Abstract
Aberrant folding of proteins in the endoplasmic reticulum activates the bifunctional transmembrane kinase/endoribonuclease Ire1. Ire1 excises an intron from HAC1 messenger RNA in yeasts and Xbp1 messenger RNA in metozoans encoding homologous transcription factors. This non-conventional mRNA splicing event initiates the unfolded protein response, a transcriptional program that relieves the endoplasmic reticulum stress. Here we show that oligomerization is central to Ire1 function and is an intrinsic attribute of its cytosolic domains. We obtained the 3.2-Å crystal structure of the oligomer of the Ire1 cytosolic domains in complex with a kinase inhibitor that acts as a potent activator of the Ire1 RNase. The structure reveals a rod-shaped assembly that has no known precedence among kinases. This assembly positions the kinase domain for trans-autophosphorylation, orders the RNase domain, and creates an interaction surface for binding of the mRNA substrate. Activation of Ire1 through oligomerization expands the mechanistic repertoire of kinase-based signalling receptors.
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Acknowledgements
We thank A. Krutchinsky for the help with MALDI instruments and for the tryptic digest analysis of Ire1KR32, F. Gruswitz for useful discussions, C. Waddling for managing the protein crystallization facility of the molecular structure group (MSG) at UCSF, and to the staff of the beamline 8.3.1 at the Advanced Light Source (Berkeley). We thank members of the Walter laboratory for critical review of the manuscript. A.V.K. is a recipient of Jane Coffin Childs fellowship. C.Z. was supported by a grant from the National Parkinson Foundation. R.M.S., J.F.-M. and P.F.E. were supported by an NIH grant RO1 GM60641. P.W. and K.M.S. are Investigators of the Howard Hughes Medical Institute.
Author Contributions A.V.K. designed and prepared protein and RNA constructs and carried out kinetic and biophysical analyses. A.V.K. and P.F.E. carried out crystallization and data collection. A.A.K. performed structure determination. J.F.-M. and A.V.K. contributed to crystallographic data processing and model building. C.Z. and K.M.S. selected and provided the kinase inhibitors. P.W. and R.M.S. supervised the work. A.V.K. and P.W. wrote the manuscript.
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Korennykh, A., Egea, P., Korostelev, A. et al. The unfolded protein response signals through high-order assembly of Ire1. Nature 457, 687–693 (2009). https://doi.org/10.1038/nature07661
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DOI: https://doi.org/10.1038/nature07661
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