Fic‐mediated deAMPylation is not dependent on homo‐dimerization and rescues toxic AMPylation in flies

Abstract

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 FicE247G mutant is lethal in Drosophila. This lethality is cell autonomous as directed expression of the mutant FicE247G 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 FicE247G mutant in Drosophila 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. We believe this mechanism leads to a molecular rheostat in the cell for induction of the UPR.Support or Funding InformationThis work was funded by National Institutes of Health (NIH) grant R01 GM115188, the Welch Foundation grant I‐1561, and Once Upon a Time…Foundation to K.O., NIH grants (EY010199 and EY021922) to H.K. and NSF Graduate Research Fellowship (1000176311) to A.T.M. K.O. is a Burroughs Welcome Investigator in Pathogenesis of Infectious Disease, a Beckman Young Investigator, and a W. W. Caruth, Jr., Biomedical Scholar and has an Earl A. Forsythe Chair in Biomedical Science.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.