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Vol. 19, Issue 7, 2802-2817, July 2008

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Dynamic Movement of the Calcium Sensor STIM1 and the Calcium Channel Orai1 in Activated T-Cells: Puncta and Distal Caps

Valarie A. Barr^*, Kelsie M. Bernot^*, Sonal Srikanth^,, Yousang Gwack^,, Lakshmi Balagopalan^*, Carole K. Regan^*, Daniel J. Helman^*, Connie L. Sommers^*, Masatsugu Oh-hora, Anjana Rao, and Lawrence E. Samelson^*

*Laboratory of Cellular and Molecular Biology, National Cancer Institute, Bethesda, MD 20892-4256; and Department of Pathology, Harvard Medical School and the Immune Disease Institute, Boston, MA 02115

Submitted February 13, 2008; Revised April 8, 2008; Accepted April 17, 2008
Monitoring Editor: Carl-Henrik Heldin

The proteins STIM1 and Orai1 are the long sought components of the store-operated channels required in T-cell activation. However, little is known about the interaction of these proteins in T-cells after engagement of the T-cell receptor. We found that T-cell receptor engagement caused STIM1 and Orai1 to colocalize in puncta near the site of stimulation and accumulate in a dense structure on the opposite side of the T-cell. FRET measurements showed a close interaction between STIM1 and Orai1 both in the puncta and in the dense cap-like structure. The formation of cap-like structures did not entail rearrangement of the entire endoplasmic reticulum. Cap formation depended on TCR engagement and tyrosine phosphorylation, but not on channel activity or Ca²⁺ influx. These caps were very dynamic in T-cells activated by contact with superantigen pulsed B-cells and could move from the distal pole to an existing or a newly forming immunological synapse. One function of this cap may be to provide preassembled Ca²⁺ channel components to existing and newly forming immunological synapses.

Present Address: Department of Physiology, David Geffen School of Medicine at UCLA, Los Angeles, CA 90095-1751.

Address correspondence to: Lawrence E Samelson (samelson{at}helixnih.gov)

Abbreviations used: APC, antigen-presenting cell; CRAC, Ca²⁺ release–activated Ca²⁺ channel; ER, endoplasmic reticulum; FRAP, fluorescence recovery after photobleaching; FRET, Förster resonance energy transfer; IS, immunological synapse; MEF, mouse embryonic fibroblast; PM, plasma membrane; TCR, T-cell receptor.