Scopus İndeksli Yayınlar Koleksiyonu / Scopus Indexed Publications Collection

Permanent URI for this collectionhttps://hdl.handle.net/11147/7148

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Author: search.filters.author.Colson, Steven D.

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  • Article
    Citation - WoS: 100
    Cholesterol Dictates the Freedom of Egf Receptors and Her2 in the Plane of the Membrane
    (Elsevier Ltd., 2013) Orr, Galya; Hu, Dehong; Özçelik, Serdar; Wiley, H. Steven; Colson, Steven D.; Opresko, Lee K.
    The flow of information through the epidermal growth factor receptor (EGFR) is shaped by molecular interactions in the plasma membrane. The EGFR is associated with lipid rafts, but their role in modulating receptor mobility and subsequent interactions is unclear. To investigate the role of nanoscale rafts in EGFR dynamics, we used single-molecule fluorescence imaging to track individual receptors and their dimerization partner, human epidermal growth factor receptor 2 (HER2), in the membrane of human mammary epithelial cells. We found that the motion of both receptors was interrupted by dwellings within nanodomains. EGFR was significantly less mobile than HER2. This difference was likely due to F-actin because its depolymerization led to similar diffusion patterns between the EGFR and HER2. Manipulations of membrane cholesterol content dramatically altered the diffusion pattern of both receptors. Cholesterol depletion led to almost complete confinement of the receptors, whereas cholesterol enrichment extended the boundaries of the restricted areas. Interestingly, F-actin depolymerization partially restored receptor mobility in cholesterol-depleted membranes. Our observations suggest that membrane cholesterol provides a dynamic environment that facilitates the free motion of EGFR and HER2, possibly by modulating the dynamic state of F-actin. The association of the receptors with lipid rafts could therefore promote their rapid interactions only upon ligand stimulation.
  • Article
    Citation - WoS: 3
    Fret Measurements Between Small Numbers of Molecules Identifies Subtle Changes in Receptor Interactions
    (SPIE, 2013) Özçelik, Serdar; Orr, Galya; Hu, Dehong; Chen, Chii-Shiarng; Reşat, Haluk; Harms, Greg S.; Opresko, Lee K.; John Wiley and Sons Inc., H. Steven; Colson, Steven D.
    Overexpression of HER2 alters the cellular behavior of EGF receptor (EGFR) and itself,with great implications on cell fate. To understand the molecular interactions underlying these alterations, we quantified the association between the two receptors by looking at efficiency changes in fluorescence resonance energy transfer (FRET) between a small number of molecules at the membrane of living cells. Human mammary epithelial (HME) cells expressing varying degrees of HER2 were studied, to identify and compare the degree of receptors interactions as a function of HER2 overexpression. A high resolution wide-field laser microscope combined with a high sensitivity cooled CCD camera was used to capture simultaneously donor and acceptor emissions. Alternating between green and red lasers every 80 msec, donor, FRET, and acceptor images were acquired and were used to calculate FRET efficiency. Automated image analysis was developed to create FRET efficiency maps from overlapping donor, acceptor and FRET images, and derive FRET efficiency histograms to quantify receptorreceptor interactions pixel by pixel. This approach enabled us to detect subtle changes in the average distance between EGFR molecules, and between EGFR and HER2. We found pre-existing EGFR homoassociations, and EGFR-HER2 heteroassociations in cells overexpressing HER2, and identified the changes in these interactions with ligand stimulation. These observations demonstrate the power of FRET measurements between small numbers of molecules in identifying subtle changes in molecular interactions in living cell.