Abstract
Most membrane and secreted proteins are glycosylated on certain asparagine (N) residues in the endoplasmic reticulum (ER), which is crucial for their correct folding and function. Protein folding is a fundamentally inefficient and error-prone process that can be easily interfered by genetic mutations, stochastic cellular events, and environmental stresses. Because misfolded proteins not only lead to functional deficiency but also produce gain-of-function cellular toxicity, eukaryotic organisms have evolved highly conserved ER-mediated protein quality control (ERQC) mechanisms to monitor protein folding, retain and repair incompletely folded or misfolded proteins, or remove terminally misfolded proteins via a unique ER-associated degradation (ERAD) mechanism. A crucial event that terminates futile refolding attempts of a misfolded glycoprotein and diverts it into the ERAD pathway is executed by removal of certain terminal α1,2-mannose (Man) residues of their N-glycans. Earlier studies were centered around an ER-type α1,2-mannosidase that specifically cleaves the terminal α1,2Man residue from the B-branch of the three-branched N-linked Man9GlcNAc2 (GlcNAc for N-acetylglucosamine) glycan, but recent investigations revealed that the signal that marks a terminally misfolded glycoprotein for ERAD is an N-glycan with an exposed α1,6Man residue generated by members of a unique folding-sensitive α1,2-mannosidase family known as ER-degradation enhancing α-mannosidase-like proteins (EDEMs). This review provides a historical recount of major discoveries that led to our current understanding on the role of demannosylating N-glycans in sentencing irreparable misfolded glycoproteins into ERAD. It also discusses conserved and distinct features of the demannosylation processes of the ERAD systems of yeast, mammals, and plants.
Highlights
Secretory and transmembrane proteins of eukaryotic organisms are synthesized on cytosolic ribosomes and enter the endoplasmic reticulum (ER) for their folding and maturation (Rapoport, 2007)
Despite rapid progress in structural understanding of the ER-associated degradation (ERAD) machinery itself (Schoebel et al, 2017; Eldeeb et al, 2020; Wu et al, 2020), our knowledge of the initial events that commit an irreparable misfolded glycoprotein to ERAD remains incomplete. It was quite clear from the early days of ERAD research that demannosylation constitutes a key step of the ERAD pathway and intensive/extensive investigation in the last quarter century have identified and biochemically characterized the α1,2-mannosidases, which generate the evolutionarily conserved ERAD N-glycan signals with exposed α1,6Man residue, and the ERAD lectins that recognize and bind such a conserved N-glycan signal
The latest discoveries of the requirement of covalent binding to members of the protein disulfide isomerases (PDIs) family for the mannosidase activities of Htm1/ER-degradation enhancing α-mannosidase-like proteins (EDEMs) and the structural preference of the yeast Htm1-Pdi1 complex for compact but partially unstructured glycoproteins over globally unstructured conformers suggested a mechanistic mimicry between the Htm1/EDEM-PDI complexes and UDP glucose:glycoprotein glucosyltransferase (UGGT), which likely compete for their binding to a misfolded protein, resulting in N-glycan demannosylation for degradation and N-glycan reglucosylation for another refolding attempt, respectively
Summary
Secretory and transmembrane proteins of eukaryotic organisms are synthesized on cytosolic ribosomes and enter the endoplasmic reticulum (ER) for their folding and maturation (Rapoport, 2007). These results were consistent with earlier findings that mutations in ALG9 or ALG12 blocked the ERAD of CPY∗ in yeast because alg or alg mutation prevents addition of an α1,6Man residue during the assembly of the Dol-PP-Glc3Man9GlcNAc2 (Figure 1) and provided a satisfactory explanation for a previous intriguing finding that Yos interacted with CPY∗ carrying Man5GlcNAc2 N-glycans in the yeast alg mutant (Szathmary et al, 2005) These in vitro MRH-oligosaccharide binding assays prompted in vivo testing of the newly discovered ERAD N-glycan signal. A detailed structural analysis of a purified Htm1-Pdi complex could shed light on the biochemical mechanism by which this disulfide-bridged ERAD “folding sensor” recognizes misfolded glycoproteins, demannosylates their N-glycans, and forces their entry into the ERAD pathway
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