An oligomeric state-dependent switch in the ER enzyme FICD regulates AMPylation and deAMPylation of BiP.

Luke A Perera,David Ron,Stephen H McLaughlin,Randy J Read,Lisa Neidhardt,Claudia Rato,Yahui Yan,Steffen Preissler

doi:10.15252/embj.2019102177

Luke A Perera, David Ron + Show 6 more

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https://doi.org/10.15252/embj.2019102177

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Abstract

AMPylation is an inactivating modification that alters the activity of the major endoplasmic reticulum (ER) chaperone BiP to match the burden of unfolded proteins. A single ER‐localised Fic protein, FICD (HYPE), catalyses both AMPylation and deAMPylation of BiP. However, the basis for the switch in FICD's activity is unknown. We report on the transition of FICD from a dimeric enzyme, that deAMPylates BiP, to a monomer with potent AMPylation activity. Mutations in the dimer interface, or of residues along an inhibitory pathway linking the dimer interface to the enzyme's active site, favour BiP AMPylation in vitro and in cells. Mechanistically, monomerisation relieves a repressive effect allosterically propagated from the dimer interface to the inhibitory Glu234, thereby permitting AMPylation‐competent binding of MgATP. Moreover, a reciprocal signal, propagated from the nucleotide‐binding site, provides a mechanism for coupling the oligomeric state and enzymatic activity of FICD to the energy status of the ER.

Highlights

In all domains of life, protein folding homeostasis is achieved by balancing the burden of unfolded proteins and the complement of chaperones
Whilst the FICD gene is necessary for BiP AMPylation, overexpression of the wild-type FICD enzyme does not result in a detectable pool of BiP-AMP in cells (Preissler et al, 2015b)
Somewhere between low-level endogenous expression, which yields physiologically regulated AMPylation, and overexpression, which precludes BiP-AMP accumulation, retrovirally rescued FICDÀ/À cells were endowed with a measure of BiP AMPylation (Figs 1A and EV1A–C)

Summary

Introduction

In all domains of life, protein folding homeostasis is achieved by balancing the burden of unfolded proteins and the complement of chaperones. In addition to well-recognised transcriptional and translational strands of the UPR (Walter & Ron, 2011), recent findings have drawn attention to the existence of rapid post-translational mechanisms that adjust the activity of the ER Hsp chaperone BiP (Preissler & Ron, 2019). Best understood amongst these is AMPylation, the covalent addition of an AMP moiety from ATP onto a hydroxyl group-containing amino acid side chain. This modification is temporally dynamic and the levels of BiP-AMP respond to changes in ER protein folding load (Preissler et al, 2015b)

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