Regulation of cell-surface receptors for human interferon-γ on the human histiocytic lymphoma cell line U937

Interferon-gamma (IFN gamma) binds to high-affinity receptors on monocytes and is rapidly internalized. This study investigates the ability of the human monocyte-like cell line, U937, to regulate the cell-surface expression of the IFN gamma receptor (IFN gamma R) during endocytosis of ligand. Recombinant IFN gamma was radiolabelled to high specific radioactivity with Bolton-Hunter reagent and used to enumerate IFN gamma R on treated U937 cells. Cells which had internalized IFN gamma for up to 3 h displayed maximal levels of IFN gamma R at all time points tested after all unlabelled IFN gamma had been acid-stripped from the cell at pH 2.78. Therefore there was no evidence of down-modulation of the receptor. After trypsin treatment of the IFN gamma R, the cells were able to synthesize and insert into the cell membrane up to 1000 IFN gamma R molecules/h after a 60 min lag. Since biosynthesis played a minor role during the first 30 min of endocytosis, I examined other possibilities to explain the lack of down-modulation of the receptor. A solubilized-receptor assay revealed the presence of an intracellular pool of receptors equal to about 25% of the number of cell surface receptors. Using trypsin to differentiate between intracellular and surface receptors, I observed that 43% of those receptors that were internalized after a 30 min exposure to IFN gamma (580 molecules) could be recycled back to the plasma membrane. In addition, equal rates of receptor decay (t1/2 = 5 h) were observed in the presence of cycloheximide with or without IFN gamma. All the data taken together suggest that during the first 30 min of endocytosis both the expression of an intracellular source of receptor and recycling of internalized receptors contribute to maintain optimal receptor expression.

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Prolonged defects of interleukin-2 production, responsiveness, and receptor expression in patients with acute lymphoblastic leukemia

Blood ◽

10.1182/blood.v73.6.1608.bloodjournal7361608 ◽

1989 ◽

Vol 73 (6) ◽

pp. 1608-1614

Author(s):

MS Borzy ◽

D Ridgway

Keyword(s):

Acute Lymphoblastic Leukemia ◽

Cell Surface ◽

Lymphoblastic Leukemia ◽

Interleukin 2 ◽

Receptor Expression ◽

Cell Surface Receptors ◽

Con A ◽

Surface Receptors ◽

Control Subjects ◽

Patient Groups

The proliferative responsiveness to, production of, and the expression of cell-surface receptors for interleukin-2 (IL-2) were examined in 14 children with acute lymphoblastic leukemia (ALL) in remission and receiving maintenance chemotherapy for 6 to 35 months; in 19 children with ALL in remission and off all therapy for 2 to 138 months; and 15 control subjects. Short-term concanavalin A (Con A)-activated, purified T lymphocytes from patients on, as well as patients off, therapy had a significantly decreased proliferative responsiveness to a saturating amount of exogenous, recombinant IL-2 as compared to control subjects (P less than 0.005 and less than 0.05, respectively). Phytohemagglutinin (PHA)-stimulated IL-2 production by peripheral blood mononuclear cells (PBMC) was also substantially decreased in both patient groups with the median values of IL-2 produced being 2.2, 2.1, and 8.1 U/mL in the on therapy, off therapy, and control groups, respectively. In addition, PHA-induced expression of cell-surface receptors for IL-2 on PBMC was significantly decreased in both patient groups as compared to control subjects (P less than 0.01). Lymphocyte proliferation to mitogens (PHA, Con A, and pokeweed mitogen) was similar in all three groups studied. These results demonstrate that substantial quantitative and qualitative abnormalities of the IL-2-T lymphocyte system are present in the majority of treated patients with ALL, not only during maintenance therapy, but also for a prolonged period after the cessation of all chemotherapy. These long-lasting defects of the IL-2 system are most likely a late effect of chemotherapy and may result in increased complications in some long- term survivors of ALL.

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Recognition of human urine α-N-acetylglucosaminidase by rat hepatocytes. Involvement of receptors specific for galactose, mannose 6-phosphate and mannose

Biochemical Journal ◽

10.1042/bj1800413 ◽

1979 ◽

Vol 180 (2) ◽

pp. 413-419 ◽

Cited By ~ 28

Author(s):

K Ullrich ◽

R Basner ◽

V Gieselmann ◽

K Von Figura

Keyword(s):

