1. Photocrosslinking-induced CRAC channel-like Orai1 activation independent of STIM1
- Author
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Maltan Lena, Weiß Sarah, Najjar Hadil, Melanie, Leopold, Lindinger Sonja, Höglinger Carmen, Höbarth Lorenz, Sallinger Matthias, Grabmayr Herwig, Berlansky Sascha, Krivic Denis, Hopl Valentina, Blaimschein Anna, Fahrner Marc, Frischauf Irene, Tiffner Adéla, and Derler Isabella
- Abstract
Ca2+ release-activated Ca2+ (CRAC) channels, indispensable for the immune system and various other human body functions, consist of two transmembrane (TM) proteins, the Ca2+-sensor STIM1 in the ER membrane and the Ca2+ ion channel Orai1 in the plasma membrane. We employ genetic code expansion in mammalian cell lines to incorporate the photocrosslinking unnatural amino acids (UAA), p-benzoyl-L-phenylalanine (Bpa) and p-azido-L-phenylalanine (Azi), into the Orai1 TM domains at different sites. Characterization of the respective UAA-containing Orai1 mutants using Ca2+ imaging and electrophysiology reveals that exposure to UV light triggers a range of effects depending on the UAA and its site of incorporation. In particular, photoactivation at A137 using Bpa in Orai1 activates Ca2+ currents that best match the biophysical properties of CRAC channels and are capable of triggering downstream signaling pathways such as nuclear factor of activated T-cells (NFAT) translocation into the nucleus without the need for the physiological activator STIM1.
- Published
- 2023
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