Cysteine cross-linking in native membranes establishes the transmembrane architecture of Ire1

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We used cysteine crosslinking to characterize the configuration of the transmembrane region of the unfolded protein response (UPR) transducer Ire1 from baker's yeast during different forms of stress. We find that different forms of ER stress converge in a single configuration of Ire1. The functionality of the individual single-cysteine variants of Ire1 was characterized by microscopy, cell-based ER stress resistance assays, and qPCR to characterize the degree of HAC1 mRNA splicing and the induction of UPR target gens..

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