Too Close for Molecular Comfort

Abstract

Release of Ca 2+ from intracellular stores leads to the rapid translocation of the endoplasmic reticulum (ER)-resident protein STIM1 (stromal interacting molecule 1) to a region of the ER near the plasma membrane. This enables the interaction of STIM1 with the Orai1 channel and the subsequent entry of Ca 2+ across the plasma membrane. Várnai et al . investigated this process in living COS-7 cells in which they created molecular bridges between the plasma membrane and the adjacent ER. They targeted fluorescently labeled constructs containing the FRB fragment of mTOR (mammalian target of rapamycin) to the cytoplasmic face of the ER, and constructs containing the FKBP-12 protein to the inner face of the plasma membrane, and induced their heterodimerization with rapamycin. This allowed visualization of stable contact zones between the ER and the plasma membrane in conjunction with visualization of fluorescently labeled STIM1 and Orai1 (or both) before and after treatment with thapsigargin (to deplete Ca 2+ stores in the ER) or stimulation of P2Y receptors with ATP. These regions of juxtaposition coincided with the regions in which STIM1 and Orai1 interacted after store depletion; however, Orai1, and the STIM1-Orai1 complex, were excluded from cross-linked contact regions in which the gap between the ER and the plasma membrane was too narrow. The authors used linkers to create bridges of different length, which indicated that STIM1 could be accommodated when the gap between the two membranes was 4 to 6 nm, whereas Orai1 required a gap of 11 to 14 nm, suggesting that it exists as part of a large macromolecular complex. P. Várnai, B. Tóth, D. J. Tóth, L. Hunyady, T. Balla, Visualization and manipulation of plasma membrane-endoplasmic reticulum contact sites indicates the presence of additional molecular components within the STIM1-Orai1 complex. J. Biol. Chem. 282 , 29678-29690 (2007). [Abstract] [Full Text]