scholarly journals A genetically encoded single-wavelength sensor for imaging cytosolic and cell surface ATP

2018 ◽  
Author(s):  
Mark Lobas ◽  
Jun Nagai ◽  
Mira T. Kronschläger ◽  
Philip Borden ◽  
Jonathan S. Marvin ◽  
...  
Keyword(s):  
Cell Surface ◽  
Protein Function ◽  
Fluorescent Protein ◽  
Cell Types ◽  
Specific Cell ◽  
Epsilon Subunit ◽  
Extracellular Signal ◽  
Intracellular Energy ◽  
Cellular Compartments

Adenosine 5’ triphosphate (ATP) is a universal intracellular energy source1 and an evolutionarily ancient2 extracellular signal3–5. Here, we report the generation and characterization of single-wavelength genetically encoded fluorescent sensors (iATPSnFRs) for imaging extracellular and cytosolic ATP from insertion of circularly permuted superfolder GFP into the epsilon subunit of F0F1-ATPase from Bacillus PS3. On the cell surface and within the cytosol, iATPSnFR1.0 responded to relevant ATP concentrations (30 μM to 3 mM) with fast increases in fluorescence. iATPSnFRs can be genetically targeted to specific cell types and sub-cellular compartments, imaged with standard light microscopes, do not respond to other nucleotides and nucleosides, and when fused with a red fluorescent protein function as ratiometric indicators. iATPSnFRs represent promising new reagents for imaging ATP dynamics.

scholarly journals Cellular Receptors Involved in KSHV Infection

Viruses ◽  
2021 ◽  
Vol 13 (1) ◽  
pp. 118
Author(s):  
Emma van der Meulen ◽  
Meg Anderton ◽  
Melissa J. Blumenthal ◽  
Georgia Schäfer
Keyword(s):  
Virus Infection ◽  
Cell Surface ◽  
Kaposi's Sarcoma ◽  
Kaposi’S Sarcoma ◽  
Cell Types ◽  
Target Cells ◽  
Specific Cell ◽  
Cellular Receptors ◽  
Diverse Range ◽  
Kshv Infection

The process of Kaposi’s Sarcoma Herpes Virus’ (KSHV) entry into target cells is complex and engages several viral glycoproteins which bind to a large range of host cell surface molecules. Receptors for KSHV include heparan sulphate proteoglycans (HSPGs), several integrins and Eph receptors, cystine/glutamate antiporter (xCT) and Dendritic Cell-Specific Intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN). This diverse range of potential binding and entry sites allows KSHV to have a broad cell tropism, and entry into specific cells is dependent on the available receptor repertoire. Several molecules involved in KSHV entry have been well characterized, particularly those postulated to be associated with KSHV-associated pathologies such as Kaposi’s Sarcoma (KS). In this review, KSHV infection of specific cell types pertinent to its pathogenesis will be comprehensively summarized with a focus on the specific cell surface binding and entry receptors KSHV exploits to gain access to a variety of cell types. Gaps in the current literature regarding understanding interactions between KSHV glycoproteins and cellular receptors in virus infection are identified which will lead to the development of virus infection intervention strategies.

scholarly journals Exo-enzymatic addition of diazirine-modified sialic acid to cell surfaces enables photocrosslinking of glycoproteins

2021 ◽  
Author(s):  
Nageswari Yarravarapu ◽  
Rohit Sai Reddy Konada ◽  
Narek Darabedian ◽  
Nichole J. Pedowtiz ◽  
Soumya N. Krishnamurthy ◽  
...  
Keyword(s):  
Sialic Acid ◽  
Cell Surface ◽  
Cell Types ◽  
Cell Surfaces ◽  
Binding Interactions ◽  
Multiple Cell ◽  
Labeling Method ◽  
Cell Surface Glycoconjugates

Glycan binding often mediates extracellular macromolecular recognition events. Accurate characterization of these binding interactions can be difficult because of dissociation and scrambling that occur during purification and analysis steps. Use of photocrosslinking methods has been pursued to covalently capture glycan-dependent interactions in situ however use of metabolic glycan engineering methods to incorporate photocrosslinking sugar analogs is limited to certain cell types. Here we report an exo-enzymatic labeling method to add a diazirine-modified sialic acid (SiaDAz) to cell surface glycoconjugates. The method involves chemoenzymatic synthesis of diazirine-modified CMP-sialic acid (CMP-SiaDAz), followed by sialyltransferase-catalyzed addition of SiaDAz to desialylated cell surfaces. Cell surface SiaDAz-ylation is compatible with multiple cell types and is facilitated by endogenous extracellular sialyltransferase activity present in Daudi B cells. This method for extracellular addition of α2-6-linked SiaDAz enables UV-induced crosslinking of CD22, demonstrating the utility for covalent capture of glycan-mediated binding interactions.

