AMPylation matches BiP activity to client protein load in the endoplasmic reticulum

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Abstract

The endoplasmic reticulum (ER) localized Hsp70 chaperone BiP affects protein folding homeostasis and the response to ER stress. Reversible inactivating covalent modification of BiP is believed to contribute to the balance between chaperones and unfolded ER proteins, but the nature of this modification has so far been hinted at indirectly. We report that deletion of FICD, a gene encoding an ER-localized AMPylating enzyme, abolished detectable modification of endogenous BiP enhancing ER buffering of unfolded protein stress in mammalian cells, whilst deregulated FICD activity had the opposite effect. In vitro, FICD AMPylated BiP to completion on a single residue, Thr⁵¹⁸. AMPylation increased, in a strictly FICD-dependent manner, as the flux of proteins entering the ER was attenuated in vivo. In vitro, Thr⁵¹⁸ AMPylation enhanced peptide dissociation from BiP 6-fold and abolished stimulation of ATP hydrolysis by J-domain cofactor. These findings expose the molecular basis for covalent inactivation of BiP.

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Steffen Preissler

Cambridge Institute for Medical Research, University of Cambridge, Cambridge, United Kingdom

Competing interests
No competing interests declared.
Cláudia Rato

Cambridge Institute for Medical Research, University of Cambridge, Cambridge, United Kingdom

Competing interests
No competing interests declared.
Ruming Chen

Cambridge Institute for Medical Research, University of Cambridge, Cambridge, United Kingdom

Competing interests
No competing interests declared.
Robin Antrobus

Cambridge Institute for Medical Research, University of Cambridge, Cambridge, United Kingdom

Competing interests
No competing interests declared.
Shujing Ding

MRC Mitochondrial Biology Unit, Cambridge, United Kingdom

Competing interests
No competing interests declared.
Ian M Fearnley

MRC Mitochondrial Biology Unit, Cambridge, United Kingdom

Competing interests
No competing interests declared.
David Ron

Cambridge Institute for Medical Research, University of Cambridge, Cambridge, United Kingdom

For correspondence
dr360@medschl.cam.ac.uk

Competing interests
David Ron, Reviewing editor, eLife.

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