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The unfolded protein response signals through high-order assembly of Ire1

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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Figure 1: Activation of Ire1 by self-association.
Figure 2: Linker controls the oligomerization and activation of Ire1.
Figure 3: Kinase inhibitors activate the RNase of wild-type Ire1.
Figure 4: Structure of the Ire1 oligomer.
Figure 5: Three interfaces of Ire1 contribute to the RNase activity.
Figure 6: The mechanism of Ire1 activation.

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Primary accessions

Protein Data Bank

Data deposits

Atomic coordinates and structure factors for the reported crystal structure have been deposited in the Protein Data Bank under accession number 3FBV.

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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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Correspondence to Alexei V. Korennykh.

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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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