Nanometer-scale organization of the alpha subunits of the receptors for IL2 and IL15 in human T lymphoma cells

Bakker, B.I. de; Bodnár, Andrea; Dijk, E.M.H.P. van; Vámosi, György; Damjanovich, Sándor; Waldmann, Thomas A.; Hulst, Niek F. van; Jenei, Attila; García-Parajo, María F.

doi:10.1242/jcs.019513

Cited by 62 publications

(47 citation statements)

References 36 publications

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“…The EGFR is organized in clusters with a range of sizes and receptor densities and with a significant fraction of receptor oligomers in the larger clusters. Similar receptor aggregation has been observed for a variety of integral membrane proteins (15,23,28).…”

Section: Discussionsupporting

confidence: 76%

“…NSOM is one of an emerging group of super-resolution methods for fluorescence imaging with subdiffraction resolution (18,19) and is based on delivering light through an aperture that has subwavelength dimensions (20,21) to overcome the diffraction limit. NSOM has recently been used for nanoscale fluorescence imaging of membrane protein clusters with spatial resolution below 100 nm in fixed cells (22)(23)(24)(25)(26)(27)(28). Using NSOM, we show that EGFR is distributed in nanometer-sized clusters in the plasma membrane and that association with lipid raft and caveolae domains regulates receptor sequestration and trafficking in response to activation and/or inhibition in HeLa adenocarcinoma cells.…”

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confidence: 96%

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Nanoscale Imaging of Epidermal Growth Factor Receptor Clustering

Abulrob

Baumann

et al. 2010

Journal of Biological Chemistry

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The development of some solid tumors is associated with overexpression of the epidermal growth factor receptor (EGFR) and often correlates with poor prognosis. Near field scanning optical microscopy, a technique with subdiffraction-limited optical resolution, was used to examine the influence of two inhibitors (the chimeric 225 antibody and tyrosine phosphorylation inhibitor AG1478) on the nanoscale clustering of EGFR in HeLa cells. The EGFR is organized in small clusters, average diameter of 150 nm, on the plasma membrane for both control and EGF-treated cells. The numbers of receptors in individual clusters vary from as few as one or two proteins to greater than 100. Both inhibitors yield an increased cluster density and an increase in the fraction of clusters with smaller diameters and fewer receptors. Exposure to AG1478 also decreases the fraction of EGFR that colocalizes with both rafts and caveolae. EGF stimulation results in a significant loss of the full-length EGFR from the plasma membrane with the concomitant appearance of low molecular mass proteolytic products. By contrast, AG1478 reduces the level of EGFR degradation. Changes in receptor clustering provide one mechanism for regulating EGFR signaling and are relevant to the design of strategies for therapeutic interventions based on modulating EGFR signaling.The plasma membrane of the cell is a complex, carefully regulated, dynamic structure that is compartmentalized into cell surface domains such as lipid rafts, caveolae, and clathrincoated pits (1). Lipid rafts are postulated to be dynamic nanodomains ranging in size from 10 to 200 nm that are enriched in cholesterol, sphingolipids, and certain proteins and that play an important role in assembling signaling complexes (2-4). Caveolae are invaginated lipid raft domains (50 -150 nm) whose stability at the plasma membrane is attributable to the formation of stable oligomers of their coat protein, caveolin-1 (5). Caveolae are thought to serve as concentrators of various signal transduction machineries. Clathrin-coated pits (100 -150 nm) internalize rapidly upon formation at the plasma membrane, and their lateral cell surface mobility is enhanced by actin cytoskeleton depolymerization (6).The epidermal growth factor receptor (EGFR), 3 a 170-kDa transmembrane glycoprotein, is one of four members of the ErbB family of receptor tyrosine kinases involved in oncogene signaling. The initial step in receptor activation involves binding of the EGF peptide to the extracellular domain leading to dimerization or activation of pre-existing dimers (7,8). Following ligand binding, the EGFR is auto-phosphorylated in several tyrosine residues of the intracellular domain, creating high affinity sites for various adaptor molecules that transmit the mitogenic signal to the Ras/MAPK signal transduction pathway (9). Activation of Ras initiates a multistep phosphorylation cascade that leads to the activation of MAPKs, ERK1, and ERK2, which regulate transcription of molecules that are linked to cell proliferation, survival, a...

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Section: Discussionsupporting

confidence: 76%

mentioning

confidence: 96%

Nanoscale Imaging of Epidermal Growth Factor Receptor Clustering

Abulrob

Baumann

et al. 2010

Journal of Biological Chemistry

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“…Garcia-Parajo and co-workers developed SNOM to a level that it can be used in aqueous solution [43] and have applied it since to various cell biological systems [42,44,45]. The small aperture of approximately 80-100 nm allows for high-resolution imaging on flat membrane regions (approx.…”

Section: (B) Technologiesmentioning

confidence: 99%

A critical survey of methods to detect plasma membrane rafts

Klotzsch

Schütz

2013

Phil. Trans. R. Soc. B

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The plasma membrane is still one of the enigmatic cellular structures. Although the microscopic structure is getting clearer, not much is known about the organization at the nanometre level. Experimental difficulties have precluded unambiguous approaches, making the current picture rather fuzzy. In consequence, a variety of different membrane models has been proposed over the years, on the basis of different experimental strategies. Recent data obtained via high-resolution single-molecule microscopy shed new light on the existing hypotheses. We thus think it is a good time for reviewing the consistency of the existing models with the new data. In this paper, we summarize the available models in ten propositions, each of which is discussed critically with respect to the applied technologies and the strengths and weaknesses of the approaches. Our aim is to provide the reader with a sound basis for his own assessment. We close this chapter by exposing our picture of the membrane organization at the nanoscale.

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“…Aperture scanning near-field optical microscopy (a-SNOM) of fluorescently labeled interleukin (IL) 2 and IL 15 alpha subunits in the membrane of human T-lymphoma cells has shown the potential of chemical high-resolution imaging of biological systems. [1] Whereas confocal fluorescence images seemed to speak for a colocalization of IL2 and IL15, high-resolution SNOM images contradicted this postulate by revealing isolated monomers as well as clusters of different sizes.…”

Section: Introductionmentioning

confidence: 99%