptor ◽

Surface Receptors ◽

Endothelial Cell Surface ◽

Heterogeneous Nature ◽

Surface Receptor ◽

Complex Relationships

SummaryThe nanoscale organization of cell surface receptors plays an important role in signaling. We determined this organization and its relation to receptor activation for VEGF Receptor-2 (VEGFR-2), a critical receptor tyrosine kinase in endothelial cells (ECs), by combining live-cell single-molecule imaging of endogenous VEGFR-2 with rigorous computational analysis. We found that surface VEGFR-2 can be mobile or immobile/confined, and monomeric or non-monomeric, with a complex interplay between the two. The mobility and interaction heterogeneity of VEGFR-2 in the basal state led to heterogeneity in the sequence of steps leading to VEGFR-2 activation by VEGF. Specifically, we found that VEGF can bind to both monomeric and non-monomeric VEGFR-2, and, when binding to monomeric VEGFR-2, promotes dimer formation but only for immobile/confined receptors. Overall, our study highlights the dynamic and heterogeneous nature of cell surface receptor organization and its complex relationships with receptor activation and signaling.

Download Full-text

Mutual Potentiation of Plant Immunity by Cell-surface and Intracellular Receptors

10.1101/2020.04.10.034173 ◽

2020 ◽

Cited By ~ 9

Author(s):

Bruno Pok Man Ngou ◽

Hee-Kyung Ahn ◽

Pingtao Ding ◽

Jonathan DG Jones

Keyword(s):

Immune System ◽

Cell Surface ◽

Pseudomonas Syringae ◽

Plant Immunity ◽

Cell Surface Receptor ◽

Hypersensitive Cell Death ◽

Surface Receptors ◽

Intracellular Receptor ◽

Surface Receptor ◽

Immune Pathways

The plant immune system involves cell-surface receptors that detect intercellular pathogen-derived molecules, and intracellular receptors that activate immunity upon detection of pathogen-secreted effectors that act inside the plant cell. Surface receptor-mediated immunity has been extensively studied but in authentic interactions between plants and microbial pathogens, its presence impedes study of intracellular receptor-mediated immunity alone. How these two immune pathways interact is poorly understood. Here, we reveal mutual potentiation between these two recognition-dependent defense pathways. Recognition by surface receptors activates multiple protein kinases and NADPH oxidases, whereas intracellular receptors primarily elevate abundance of these proteins. Reciprocally, the intracellular receptor-dependent hypersensitive cell death response is strongly enhanced by activation of surface receptors. Activation of either immune system alone is insufficient to provide effective resistance against Pseudomonas syringae. Thus, immune pathways activated by cell-surface and intracellular receptors mutually potentiate to activate strong defense that thwarts pathogens. By studying the activation of intracellular receptors in the absence of surface receptor-mediated immunity, we have dissected the relationship between the two distinct immune systems. These findings reshape our understanding of plant immunity and have broad implications for crop improvement.

Download Full-text

The Arabidopsis receptor kinase STRUBBELIG undergoes clathrin-dependent endocytosis

Journal of Experimental Botany ◽

10.1093/jxb/erz190 ◽

2019 ◽

Vol 70 (15) ◽

pp. 3881-3894 ◽

Cited By ~ 8

Author(s):

Jin Gao ◽

Ajeet Chaudhary ◽

Prasad Vaddepalli ◽

Marie-Kristin Nagel ◽

Erika Isono ◽

...

Keyword(s):

Cell Surface ◽

Fluorescent Protein ◽

Growth Patterns ◽

Cell Surface Receptor ◽

Receptor Kinase ◽

Surface Receptors ◽

Surface Receptor ◽

Green Fluorescent ◽

Golgi Network

AbstractSignaling mediated by cell surface receptor kinases is central to the coordination of growth patterns during organogenesis. Receptor kinase signaling is in part controlled through endocytosis and subcellular distribution of the respective receptor kinase. For the majority of plant cell surface receptors, the underlying trafficking mechanisms are not characterized. In Arabidopsis, tissue morphogenesis requires the atypical receptor kinase STRUBBELIG (SUB). Here, we studied the endocytic mechanism of SUB. Our data revealed that a functional SUB–enhanced green fluorescent protein (EGFP) fusion is ubiquitinated in vivo. We further showed that plasma membrane-bound SUB:EGFP becomes internalized in a clathrin-dependent fashion. We also found that SUB:EGFP associates with the trans-Golgi network and accumulates in multivesicular bodies and the vacuole. Co-immunoprecipitation experiments revealed that SUB:EGFP and clathrin are present within the same protein complex. Our genetic analysis showed that SUB and CLATHRIN HEAVY CHAIN (CHC) 2 regulate root hair patterning. By contrast, genetic reduction of CHC activity ameliorates the floral defects of sub mutants. Taken together, the data indicate that SUB undergoes clathrin-mediated endocytosis, that this process does not rely on stimulation of SUB signaling by an exogenous agent, and that SUB genetically interacts with clathrin-dependent pathways in a tissue-specific manner.

