Using magnetic probes to study receptor clustering in live cells

Burcu Celikkol, Alessandra Cambi, Carl Figdor, Vinod Subramaniam, Johannes S. Kanger

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Abstract

During pathogen recognition T-Cell Receptors form microclusters which are believed to be the central signalling units. These structures could hold the secret behind the exceptional sensitivity of T-Cells in distinguishing single triggering ‘agonist’ peptides against a background of thousands. We have developed a biophysical approach based on magnetic tweezers that allows us to study the players involved in these receptor clusters and their dynamics. We use antibody functionalized magnetic beads to target CD3, a subunit of the TCR Complex to induce TCR clustering. Using magnetic tweezers, we move the beads along the cell membrane and simultaneously measure trafficking of co-receptors and proteins involved in the complex using confocal fluorescence microscopy and fluorescence recovery after photobleaching (FRAP). We study co-receptor CD6, which is considered a co-stimulator for cell activation during cluster formation. Our findings suggest that while CD6 is not physically associated with TCR complex, it gets recruited into the TCR clusters. There it is partially immobilized and moves along as clusters are displaced. The diffusion coefficient of CD6 is higher in bead-stimulated cells, whereas CD6 outside clusters diffuse faster than those within clusters. We are also downscaling this method to induce formation of receptor nanoclusters, in order to explore the effects of physical receptor oligomerization on the activity of TCR and Epidermal Growth Factor Receptors.
Original languageEnglish
Pages (from-to)20a-20a
JournalBiophysical journal
Volume106
Issue number2, Suppl. 1
DOIs
Publication statusPublished - 16 Feb 2014
Event58th Annual Biophysical Society Meeting 2014 - San Francisco, United States
Duration: 15 Feb 201419 Feb 2014
Conference number: 58

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