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Communication between N terminus and loop2 tunes Orai activation

Ca(2+) release-activated Ca(2+) (CRAC) channels constitute the major Ca(2+) entry pathway into the cell. They are fully reconstituted via intermembrane coupling of the Ca(2+)-selective Orai channel and the Ca(2+)-sensing protein STIM1. In addition to the Orai C terminus, the main coupling site for S...

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Detalles Bibliográficos
Autores principales: Fahrner, Marc, Pandey, Saurabh K., Muik, Martin, Traxler, Lukas, Butorac, Carmen, Stadlbauer, Michael, Zayats, Vasilina, Krizova, Adéla, Plenk, Peter, Frischauf, Irene, Schindl, Rainer, Gruber, Hermann J., Hinterdorfer, Peter, Ettrich, Rüdiger, Romanin, Christoph, Derler, Isabella
Formato: Online Artículo Texto
Lenguaje:English
Publicado: American Society for Biochemistry and Molecular Biology 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5787804/
https://www.ncbi.nlm.nih.gov/pubmed/29237733
http://dx.doi.org/10.1074/jbc.M117.812693
Descripción
Sumario:Ca(2+) release-activated Ca(2+) (CRAC) channels constitute the major Ca(2+) entry pathway into the cell. They are fully reconstituted via intermembrane coupling of the Ca(2+)-selective Orai channel and the Ca(2+)-sensing protein STIM1. In addition to the Orai C terminus, the main coupling site for STIM1, the Orai N terminus is indispensable for Orai channel gating. Although the extended transmembrane Orai N-terminal region (Orai1 amino acids 73–91; Orai3 amino acids 48–65) is fully conserved in the Orai1 and Orai3 isoforms, Orai3 tolerates larger N-terminal truncations than Orai1 in retaining store-operated activation. In an attempt to uncover the reason for these isoform-specific structural requirements, we analyzed a series of Orai mutants and chimeras. We discovered that it was not the N termini, but the loop2 regions connecting TM2 and TM3 of Orai1 and Orai3 that featured distinct properties, which explained the different, isoform-specific behavior of Orai N-truncation mutants. Atomic force microscopy studies and MD simulations suggested that the remaining N-terminal portion in the non-functional Orai1 N-truncation mutants formed new, inhibitory interactions with the Orai1-loop2 regions, but not with Orai3-loop2. Such a loop2 swap restored activation of the N-truncation Orai1 mutants. To mimic interactions between the N terminus and loop2 in full-length Orai1 channels, we induced close proximity of the N terminus and loop2 via cysteine cross-linking, which actually caused significant inhibition of STIM1-mediated Orai currents. In aggregate, maintenance of Orai activation required not only the conserved N-terminal region but also permissive communication of the Orai N terminus and loop2 in an isoform-specific manner.