scholarly journals Pituitary Adenylate Cyclase-Activating Polypeptide Protects Glomerular Podocytes from Inflammatory Injuries

2015 ◽  
Vol 2015 ◽  
pp. 1-10 ◽  
Author(s):  
Kenichi Sakamoto ◽  
Kyoko Kuno ◽  
Minoru Takemoto ◽  
Peng He ◽  
Takahiro Ishikawa ◽  
...  
Keyword(s):  
Adenylate Cyclase ◽  
Treatment Options ◽  
Cell Types ◽  
Specific Cell ◽  
Monocyte Chemoattractant Protein 1 ◽  
Extracellular Signal Regulated Kinase ◽  
Extracellular Signal ◽  
Pituitary Adenylate Cyclase ◽  
End Stage ◽  
Anti Inflammation

Diabetic nephropathy (DN) is a leading cause of end-stage kidney disease; however, there are few treatment options. Inflammation plays a crucial role in the initiation and/or progression of DN. Pituitary adenylate cyclase-activating polypeptide (PACAP) is a neuropeptide, which was originally isolated from the ovine hypothalamus and reportedly has diverse biological functions. It has been reported that PACAP has renoprotective effects in different models of kidney pathology. However, the specific cell types within the kidney that are protected by PACAP have not yet been reported. In this study, we localized VPAC1, one of the PACAP receptors, to glomerular podocytes, which also reportedly has crucial roles not only in glomerular physiology but also in pathology. PACAP was effective in the downregulation of proinflammatory cytokines, such as monocyte chemoattractant protein-1 (MCP-1) and interleukin-6, which had been induced by the activation of toll-like receptor (TLR) with lipopolysaccharide. PACAP also had downregulated the expression of MCP-1 through the protein kinase A signaling pathway; this led to the attenuation of the activation of extracellular signal-regulated kinase and nuclear factor-kappa B signaling. Our results suggested that PACAP could be a possible treatment option for DN through the use of anti-inflammation effects on glomerular podocytes.

scholarly journals Isolation of an adhesion-mediating protein from chick neural retina adherons.

1985 ◽  
Vol 101 (3) ◽  
pp. 1071-1077 ◽  
Author(s):  
D Schubert ◽  
M LaCorbiere
Keyword(s):  
Cell Surface ◽  
Heparan Sulfate ◽  
Cultured Cells ◽  
Cell Types ◽  
Neural Retina ◽  
Cell Surface Receptor ◽  
Isolation And Characterization ◽  
Surface Receptor ◽  
Adhesive Interactions

Adherons are high molecular weight glycoprotein complexes which are released into the growth medium of cultured cells. They mediate the adhesive interactions of many cell types, including those of embryonic chick neural retina. The cell surface receptor for chick neural retina adherons has been purified, and shown to be a heparan sulfate proteoglycan (Schubert, D., and M. LaCorbiere, 1985, J. Cell Biol., 100:56-63). This paper describes the isolation and characterization of a protein in neural retina adherons which interacts specifically with the cell surface receptor. The 20,000-mol-wt protein, called retinal purpurin (RP), stimulates neural retina cell-substratum adhesion and prolongs the survival of neural retina cells in culture. The RP protein interacts with heparin and heparan sulfate, but not with other glycosaminoglycans. Monovalent antibodies against RP inhibit RP-cell adhesion as well as adheron-cell interactions. The RP protein is found in neural retina, but not in other tissues such as brain and muscle. These data suggest that RP plays a role in both the survival and adhesive interactions of neural retina cells.

Binding of peanut lectin to specific epithelial cell types in kidney

1982 ◽  
Vol 242 (1) ◽  
pp. C117-C120 ◽  
Author(s):  
M. LeHir ◽  
B. Kaissling ◽  
B. M. Koeppen ◽  
J. B. Wade
Keyword(s):  
Cell Types ◽  
Rabbit Kidney ◽  
Specific Cell ◽  
Luminal Membrane ◽  
Isolation And Characterization ◽  
Collecting Ducts ◽  
Epithelial Membranes ◽  
Outer Stripe ◽  
Specific Affinity

The binding of peanut agglutinin (PNA) to epithelial membranes of the rabbit kidney was evaluated at the light- and electron-microscope level using PNA conjugated to horseradish peroxidase. In the renal cortex and outer stripe of the medulla PNA appears to bind exclusively to the luminal membrane of intercalated cells in connecting tubules and collecting ducts. PNA also binds to the thin descending limb of the loop of Henle in the inner stripe and inner zone of the medulla. This very specific affinity of PNA should be useful in the isolation and characterization of specific cell types in cytologically heterogeneous epithelia.

scholarly journals Genetically modified adenoviral vector with the protein transduction domain of Tat improves gene transfer to CAR-deficient cells