Cell Surface ◽

Human Urine ◽

Rat Hepatocytes ◽

Cell Surface Receptor ◽

Cell Surface Receptors ◽

Minor Role ◽

Isolated Rat Hepatocytes ◽

Surface Receptors ◽

Terminal Galactose ◽

Mannose 6 Phosphate

Adsorptive endocytosis of alpha-N-acetylglucosaminidase from human urine by isolated rat hepatocytes is inhibited by glycoproteins, polysaccharides and sugars that are known to bind to cell-surface receptors specific for either terminal galactose/N-acetylgalactosamine residues, terminal mannose residues or mannose 6-phosphate residues. Recognition of alpha-N-acetylglucosaminidase by a cell-surface receptor specific for terminal galactose/N-acetylgalactosamine residues is supported by the observations (a) that neuraminidase pretreatment of the enzyme enhances endocytosis, (b) that beta-galactosidase treatment decreases endocytosis and (c) that neuraminidase pretreatment of hepatocytes decreases alpha-N-acetylglucosaminidase endocytosis. Recognition of alpha-N-acetylglucosaminidase via receptors recognizing mannose 6-phosphate residues is lost after treatment of the enzyme with alkaline phosphatase and endoglucosaminidase H. The effect of endoglucosaminidase H supports the view that the mannose 6-phosphate residues reside in N-glycosidically linked oligosaccharide side chains of the high-mannose type. The weak inhibition of endocytosis produced by compounds known to interact with cell-surface receptors specific for mannose residues suggests that this recognition system plays only a minor role in the endocytosis of lysosomal alpha-N-acetylglucosaminidase by hepatocytes.

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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.

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Glycosylation of human proteinase-activated receptor-2 (hPAR2): role in cell surface expression and signalling

Biochemical Journal ◽

10.1042/bj20020706 ◽

2002 ◽

Vol 368 (2) ◽

pp. 495-505 ◽

Cited By ~ 82

Author(s):

Steven J. COMPTON ◽

Sabrina SANDHU ◽

Suranga J. WIJESURIYA ◽

Morley D. HOLLENBERG

Keyword(s):

Cell Surface ◽

Western Blot ◽

Molecular Mass ◽

Cell Line ◽

Blot Analysis ◽

Western Blot Analysis ◽

Mutant Cell ◽

Surface Expression ◽

Receptor Expression ◽

Cell Surface Expression

We have analysed the role of N-linked glycosylation in regulating human proteinase-activated receptor-2 (hPAR2) expression and function. Epitope-tagged wild-type hPAR2 (wt-hPAR2) or hPAR2 that lacked glycosylation sequons (following site-directed mutagenesis) in either the N-terminus [hPAR2N30A (Asn30→Ala)], extracellular loop 2 [ECL2; hPAR2N222Q (Asn222→Gln) or hPAR2N222A (Asn222→Ala)] or both (hPAR2N30A,N222A or hPAR2N30A,N222Q) were expressed in the Chinese-hamster ovary (CHO) fibroblast cell line, Pro5. Western blot analysis of wt-hPAR2 showed mature wt-hPAR2 to have a molecular mass of 55—100kDa, and 33—48kDa following N-glycosidase F deglycosylation. FACS analysis and immunocytochemistry of the wt-hPAR2 and PAR2 mutant cell lines revealed that removal of both glycosylation sequons decreases (50% of wt-hPAR2) cell surface expression. Western blot analysis indicated that both N-linked sites are glycosylated. In functional studies, hPAR2N30A displayed a selective and significant increase in sensitivity towards tryptase. Interestingly, hPAR2N222A displayed a loss in sensitivity towards all PAR2 agonists tested. However, further analysis revealed receptor sensitivity to alanine mutations in this domain, as the more conservative substitution hPAR2N222Q displayed no change in response to PAR2 agonists. hPAR2N30A,N222Q displayed increased sensitivity towards tryptase, but a loss in sensitivity towards trypsin and the synthetic peptide SLIGRL-NH2, although this loss in sensitivity towards trypsin and SLIGRL-NH2 was secondary to changes in cell-surface expression. Finally, expression of sialic-acid-deficient wt-hPAR2 in the CHO Lec2 glycosylation-deficient mutant cell line, showed a 40kDa loss in molecular mass, in addition to a marked and selective increase in sensitivity towards tryptase. We conclude that hPAR2 N-linked glycosylation and sialylation regulates receptor expression and/or signalling.

