Fic-mediated deAMPylation is not dependent on homodimerization and rescues toxic AMPylation in flies.

Protein chaperones play a critical role in proteostasis. The activity of the major endoplasmic reticulum chaperone BiP (GRP78) is regulated by Fic-mediated AMPylation during resting states. By contrast, during times of stress, BiP is deAMPylated. Here, we show that excessive AMPylation by a constitutively active Fic mutant is lethal in This lethality is cell-autonomous, as directed expression of the mutant Fic to the fly eye does not kill the fly but rather results in a rough and reduced eye. Lethality and eye phenotypes are rescued by the deAMPylation activity of wild-type Fic. Consistent with Fic acting as a deAMPylation enzyme, its activity was both time- and concentration-dependent. Furthermore, Fic deAMPylation activity was sufficient to suppress the AMPylation activity mediated by the constitutively active Fic mutant in S2 lysates. Further, we show that the dual enzymatic activity of Fic is, in part, regulated by Fic dimerization, as loss of this dimerization increases AMPylation and reduces deAMPylation of BiP.