2009 ◽  
Vol 29 (2) ◽  
pp. 103-109 ◽  
Author(s):  
Shihai Liu ◽  
Qinwen Mao ◽  
Weifeng Zhang ◽  
Xiaojing Zheng ◽  
Ye Bian ◽  
...  
Keyword(s):  
Gene Transfer ◽  
Cell Surface ◽  
Fluorescent Protein ◽  
Function Analysis ◽  
Cell Types ◽  
Transduction Efficiency ◽  
Target Cells ◽  
Protein Transduction ◽  
Protein Transduction Domain ◽  
Green Fluorescent

The transduction efficiency of Ad (adenovirus) depends, to some extent, on the expression level of CAR (coxsackievirus and Ad receptor) of a target cell. The low level of CAR on the cell surface is a potential barrier to efficient gene transfer. To overcome this problem, PTD.AdeGFP (where eGFP is enhanced green fluorescent protein) was constructed by modifying the HI loop of Ad5 (Ad type 5) fibre with the Tat (trans-activating) PTD (protein transduction domain) derived from HIV. The present study showed that PTD.AdeGFP significantly improved gene transfer to multiple cell types deficient in expression of CAR. The improvement in gene transfer was not the result of charge-directed binding between the virus and the cell surface. Although PTD.AdeGFP formed aggregates, it infected target cells in a manner different from AdeGFP aggregates precipitated by calcium phosphate. In addition, PTD.AdeGFP was able to transduce target cells in a dynamin-independent pathway. The results provide some new clues as to how PTD.AdeGFP infects target cells. This new vector would be valuable in gene-function analysis and for gene therapy in cancer.

scholarly journals Adenovirus Vectors Target Several Cell Subtypes of Mammalian Inner Ear In Vivo

2016 ◽  
Vol 2016 ◽  
pp. 1-8 ◽  
Author(s):  
Yilai Shu ◽  
Yong Tao ◽  
Wenyan Li ◽  
Jun Shen ◽  
Zhengmin Wang ◽  
...  
Keyword(s):  
Inner Ear ◽  
Fluorescent Protein ◽  
Cell Types ◽  
Sensory Epithelium ◽  
Specific Cell ◽  
Supporting Cells ◽  
Auditory Hair Cells ◽  
A Genome ◽  
Adenovirus Vectors

Mammalian inner ear harbors diverse cell types that are essential for hearing and balance. Adenovirus is one of the major vectors to deliver genes into the inner ear for functional studies and hair cell regeneration. To identify adenovirus vectors that target specific cell subtypes in the inner ear, we studied three adenovirus vectors, carrying a reporter gene encoding green fluorescent protein (GFP) from two vendors or with a genome editing gene Cre recombinase (Cre), by injection into postnatal days 0 (P0) and 4 (P4) mouse cochlea through scala media by cochleostomy in vivo. We found three adenovirus vectors transduced mouse inner ear cells with different specificities and expression levels, depending on the type of adenoviral vectors and the age of mice. The most frequently targeted region was the cochlear sensory epithelium, including auditory hair cells and supporting cells. Adenovirus with GFP transduced utricular supporting cells as well. This study shows that adenovirus vectors are capable of efficiently and specifically transducing different cell types in the mammalian inner ear and provides useful tools to study inner ear gene function and to evaluate gene therapy to treat hearing loss and vestibular dysfunction.

scholarly journals Isolation and immunologic characterization of a human. B-lymphocyte-specific, cell surface antigen.

1976 ◽  
Vol 144 (1) ◽  
pp. 98-112 ◽  
Author(s):  
R E Humphreys ◽  
J M McCune ◽  
L Chess ◽  
H C Herrman ◽  
D J Malenka ◽  
...  
Keyword(s):  
Cell Surface ◽  
Cell Lines ◽  
B Lymphocyte ◽  
Surface Protein ◽  
Specific Cell ◽  
Cell Surface Protein ◽  
Lymphoblast Cell Line ◽  
Lymphocyte Antigens ◽  
Specific Cell Surface

In addition to HL-A antigens, another cell surface protein complex has been obtained from membranes of the human B-lymphoblast cell line IM-1. This complex which was solubilized with papain, consisted of polypeptides of 23,000 and 30,000 daltons (p23, 30). Rabbit antisera to this material precipitated from [35S]methionine-labeled detergent-solubilized cells, three proteins of 39,000, 34,000, and 29,000 daltons. These antisera were specifically cytotoxic for B lymphocytes of peripheral blood, for B-lymphoblast cell lines, and for EAC rosette receptor-positive surface Ig-negative (Null) lymphocytes. The p23,30 complex was not present on T lymphocytes, EAC rosette receptor-negative Null lymphocytes, or platelets. In addition, the p23,30 complex from several cell lines inhibited alloantisera from multiparous Amish women which had been shown to recognize non-HL-A, B-lymphocyte antigens. Some other properties of the anti-p23,30 sera antisera were described.

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