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Prolonged defects of interleukin-2 production, responsiveness, and receptor expression in patients with acute lymphoblastic leukemia

Blood ◽

10.1182/blood.v73.6.1608.1608 ◽

1989 ◽

Vol 73 (6) ◽

pp. 1608-1614 ◽

Cited By ~ 14

Author(s):

MS Borzy ◽

D Ridgway

Keyword(s):

Acute Lymphoblastic Leukemia ◽

Cell Surface ◽

Lymphoblastic Leukemia ◽

Interleukin 2 ◽

Receptor Expression ◽

Cell Surface Receptors ◽

Con A ◽

Surface Receptors ◽

Control Subjects ◽

Patient Groups

Abstract The proliferative responsiveness to, production of, and the expression of cell-surface receptors for interleukin-2 (IL-2) were examined in 14 children with acute lymphoblastic leukemia (ALL) in remission and receiving maintenance chemotherapy for 6 to 35 months; in 19 children with ALL in remission and off all therapy for 2 to 138 months; and 15 control subjects. Short-term concanavalin A (Con A)-activated, purified T lymphocytes from patients on, as well as patients off, therapy had a significantly decreased proliferative responsiveness to a saturating amount of exogenous, recombinant IL-2 as compared to control subjects (P less than 0.005 and less than 0.05, respectively). Phytohemagglutinin (PHA)-stimulated IL-2 production by peripheral blood mononuclear cells (PBMC) was also substantially decreased in both patient groups with the median values of IL-2 produced being 2.2, 2.1, and 8.1 U/mL in the on therapy, off therapy, and control groups, respectively. In addition, PHA-induced expression of cell-surface receptors for IL-2 on PBMC was significantly decreased in both patient groups as compared to control subjects (P less than 0.01). Lymphocyte proliferation to mitogens (PHA, Con A, and pokeweed mitogen) was similar in all three groups studied. These results demonstrate that substantial quantitative and qualitative abnormalities of the IL-2-T lymphocyte system are present in the majority of treated patients with ALL, not only during maintenance therapy, but also for a prolonged period after the cessation of all chemotherapy. These long-lasting defects of the IL-2 system are most likely a late effect of chemotherapy and may result in increased complications in some long- term survivors of ALL.

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Fibrillar organization of fibronectin is expressed coordinately with cell surface gangliosides in a variant murine fibroblast.

The Journal of Cell Biology ◽

10.1083/jcb.102.5.1898 ◽

1986 ◽

Vol 102 (5) ◽

pp. 1898-1906 ◽

Cited By ~ 46

Author(s):

S Spiegel ◽

K M Yamada ◽

B E Hom ◽

J Moss ◽

P H Fishman

Keyword(s):

Cell Surface ◽

Cholera Toxin ◽

Cell Attachment ◽

Specific Radioactivity ◽

Cell Types ◽

Concomitant Treatment ◽

Matrix Assembly ◽

Surface Receptors ◽

High Specific Radioactivity ◽

Fibrillar Network

NCTC 2071A cells, a line of transformed murine fibroblasts, grow in serum-free medium, are deficient in gangliosides, synthesize fibronectin, but do not retain and organize it on the cell surface. When the cells are exposed to exogenous gangliosides, fibrillar strands of fibronectin become attached to the cell surface. A morphologically distinct variant of NCTC 2071A cells was observed to both retain cell surface fibronectin and organize it into a fibrillar network when the cells were stained with anti-fibronectin antibodies and a fluorescent second antibody. A revertant cell type appeared to resemble the parental NCTC 2071A cells in terms of morphology and fibronectin organization. All three cell types were subjected to mild NaIO4 oxidation and reduction with KB3H4 of very high specific radioactivity in order to label the sialic acid residues of surface gangliosides. The variant had much more surface gangliosides than the parental, particularly more complex gangliosides corresponding to GM1 and GD1a. The surface gangliosides of the revertant were intermediate between the parental and the variant. By using sialidase, which hydrolyzes GD1a to GM1, and 125I-labeled cholera toxin, which binds specifically to GM1, the identity and levels of these gangliosides were confirmed in the three cell types. When variant cells were exposed to sialidase for 2 d, there appeared to be little change in fibronectin organization. Concomitant treatment of the cells with the B subunit of cholera toxin, which bound to all the surface GM1 including that generated by the sialidase, however, eliminated the fibrillar network of fibronectin. In addition, exposure of the variant cells to a 70,000-mol-wt fragment of fibronectin, which lacks the cell attachment domain but contains a matrix assembly domain, inhibited the formation of fibers. Finally, all three cell types were assayed for their ability to attach to and spread on fibronectin-coated surfaces; no significant differences were found. Our results further establish that the ability of a cell to organize fibronectin into an extracellular matrix is dependent on certain gangliosides, but they also indicate that cell adhesion to fibronectin is independent of these gangliosides. We suggest that matrix organization and cell attachment and spreading are based on separate mechanisms and that these functions are associated with different cell surface "receptors."

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Prognostic and Therapeutic Utility of Variably Expressed Cell Surface Receptors in Osteosarcoma

Sarcoma ◽

10.1155/2021/8324348 ◽

2021 ◽

Vol 2021 ◽

pp. 1-10

Author(s):

Yoav Zvi ◽

Elif Ugur ◽

Brian Batko ◽

Jonathan Gill ◽

Michael Roth ◽

...