Download Full-text

Centrosomal P4.1-associated protein (CPAP) positively regulates endocytic vesicular transport and lysosome targeting of EGFR

Scientific Reports ◽

10.1038/s41598-021-91818-8 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Radhika Gudi ◽

Viswanathan Palanisamy ◽

Chenthamarakshan Vasu

Keyword(s):

Cell Surface ◽

Vesicular Transport ◽

Critical Role ◽

Cell Surface Receptor ◽

Regulatory Effect ◽

Cellular Functions ◽

Surface Receptors ◽

Egfr Activation ◽

Early Endosomes ◽

Surface Receptor

AbstractCentrosomal P4.1-associated protein (CPAP) plays a critical role in restricting the centriole length in human cells. Here, we report a novel, positive regulatory influence for CPAP on endocytic vesicular transport (EVT) and lysosome targeting of internalized-cell surface receptor EGFR. We observed that higher CPAP levels cause an increase in the abundance of multi-vesicular body (MVB) and EGFR is detectable in CPAP-overexpression induced puncta. The surface and cellular levels of EGFR are higher under CPAP deficiency and lower under CPAP overexpression. While ligand-engagement induced internalization or routing of EGFR into early endosomes is not influenced by cellular levels of CPAP, we found that targeting of ligand-activated, internalized EGFR to lysosome is impacted by CPAP levels. Transport of ligand-bound EGFR from early endosome to late endosome/MVB and lysosome is diminished in CPAP-depleted cells. Moreover, CPAP depleted cells appear to show a diminished ability to form MVB structures upon EGFR activation. These observations suggest a positive regulatory effect of CPAP on EVT of ligand-bound EGFR-like cell surface receptors to MVB and lysosome. Overall, identification of a non-centriolar function of CPAP in endocytic trafficking provides new insights in understanding the non-canonical cellular functions of CPAP.

Download Full-text

SPaRTAN, a computational framework for linking cell-surface receptors to transcriptional regulators

Nucleic Acids Research ◽

10.1093/nar/gkab745 ◽

2021 ◽

Vol 49 (17) ◽

pp. 9633-9647

Author(s):

Xiaojun Ma ◽

Ashwin Somasundaram ◽

Zengbiao Qi ◽

Douglas J Hartman ◽

Harinder Singh ◽

...

Keyword(s):

Cell Surface ◽

Single Cell ◽

Transcriptional Regulators ◽

Cell Types ◽

Receptor Expression ◽

Cell Surface Receptor ◽

Cell Surface Receptors ◽

Surface Receptors ◽

Surface Receptor ◽

Context Specific

Abstract The identity and functions of specialized cell types are dependent on the complex interplay between signaling and transcriptional networks. Recently single-cell technologies have been developed that enable simultaneous quantitative analysis of cell-surface receptor expression with transcriptional states. To date, these datasets have not been used to systematically develop cell-context-specific maps of the interface between signaling and transcriptional regulators orchestrating cellular identity and function. We present SPaRTAN (Single-cell Proteomic and RNA based Transcription factor Activity Network), a computational method to link cell-surface receptors to transcription factors (TFs) by exploiting cellular indexing of transcriptomes and epitopes by sequencing (CITE-seq) datasets with cis-regulatory information. SPaRTAN is applied to immune cell types in the blood to predict the coupling of signaling receptors with cell context-specific TFs. Selected predictions are validated by prior knowledge and flow cytometry analyses. SPaRTAN is then used to predict the signaling coupled TF states of tumor infiltrating CD8+ T cells in malignant peritoneal and pleural mesotheliomas. SPaRTAN enhances the utility of CITE-seq datasets to uncover TF and cell-surface receptor relationships in diverse cellular states.

Download Full-text