Abstract
The maintenance of endoplasmic reticulum (ER) homeostasis is a critical aspect of determining cell fate and requires a properly functioning unfolded protein response (UPR). We have discovered a previously unknown role of a post-translational modification termed adenylylation/AMPylation in regulating signal transduction events during UPR induction. A family of enzymes, defined by the presence of a Fic (filamentation induced by cAMP) domain, catalyzes this adenylylation reaction. The human genome encodes a single Fic protein, called HYPE (Huntingtin yeast interacting protein E), with adenylyltransferase activity but unknown physiological target(s). Here, we demonstrate that HYPE localizes to the lumen of the endoplasmic reticulum via its hydrophobic N terminus and adenylylates the ER molecular chaperone, BiP, at Ser-365 and Thr-366. BiP functions as a sentinel for protein misfolding and maintains ER homeostasis. We found that adenylylation enhances BiP's ATPase activity, which is required for refolding misfolded proteins while coping with ER stress. Accordingly, HYPE expression levels increase upon stress. Furthermore, siRNA-mediated knockdown of HYPE prevents the induction of an unfolded protein response. Thus, we identify HYPE as a new UPR regulator and provide the first functional data for Fic-mediated adenylylation in mammalian signaling.
Highlights
Adenylylation/AMPylation by Fic proteins alters cellular signaling
HYPE⌬N45-E234G Is a Highly Active Adenylyltransferase That Autoadenylylates at Thr-183, Ser-79, and Thr-80 in Vitro—HYPE is a 52-kDa protein that is expressed in all tissues, albeit at very low levels [5]
SMART (Simple Molecular Architecture Research Tool) analysis of the domain architecture of HYPE predicts an N-terminal signal peptide and/or a hydrophobic stretch of amino acids that could specify a transmembrane sequence (Fig. 1A). This is followed by a tetratricopeptide repeat (TPR) domain that is typically implicated in proteinprotein interactions and a Fic domain at its C terminus (Fig. 1A)
Summary
Adenylylation/AMPylation by Fic proteins alters cellular signaling. HYPE, the sole human Fic protein, is an adenylyltransferase. Significance: Adenylylation/AMPylation is a new mode of UPR regulation This is the first demonstration of a physiological role for human HYPE. The human genome encodes a single Fic protein, called HYPE (Huntingtin yeast interacting protein E), with adenylyltransferase activity but unknown physiological target(s). We identify HYPE as a new UPR regulator and provide the first functional data for Fic-mediated adenylylation in mammalian signaling. HYPE belongs to the Fic (filamentation induced by cAMP) family of enzymes that uses cellular sources of nucleotides to carry out various post-translational modifications [3,4,5] Predominant among these post-translational modifications is the covalent addition of AMP to target substrates such as mammalian Rho GTPases [4,5,6,7,8]. Our study is the first functional demonstration of the physiological significance of Fic-mediated adenylylation in mammalian cell signaling
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