Keyword(s):

Overall Survival ◽

Growth Factor ◽

Cell Surface ◽

Cell Lines ◽

Growth Factor Receptor ◽

Receptor Expression ◽

Cell Surface Receptors ◽

Surface Receptors ◽

Variable Expression ◽

Her 2

Background. Six cell surface receptors, human epidermal growth factor receptor-2 (Her-2), platelet-derived growth factor receptor-β (PDGFR-β), insulin-like growth factor-1 receptor (IGF-1R), insulin receptor (IR), c-Met, and vascular endothelial growth factor receptor-3 (VEGFR-3), previously demonstrated variable expression across varying patient-derived and standard osteosarcoma (OS) cell lines. The current study sought to validate previous expression patterns and evaluate whether these receptors offer prognostic and/or therapeutic value. Methods. Patient-derived OS cell lines (n = 52) were labeled with antibodies to Her-2, PDGFR-β, IGF-1R, IR, c-Met, and VEGFR-3. Expression was characterized using flow cytometry. The difference in geometric mean fluorescent intensity (geoMFIdiff = geoMFIpositive − geoMFInegative) was calculated for each receptor across all cell lines. Receptor expression was categorized as low (Q1), intermediate (Q2, Q3), or high (Q4). The event-free survival (EFS) and overall survival for the six cell surface receptors were estimated by the Kaplan–Meier method. Differences in hazard for EFS event and overall survival event for patients in each of the three expression levels in each of the six cell surface receptors were assessed using the log-rank test. Results. All 6 receptors were variably expressed in the majority of cell lines. IR and PDGFR-β expressions were found to be significant predictors for EFS amongst patients with nonmetastatic disease ( p = 0.02 and 0.01, respectively). The hazard ratio for EFS was significantly higher between high IR and intermediate IR expression (HR = 2.66, p = 0.02 ), as well as between high PDGFR-β and intermediate PDGFR-β expression (HR = 5.68, p = 0.002 ). Her-2, c-Met, IGF-1R, and VEGFR-3 were not found to be significant predictors for either EFS or overall survival. Conclusion. The six cell surface receptors demonstrated variable expression across the majority of patient-derived OS cell lines tested. Limited prognostic value was offered by IR and PDGFR-β expression within nonmetastatic patients. The remaining receptors do not provide clear prognostic utility. Nevertheless, their consistent, albeit variable, surface expression across a large panel of patient-derived OS cell lines maintains their potential use as future therapeutic targets.

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Diversity of receptor expression in central and peripheral mouse neurons estimated from single cell RNA sequencing

10.1101/2021.01.22.427766 ◽

2021 ◽

Author(s):

Andi Wangzhou ◽

Candler Paige ◽

Pradipta R. Ray ◽

Gregory Dussor ◽

Theodore J. Price

Keyword(s):

T Cells ◽

Cell Surface ◽

Single Cell ◽

Rna Sequencing ◽

Sympathetic Neurons ◽

Receptor Expression ◽

Cell Surface Receptors ◽

Surface Receptors ◽

Single Cell Rna Sequencing ◽

Cns Neurons

AbstractBecause somatosensory PNS neurons, in particular nociceptors, are specially tuned to be able to detect a wide variety of both exogenous and endogenous signals, it is widely assumed that these neurons express a greater variety of receptor genes. Because cells detect such signals via cell surface receptors, we sought to formally test the hypothesis that PNS neurons might express a broader array of cell surface receptors than CNS neurons using existing single cell RNA sequencing resources from mouse. We focused our analysis on ion channels, G-protein coupled receptors (GPCRS), receptor tyrosine kinase and cytokine family receptors. In partial support of our hypothesis, we found that mouse PNS somatosensory, sympathetic and enteric neurons and CNS neurons have similar receptor expression diversity in families of receptors examined, with the exception of GPCRs and cytokine receptors which showed greater diversity in the PNS. Surprisingly, these differences were mostly driven by enteric and sympathetic neurons, not by somatosensory neurons or nociceptors. Secondary analysis revealed many receptors that are very specifically expressed in subsets of PNS neurons, including some that are unique among neurons for nociceptors. Finally, we sought to examine specific ligand-receptor interactions between T cells and PNS and CNS neurons. Again, we noted that most interactions between these cells are shared by CNS and PNS neurons despite the fact that T cells only enter the CNS under rare circumstances. Our findings demonstrate that both PNS and CNS neurons express an astonishing array of cell surface receptors and suggest that most neurons are tuned to receive signals from other cells types, in particular immune cells.

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